All posts in Research and HD

The Stanford “Center of Excellence”

Stanford has recently been recognized by the Huntington’s Disease Society of America (HDSA) as a “Center of Excellence,” a designation reserved for medical clinics providing the highest standard of care to Huntington’s disease (HD) patients. Two neurologists, Drs. Veronica Santini and Sharon Sha, direct the Stanford Multidisciplinary HD and Ataxia clinic, which has received the designation. Dr. Santini specializes in Movement Disorders, while Dr. Sha specializes in Memory Disorders, and together they lead a team of diverse experts to treat the wide variety of symptoms associated with HD.

The HDSA “Center of Excellence” (COE) is a competitive designation requiring a lengthy application process. The title is reserved for clinics that meet the criteria for exceptional quality of care as recommended by the HDSA. In some circumstances, it also allows for additional funding of HD-related services. Although the clinic has provided extraordinary care for patients with HD for many years, they were only newly designated as a COE in January of 2015. For Dr. Santini, being a COE means providing the “gold” standard of care to her patients with Huntington’s disease. This means providing the total emotional, cognitive, psychosocial, and physical supports that each patient needs. Stanford is able to provide this holistic care through a multidisciplinary team approach with a large and dedicated team (listed below), made up of a genetic counselor, a clinical social worker, a neuropsychiatrist, and a nurse coordinator. The team also has speech, occupational, and physical therapists on call and with plans to move into the new neuroscience building on Stanford’s campus, these services will be available at each clinic visit. The clinic would not be able to offer these wonderful services to patients without the generosity of a local private donor. The new designation of Stanford as a COE reinforces the clinic as a beacon of treatment and information for patients and families throughout the Northern California area. The COE designation also makes the Stanford clinic a place for education of clinical trial participation for new therapeutics and research. Dr. Santini and Dr. Sha wish to begin offering clinical studies in shortly.

The large team at Stanford holds their HD and Ataxia clinic every Friday. The clinic is now located on the third floor of the Stanford hospital and clinics, but will be moving to the new neurosciences expansion that is currently under construction and is due to open in November of 2015. The HD and Ataxia clinic will be on the first floor providing better accessibility for HD patients and other patients with limited mobility and movement disorders. The open floor plan of the new clinic will better allow patients to see multiple specialists on the HD team per visit, further reinforcing the holistic care model, for which the clinic was already known.

A conversation with Dr. Santini revealed that the collaborative nature of team is what is most striking about the Stanford COE. Dr. Santini says that each member shows incredible initiative in their roles and everyone has their heart in it to take the best possible care of each patient.

For more information on the new Stanford “Center of Excellence”, visit their website by clicking here:


List of Team Members:

Veronica Santini, MD, MA, Clinical Assistant Professor of Neurology

Sharon Sha, MD, MS, Clinical Assistant Professor of Neurology

Vala Paladottir, MD, Movement Disorders Fellow

Victoria Tanoury, RN, CNRN, Clinical Nurse Coordinator

Carly Siskind, MS, LCGC, Genetic Counselor

Andrea Kwan, MS LGC, Genetic Counselor

Amee Jaiswal, MSW, Clinical Social Worker

Sepidedeh Bajestan, MD, PhD, Neuropsychiatrist

John Barry, MD, Neuropsychiatrist


Research and HD: Table of Contents

Research Basics

Research Updates

 HD Scientists

Research Institutions


Research Updates: Table of Contents

Research Updates


Inhibition of mitochondrial protein import by mutant huntingtin

Inhibition of mitochondrial protein import by mutant huntingtin

Research has shown that mitochondrial dysfunction is associated with neuronal loss in Huntington’s disease (HD). However, it is unclear how mutant huntingtin (Htt) may cause such dysfunction. Researchers at University of Pittsburg and Washington University have discovered evidence of a direct relationship between mutant Htt and the mitochondrial protein import machinery1. This study has many implications for HD research including the development of mitochondrial protein import-based therapies.


Mitochondria are specialized subunits within a cell often referred to as the ‘energy powerhouse’ as they are responsible for conversion of nutrients into energy for the cell. Mitochondria can be found in every cell of the human body (with the exception of red blood cells) and they are also responsible for calcium homeostasis and the regulation of apoptosis (programmed cell death) 2.

Unfortunately, mitochondria may not function properly due to the obstruction of important pathways, which can compromise the energy production of the cell. When mitochondria in the brain malfunction, less energy is generated within the neuronal cells, which can lead to cell injury, and eventually cell death. Therefore, neuronal cells need healthy mitochondria to survive. There is evidence that mitochondrial dysfunction may be a critical driver of HD pathophysiology1.

The mitochondrial protein import pathway is crucial for healthy mitochondrial function since proteins are often created in the cytoplasm before traveling through several complexes to make their way to the mitochondria. To stress the importance of this pathway, mitochondria contain approximately 1,500 different proteins, yet 99% of which are encoded by the nuclear genome.

The Study^

Initially, the scientists observed that mutant Htt protein, unlike normal Htt protein, was present within brain mitochondria of HD patients. They hypothesized the existence of an interaction between mHtt and some mitochondrial proteins. The researchers conducted a series of experiments using mice models and cell lines to determine what that interaction is exactly and if that interaction negatively affects the mitochondria function.

In order to further understand how this deficiency in mitochondrial protein import is occurring, the scientists further investigated the role of Htt protein using an in vitro protein import assay with a radiolabeled precursor matrix protein (pOTC) which allows the measurement of the import activity for many mitochondrial proteins. They performed this assay in normal mitochondria in the presence of either normal (23Q) or mutant (97Q) recombinant Htt fusion proteins (shorter N-terminal Htt fragments that have been tagged to allow for purification and increased expression3) and observed that, if the Htt fragment carries a polyQ expansion, it will inhibit the uptake of preproteins into the mitochondria through direct association with the TIM23 mitochondrial protein import complex1. (The TIM23 complex assists in the movements of certain proteins across the inner mitochondrial membrane into the mitochondrial matrix.) The mutant Htt does, in fact, cause mitochondrial dysfunction by interfering with the mitochondria’s protein import function.

To understand how to prevent such dysfunction, the scientists overexpressed the TIM23 complex of the mitochondria. This resulted in more pre-proteins being able to travel through the membrane and into the mitochondria, and consequently, successfully limited the problematic cell death caused by the mutant Htt. This observation confirms that the purposeful overexpression of the TIM23 complex may actually be a key therapeutic target as cell death could be avoided despite a presence of mutant Htt.

It is important to note that there is a brain-specific reduction in the activity of protein import in neuronal mitochondria, rather than an increased sensitivity to import dysfunction, as previously hypothesized. Because of the energetic demands of synaptic transmission, synaptosomal mitochondria might be more sensitive to changes in protein import.

Finally, since HD is an age-dependent progressive neurodegenerative disease, the researchers decide to investigate how age-related insults like oxidative stress (link to definition) might compound the progression of the disease. For this experiment, mitochondrial protein import activity was observed in the presence of a sub lethal dose of hydrogen peroxide. This dosage did not impact the wild type neurons. However, it did significantly decrease import activity in the 195CAG HD neurons.


These results suggest that mutant Htt inhibits mitochondrial protein import via a direct interaction with the import machinery and have important implications in respect to the development of future HD therapies. Scientists can now apply these findings to target mechanisms that prevent the mutant Htt from blocking mitochondrial protein import in an attempt to prevent or delay neuronal cell death due to mitochondrial dysfunction. Such therapeutic approach could have the potential to slow down the progression of the disease and reduce the need to focus on the more difficult task of neurogenesis and network repair.

For Further Reading^

1.Yano, Hiroko, et al. “Inhibition of mitochondrial protein import by mutant huntingtin.” Nature neuroscience (2014).

2.Wiedemann, Nils, Ann E. Frazier, and Nikolaus Pfanner. “The protein import machinery of mitochondria.” Journal of Biological Chemistry 279.15 (2004): 14473-14476.

3. “GST-tagged Proteins – Production and Purification.” GST-tagged Proteins. N.p., n.d. Web. 02 July 2014.

4. . Johri, A. & Beal, M.F. Antioxidants in Huntington’s disease. Biochem. Biophys. Acta 1822, 664–674 (2012).

K. Powers 2014


Emotional Recognition Deficits

It is well documented, as reviewed by Labuschagne et al. (2012), that Huntington’s disease (HD) patients often have difficulty recognizing facial cues and understanding emotions of other individuals1, especially regarding the emotion of disgust2. Such emotional deficits can greatly diminish their ability to communicate, which may result in aggravated tension and stress between patients and caregivers.

In order to understand the potential spectrum of emotional recognition deficits in HD and its relationship to certain medications, a recent study conducted as part of a multi-site international project (TRACK-HD) examined emotion recognition in relation to the use of neuroleptic and selective serotonin reuptake inhibitor (SSRI) medications1.

What are neuroleptic and SSRI medications?^

Neuroleptic drugs are anti-psychotic medications that affect one’s cognition and behavior. These drugs have the potential to cause apathy, reduced range of emotion, confusion and agitation3. Physicians prescribe HD patients neuroleptics in order to treat chorea and behavioral disturbances such as irritability and anger outbursts, and less often for psychotic symptoms1.

SSRIs are anti-depressant medications, which, on top of being used for depression treatment, may also be used to subdue behavioral issues such as irritability and aggression3.

In clinical trials focusing on diseases other than HD, these medications have been found to reduce emotional capabilities and cause symptoms such as affective indifference, emotional blunting, reduced facial expressiveness, and reduced intensity and frequency of emotional experiences1. The aim of this study was to determine if these observations are applicable to the HD patients.


The researchers identified 344 participants that ranged from premanifest patients (individuals which have already been diagnosed as carriers of the HD mutation, but have yet to start displaying any signs of HD-related symptoms ; n=115), early HD patients (n=113), and controls (n=116). The participants were between 18 and 65 years of age with no history of other neurological illnesses. Using six emotional expressions and a neutral expression, the researchers examined how these three different groups would identify and respond to the emotions demonstrated. Additionally, the researchers focused on the subset of early HD patients who were taking neuroleptic or SSRI medications in order to compare the emotion recognition performance with that of early HD patients who were not taking these medications.

In order to gage emotional recognition, the researchers utilized a computer tablet to show facial stimuli from the Ekman and Friesen face stimulus set5. Each experimental trial included a single face showing one emotional expression (anger, disgust, fear, happiness, sadness, or surprise) or a neutral expression. Subsequently, participants were advised to press one of the seven emotional word response labels displayed below the facial stimuli, allowing them ample time to signify the emotion of the face. Each participant performed 70 experimental trials.


Overall, symptomatic participants were significantly impaired when it came to recognizing the following individual emotions: anger, fear, and surprise. This impairment is greater than that observed in the unaffected controls and the premanifest groups. Those in the early stage of HD who were taking the neuroleptics were significantly less accurate at recognizing the emotions of fear, happiness and sadness than the unmedicated group. However, members of the early HD group who were taking SSRI medication had results that correlated with improved facial recognition, particularly for disgust and sadness emotions.


This study showed that neuroleptic use by HD patients was associated with worse facial emotion recognition, whereas SSRI use was associated with better emotion recognition. These findings are of high significance for HD patient care since medications commonly prescribed to patients may difficult communication and affect social interactions. HD patients are more susceptible to misinterpret neutral events that are actually irrelevant as something that is significant, leading to issues such as depression. Prescribing medication-targeting symptoms such as depression seems to further alter the ability of those affected to communicate or read emotional cues from those around them. Therefore, a better understanding of the effects of neuroleptics and SSRIs is of great importance.

Further reading^

1. Labuschagne, Izelle, et al. “Emotional face recognition deficits and medication effects in pre-manifest through stage-II Huntington’s disease.” Psychiatry research 207.1 (2013): 118-126.

This scientific article is the primary article describing the experiment regarding emotional face recognition issues among different stages of Huntington’s disease.

2. Kipps, C. M., et al. “Disgust and happiness recognition correlate with anteroventral insula and amygdala volume respectively in preclinical Huntington’s disease.” Journal of cognitive neuroscience 19.7 (2007): 1206-1217.

This study by Kipps et al. focuses on some of the first research regarding emotional recognition in Huntington’s disease patients.

3. “Depression (major Depressive Disorder).” Selective Serotonin Reuptake Inhibitors (SSRIs). Mayo Clinic, n.d. Web. 02 July 2014.

The Mayo Clinic describes the use of SSRIs in the treatment of depression.

4. Cubeddu, Richard Finkel, Michelle A. Clark, Luigi X. (2009). Pharmacology (4th ed.). Philadelphia: Lippincott Williams & Wilkins. p. 151.ISBN 9780781771559

This resource profiles the function and use of psychiatric drugs such as neuroleptics and SSRI.

5. Ekman, Paul. “Facial expressions.” Handbook of cognition and emotion 16 (1999): 301-320.

This handbook covers various tools, techniques and practices for studying cognition and emotion.

K. Powers 2014


Trojan Therapy

The blood-brain barrier (BBB) is a layer of cells that block most molecules from entering the brain in order to protect this sensitive organ from external invaders. It does its job so well that delivering treatments to the brain is extremely difficult and poses a great challenge to drug development.




Researchers from the University of Nebraska Medical Center, the University of North Carolina, and Lomonosov Moscow State University in Russia have found a potential solution. In early 2013, these collaborators announced that they were successful in delivering an enzyme to the site of cell death within the brain of a mouse experiencing poor motor function characteristic of Parkinson’s disease.  The researchers accomplished targeted delivery through a method called Trojan therapy. This article will explain the concept of Trojan therapy, as well as its potential application for Huntington’s disease.

What is Trojan Therapy?^

The method for drug delivery is named for the Trojan War tale, during which one army snuck past the defenses of their enemy by disguising themselves within large wooden horses. The horse was presented as a gift and when it was brought behind the city’s defenses the army used the element of surprise to defeat the enemy forces.

Trojan therapy works in a similar fashion. The blood-brain barrier serves as a fortress wall for the brain. In this case, researchers disguised potential treatments within macrophages, immune cells that are allowed entry past the blood-brain barrier, making it possible to deliver the targeted therapy straight to the cells that need it.

The scientists utilized an antioxidant enzyme (proteins that combat oxidative free radicals within the body) known as catalase, to prevent cell death within the mouse brain affected by Parkinson’s disease. They engineered macrophages to carry the genetic material for catalase past the blood-brain barrier and into the brain. Once inside the brain, the macrophages attached to dying neurons, stimulating signals that called for backup to repair these neurons.

The Science behind Trojan Therapy^

Oxidative stress can lead to cell damage by producing free radicals, highly reactive molecules that can indiscriminately react with and damage cellular components. Reducing excess free radicals with antioxidants is an attractive possible method to reducing inflammation caused by cell damage in Parkinson’s brains. In the past, introducing antioxidant therapies has proven unsuccessful because the treatments were not able to overcome the blood-brain barrier.

In order for this type of therapy to function properly, blood-borne macrophages must be able to carry antioxidant proteins across the blood-brain barrier to affected areas of the brain. In this study, the scientists loaded the macrophages with nanozymes, a type of enzyme that was used to clear out free radicals from the brain.


Once the macrophage finds its way into the brain, there are three options for the nanozyme to reach its specific target in the brain:

1) Transient fusion of cellular membranes

2) Formation of macrophage bridges (physical structures that repair cellular gaps) and cellular movement mechanisms and,

3) Release of exosomes – large vesicles or storage pockets in the cell – containing the nanozyme. 1

Exosomes appear to be most promising for targeting cell death as research has shown they are excellent long-distance cellular transporters. They effectively transport many biological molecules involved in making proteins from the information contained in genetic material, including proteins, mRNA, and microRNA (Zomer et al, 2010). Researchers engineered macrophages to release exosomes containing DNA, mRNA, transcription factors (proteins that attach to DNA and control transcription of genetic information to messenger RNA) and an encoded catalase protein to assist in reducing inflammation of the brain. In Parkinson’s disease mouse models, the researchers discovered that sustained production of catalase resulted in inflammatory and neuroprotective outcomes, meaning cell death and brain atrophy was not as likely in those mouse models treated by the macrophage Trojan therapy.

The expression of the encoded catalase included within the macrophage increased over time, while DNA levels remained constant, which has implications for the therapeutic efficacy. An antioxidant with greater positive outcomes might be able to be transferred to needy neurons through this mechanism of delivery.

Conclusion of Findings^

The control group for this experiment utilized mouse models without inflammation.  The experimental group included mice exhibiting motor signs of PD. After injecting the PD mice with the macrophages, noticeable changes occurred. The fluorescent markers used to tag the progress of the materials packaged within the macrophage showed thorough coverage of the brain, especially to regions of inflammation where cell repair support was needed. These mice were able to perform just as well as the control mice on balance tests after receiving the Trojan therapy.

Oxidative stress can lead to cell damage. Reducing excess free radicals is an attractive method of reducing inflammation in PD brains. In the past, these methods have proven unsuccessful due a lack of mechanisms to deliver therapeutic drugs across the blood-brain barrier. The research conducted by this collaboration of scientists is the first to find a potential route for overcoming these obstacles. In fact, this method might be more effective than traditional viral vector gene therapy used for these purposes.

The research by the University of North Carolina and Lomonosov Moscow State University highlights various issues faced by clinical researchers. As outlined in this article, ability to cross the blood-brain barrier plays a major role in therapy design. Additionally, as the brain is a very sensitive organ, it is difficult to gauge the toxicity thresholds for various treatments and medications. Exosomes, unfortunately, are not a silver bullet either. The efficiency of loading drugs into exosomes is a challenge, as well as the difficulty in targeting the exosome to a particular cell type or organ.

Scientist Andrew Feigin of Hofstra North Shore-LIJ School of Medicine is currently leading a human-based clinical trial in which he is trying to use virus-delivered gene therapy for PD. This method has more sustained impacts than macrophages, but is much harder to get through the brain as the immune system fights these foreign viruses trying to pass the blood-brain barrier.

Researcher Elena Batrakova of University of North Carolina aims to deliver growth factors into the brain utilizing macrophages, in order to stimulate the growth of neurons. She believes that macrophages are the solution to reversing neuronal death in patients with PD. To-date this research has only been conducted in mouse models. In order to start human trials, scientists need to prove the safety of this therapy, as well as assure this therapy can be repeated successfully. This process could take several more years before human trials begin.

What does this research means for HD?^

While this study focuses on Parkinson’s disease, overcoming the challenge of effectively delivering medicine across the blood-brain barrier could be useful in preventing damage or repairing brain cells in any of the neurodegenerative disorders, including Huntington’s disease. If HD scientists develop drugs or treatments that could slow down or reverse neuronal death in Huntington’s disease patients, macrophage-based delivery, as pioneered in this research, could be a method for getting that medication across the blood brain barrier and into regions of the brain where the medication is needed. If a method of drug delivery, such as Trojan therapy, could be proven safe, effective, and broadly applicable, it would allow scientists to concentrate on drug development.

For Further Reading

1. Haney, M. J., Zhao, Y., Harrison, E. B., Mahajan, V., Ahmed, S., He, Z., … & Batrakova, E. V. (2013). Specific Transfection of Inflamed Brain by Macrophages: A New Therapeutic Strategy for Neurodegenerative Diseases. PloS one, 8(4), e61852.
2. Ahmed, Abdul-Kareem. “A Trojan Treatment for Parkinson’s Disease.” MIT Technology Review. MIT, 14 Oct. 2013. Web. 04 Nov. 2013.
3. Zomer A, Vendrig T, Hopmans ES, van Eijndhoven M, Middeldorp JM, et al. (2010) Exosomes: Fit to deliver small RNA. Commun Integr Biol 3: 447–450.

KP 12/6/13 More

Transgenic HD Monkey Models


This article will discuss the advantages and disadvantages of using transgenic monkeys to model Huntington’s disease (HD). Most HD animal research utilizes mouse models of the disease. While there is much that we can learn from mice, animals that are more similar to humans, such as monkeys, could offer more pertinent insights into HD and serve as brand new and promising avenues for HD research. Along these lines, transgenic rhesus macaques carrying a human mutant huntingtin gene have been developed at Yerkes Primate Center in Atlanta, Ga. In addition, transgenic marmosets carrying a fluorescent protein have recently been developed at Keio University, in Japan. Researchers plan to use marmosets to create either a Parkinson’s or HD model in the near future. The new models offer a host of advantages that no other animal model has provided, but they are also constrained by cost, ethical considerations, and time.

Advantages of using a Monkey Model

Monkeys are more genetically similar to humans than rodents. As a result, they have a similar lifespan, metabolism, and physiology to humans. For these reasons, monkeys will probably be better models for monitoring disease progression and the effectiveness of experimental drugs. The transgenic monkeys with the mutant huntingtin gene exhibit an HD phenotype closer to humans and more closely mirror the physical, behavioral, and cognitive symptoms of the disease than any other HD animal model so far.

A variety of established behavioral and cognitive tests are used to assess the monkeys. One such test is the HD primate model scale, modified from the HD scale used for humans. The scale ranges from 0 to 80, in which 80 describes the most severe symptoms and is used to track the number of involuntary movements in the transgenic rhesus macaques. The test shows that the monkeys display chorea and dystonia more clearly than many HD mouse models.

In addition to physical tests, non-invasive fMRI procedures are used to monitor neurodegeneration, and intranuclear huntingtin inclusions, as well as other features of the disease at the neural level. Monkeys have larger brains than rodents, so neural changes can be monitored more accurately. Moreover, a germline of macaque pluripotent stem cells with the mutant huntingtin gene has been developed and is currently being studied by the same researchers. These stem cells can be coaxed into becoming neurons, which could then demonstrate the neural symptoms of the disease. Ethical concerns have prevented the development of a similar human germline, so the transgenic rhesus monkeys provide both an in vivo and in vitro avenue to study the neural progression of HD.

Disadvantages of using a Monkey Model

Many of the advantages of monkey models come hand in hand with their problems. Primate models are more expensive, take more time, and raise more ethical concerns than mouse models. The rhesus macaque for example has one baby at a time, a gestation period of 150 to 160 days, and a long puberty of 3 to 4 years.  The constraints and labor involved in natural and artificial reproduction make the cost of transgenic monkeys significantly higher than transgenic mice. Similarly, monkeys are much larger than mice, so their housing and food costs are more expensive. Because monkeys display HD symptoms several years after they are born, these costs for monkeys are exponentially greater than those for mice. Because macaques and marmosets are similar to humans in terms of emotions, cognition, and behavior, the use of monkeys in lab research raises more ethical concerns and media interest than other animal models. The monkeys are protected by stringent ethical guidelines and procedures determined by the National Institute of Health and the Institutional Animal Care and Use Committee. The use of monkeys in research sparks public interest more than rodents, which can bring both positive and negative attention to transgenic primate research and HD animal model research in general.

Different types of monkeys have their advantages and disadvantages. The rhesus macaque is more closely related to humans than the marmoset but the reproductive traits of the marmoset make it an attractive model in its own right.  The marmoset can have 80 babies over its lifetime, compared to that of 10 offspring for the macaque. The marmoset also has a shorter pregnancy and a faster sexual maturation, making the marmoset model perhaps a more efficient model for studying HD. The marmoset is also smaller making housing and feeding them more manageable economically. However, the marmoset has a smaller brain, making it more difficult to track neurodegeneration on an MRI. Both of these monkey species have far longer pregnancies, pubertal periods, and fewer offspring than a female mouse, who can have up to 10 litters of 3 to 14 mice per year. However, animal models more similar to humans could not only accelerate discoveries in the field, but could potentially reduce the total number of animal models needed in research.

Method of Transgenesis

There are a variety of animal models that are designed through genetic engineering in order to study HD. The only type of monkey model so far is the transgenic model. In this type of model, a transgene is integrated into the animal’s genome. An example of the method used for the transgenic macaques is as follows.  A mutant human huntingtin gene with 84 CAG repeats is inserted into rhesus macaque egg cells through a viral vector. The viral vector is a modified lentivirus, which is a virus commonly used for gene delivery because it infects non-dividing cells. The lentivirus contains the gene encoding for the mutant huntingtin gene as well as a gene encoding green fluorescence protein (GFP) to serve as a marker, so that the scientists can tell whether or not the transgene was integrated into the genome of the monkey. Many of these HD eggs are then artificially fertilized and implanted into female surrogate monkeys. The gene is integrated randomly into the embryo’s genome through reverse transcriptase. Of the HD monkeys that are born, each will vary in terms of the location of the human mutant huntingtin gene on the chromosomes, the number of CAG repeats, and severity of the HD phenotype. Essentially, HD manifests differently in each monkey due to the nature of the transgenic procedure. However, the mutant huntingtin gene is still heritable and dominant despite its variable location in the chromosomes so natural and artificial offspring of transgenic monkeys can also be used to study HD.

The advantages of transgenesis through the lentivirus vector are that it is very effective and that HD animal models can be produced in high yield. However, through this method the mutant human huntingtin gene could be integrated anywhere in the genome rather than where the gene is normally located. Some knock-in mice have been produced in which the human gene replaces, at least in part, the mouse’s huntingtin gene in the correct location on the chromosome. They should, in theory, be more accurate models than transgenic mice. A knock-in monkey is theoretically feasible, but the laboratory techniques are not yet efficient enough to make this cost-effective.

Current and Future Research

There are currently only two branches of research on transgenic non-human primates at this time: macaques and marmosets. The first successful transgenic monkey was developed in 2001, when Dr. Anthony Chan and his colleagues at Oregon Regional Primate Research Center inserted a GFP gene into an embryo of a rhesus macaque via a retrovirus.  Chan then moved to Emory University where he developed the first HD macaque model in 2008. Five transgenic monkeys were born and four of those expressed HD symptoms. The macaques had integrated human mutant huntingtin genes that varied in CAG repeats and integration location. Two of these monkeys died within the same day of birth, probably because they had longer CAG repeats. This feature is known to cause a quicker onset of symptoms in humans, and thus expressed a more severe phenotype –motor impairment and difficulty breathing– than the others.  They both also had multiple mutant huntingtin integration sites, which may have lead to the overexpression of the huntingtin gene as well. One monkey is still living today. Its huntingtin protein had only 29 CAG repeats, within the range of a normal huntingtin gene, so it will probably not develop the disease. The other two monkeys had 83 and 84 CAG repeats and lived long enough to be studied by the researchers. All the monkeys were studied based on their physical behaviors of dystonia, chorea, difficulty swallowing and difficulty breathing. These symptoms manifested in varying degrees in all monkeys except the monkey with 29 CAG repeats.

The lab now has a second generation of monkeys that are currently being studied. They are from the same germline of one of the monkeys from the first generation, meaning they are genetically related. Though the research is unpublished, the new group of monkeys model HD even better, reflecting a delayed onset of the disease and milder phenotypes. The macaques’ brains will be studied through fMRI, which is non-invasive. The macaques could potentially be used to study promising HD medications.

In 2009, Japanese researchers developed the first transgenic marmoset model expressing GFP. Five monkeys made up the first generation of transgenic marmosets. The marmosets glow under a specific wavelength of light because, like the macaques, they carry the GFP gene derived from jellyfish DNA. The researchers hope to use the same techniques to integrate genes coding for Parkinson’s, but are also considering HD. Because the marmosets reproduce more frequently and in larger numbers, transgenic marmosets could even further accelerate HD research.

HD is easier to study than some other diseases because it involves a mutation in one gene, where as Parkinson’s, ALS, and Alzheimer’s have more complex genetic origins. So HD is, in many ways, a gateway for all neurodegenerative research.  As more labs start working with transgenic non-human primate models, advances in understanding the progression, physiology, and neurobiology of HD are sure to follow. Because HD monkeys are also an intermediate between mouse and humans, they serve as a new avenue for drug research, which could reduce the time it takes to bring a new drug to market. The new monkey models put HD research on the cusp of major breakthroughs for both understanding and treating the disease.

Further Reading

Anthony W.S. Chan, Pei-Haun Cheng, Adam Neumann and Jin-Jing Yang (2010) Reprogramming Huntington Monkey Skin Cells into Pluripotent Stem Cells. Cellular Reprogramming (In Press). Dense and unnecessary to read.

Anthony W.S. Chan (2009). Transgenic primate research paves the path to a better animal model: are we a step closer to curing inherited human genetic disorders? J Mol Cell Biol 1(1):13-14. PMID:19671628. Great article that discusses the advantages of a monkey model and is easy to read.

Anthony W.S. Chan and Shang-Husn Yang (2009). Generation of Transgenic Monkeys with Human Inherited Genetic Disease. Methods 2009 May 23 [Epub ahead of print] PMID: 19467335. Scientific research and difficult read but helpful for understanding setting up a germline.

Bachevalier, Stuart M.  Zola, Shihua Li, Xiao-Jiang Li and Anthony WS Chan (2008) Toward a Transgenic Model of Huntington’s Disease in the Nonhuman Primate. Nature 453(7197): 921-924. PMID:18488016. Scientific and dense but excellent description of methods used for transgenesis.

B.Snyder, A. M. Chiu, D. Prockop and Anthony W.S. Chan (2010) Human Multipotent Stromal Cells (MSCs) Increase Neurogenesis and Decrease Atrophy of the Striatum in a Transgenic Mouse Model for Huntington’s Disease. Dense and unnecessary to read.

Chan, A. W. Personal Interview.  4 Jan. 2013.

Cyranoski, David. “Marmoset Model Takes Centre Stage.” Nature Publishing Group, 27 May 2009. Web. Good summary article of more difficult published research.

Nature 459, 523-527 (28 May 2009) | doi:10.1038/nature08090; Received 27 September 2008; Accepted 30 April 2009. Difficult but worthwhile read on the marmoset models.



Neuroimaging refers to techniques that produce images of the brain without requiring surgery, incision of the skin, or any direct contact with the inside of the body.  Because these technologies enable noninvasive visualization of the structure and functionality of the brain, neuroimaging has become a powerful tool for both research and medical diagnosis.  Although still relatively young, the field of neuroimaging has rapidly advanced over the years due to breakthroughs in technology and computational methods.  Applications of neuroimaging techniques have likewise become far-reaching.

This article will cover three of the most common techniques in neuroimaging: computerized tomography (CT), functional magnetic resonance imaging (fMRI), and positron emission tomography (PET).  You will likely come across at least one of these techniques while reading about Huntington’s disease (HD) news and research.  For each technique, the article will discuss its working mechanism, its pros and cons, and its relevance to HD.

Computerized tomography (CT)^

Computerized tomography (CT) is a medical imaging procedure that uses x-rays to generate detailed pictures of structures inside the body.  It has become a valuable tool for medical diagnosis and for planning, guiding, and monitoring therapy.

Attribution: Aaron G. Filler, MD, PhDA CT scan of a human brain. (Source: Aaron G. Filler, MD, PhD)

How CT works^

A CT scan uses X-rays positioned at different angles to create cross-sectional images of the brain.  During a CT scan, a movable X-ray source is rotated around the patient’s head.  Detectors record information about the intensity of the rays transmitted through the head at each angle, which is sort of like taking a series of two-dimensional snapshots.  The computer then uses an algorithm to combine all the individual snapshots from different axes of rotation of the source together and to reconstruct them into a 3-D, cross-sectional image.  By allowing doctors and researchers to view the full volume of the head, CT scans can provide much more information about the brain than traditional X-ray scans, which only offer a two-dimensional representation of the brain.

Advantages and Disadvantages of CT^

CT scans are painless, fast, and cost-effective.  CT can image bone, blood vessels, and soft tissue simultaneously and provides very detailed images of many types of tissue.  There are, however, several risks associated with the use of CT.  The main risks, according to the US Food and Drug Administration, are:

  • An increased lifetime risk of cancer due to x-ray radiation exposure
  • Possible allergic reactions or kidney failure due to contrast agent, or “dye” that may be used in some cases to improve visualization
  • The need for additional follow-up tests after receiving abnormal test results or to monitor the effect of a treatment on disease

You and your physician should decide whether or not the benefits of getting the added information from a CT scan outweigh the possible risks.

CT and Huntington’s disease^

CT cannot by itself be used to diagnose HD, since other disorders or conditions can result in similar anatomical change in the brain; often it is used along with family history and medical records in order to provide a more detailed diagnosis.  (The same is true of MRI scans, which will be discussed in the upcoming section.) Doctors use CT (or MRI) to evaluate a patient with HD by checking for characteristic degeneration in the brain and ruling out other possible disorders.  A CT scan for a patient in the early stages of HD may appear normal, but a CT scan for a patient in advanced stages of the disease can often reveal significant degeneration – specifically, reduction in volume of certain regions of the basal ganglia.

Functional Magnetic Resonance Imaging (fMRI)^

Functional magnetic resonance imaging (fMRI) is a specialized form of MRI that indirectly provides information on brain activity by measuring changes in blood flow in the brain.

Public domain, NASA

An fMRI scan of a human brain. (Public domain)

How MRI works^

A traditional MRI scanner contains a very strong electromagnet, which generates a strong magnetic field inside the scanner.  When turned on, this causes randomly spinning protons, or positively charged particles, in the brain (in this case hydrogen protons from water molecules) to align themselves with the direction of the field.  (Think of how a compass needle at rest is aligned with the Earth’s magnetic field, which is why it always points north.) However, the protons still spin while in alignment in the field, and the axis of their spin isn’t perfectly parallel to the direction of the field – rather, the protons behave like wobbling tops. The rate at which the protons wobble is called resonance.

We can also think of resonance as the ability of a system to absorb energy delivered at a particular frequency.  To illustrate this concept, imagine that you are pushing a small child on a playground swing.  If you push the child too quickly or too slowly, the child won’t rise very high.  But if you give the child enough pushes at just the right frequency, the child will rise to the maximum height attainable due to the energy you have provided by pushing.  This frequency is called the resonant frequency of the swing, which is the rate at which the swing will naturally oscillate.  The swing absorbs the most energy when it is pushed at exactly this frequency.  Similarly, protons placed in a strong magnetic field will efficiently absorb energy when the energy is delivered at a particular resonant frequency, namely the rate of “wobbling.”  In the case of MRI, radio waves provide the “push” necessary to move the protons.

During a process called excitation, the MRI scanner emits energy in the form of radio waves at precisely the resonant frequency of protons.  These radio waves knock the protons out of alignment, causing them to flip and spin on their opposite ends.  In order to flip over, the protons have to absorb some energy from the radio waves.  When the radio signal is turned off, the protons flip back around to their original alignment, and in the process release the energy they have absorbed.  This released energy is called the MR (magnetic resonance) signal and can be measured by electromagnetic detectors around the subject.  A computer receives the MR signals as mathematical data and compiles them into an image.

Although fMRI uses the same principles as MRI, there is one important distinction.  MRI (often called structural MRI) reveals brain anatomy, while fMRI reveals brain function, often in the form of neural activity.  For this reason, fMRI is very useful for neuroscience and clinical research, while MRI is traditionally used for clinical characterization in the same manner as CT scans.

However, fMRI doesn’t reveal neural activity directly.  Recall that when neurons are active, they fire action potentials and send messages in the form of neurotransmitters to other neurons. (To read more about this, click here.) While other imaging methods such as EEG (electroencephalography) can measure this activity directly, they often come with severe limitations such as poor spatial resolution.  Instead, fMRI studies neural activity indirectly by measuring the BOLD (Blood Oxygenation Level Dependent) signal.  This method is based on the fact that hemoglobin carrying a bound oxygen molecule (oxyhemoglobin) in the bloodstream emits a different MR signal than oxygen-depleted hemoglobin (deoxyhemoglobin).  When parts of the brain become active, such as when a person is carrying out a cognitive task, they use up more oxygen than relatively inactive parts of the brain.  You might think that this would result in a relative decrease in the level of oxygen in the active areas compared to the inactive areas, but actually the reverse happens!  Because oxygen is being depleted, the brain compensates for the loss by increasing the flow of oxygenated blood to the active area.  There is a slight overcompensation, which causes an increased oxyhemoglobin to deoxyhemoglobin ratio.  As this ratio increases, the BOLD signal gets stronger.  Essentially the BOLD signal measures the ratio of oxyhemoglobin to deoxyhemoglobin in the brain, which enables researches to indirectly gauge neural activity: higher neural activity = higher oxyhemoglobin to deoxyhemoglobin ratio = stronger BOLD signal.

Advantages and Disadvantages of fMRI and MRI^

MRI and fMRI both involve no radiation, and there are no known side effects caused by the magnetic fields and radio waves.  The main risk with MRI is that the presence of metal in the body (such as pacemakers or other implants) can be a safety hazard.  fMRI is also relatively inexpensive, non-invasive, widely available, and provides excellent spatial resolution and good temporal resolution.  Largely because of these factors, fMRI has come to predominate in the field of neuroimaging research.

fMRI and Huntington’s Disease^

Use of fMRI or MRI scans in HD patients sometimes requires sedation since both techniques require remaining extremely still and the results can be ruined by small movements.  It is thought that fMRI can potentially be used as an indicator, or imaging biomarker, of early neuronal dysfunction in individuals before the onset of HD (a stage called pre-HD).  A 2004 study conducted with pre-HD patients demonstrated that fMRI could detect abnormalities in brain activity more than 12 years before the estimated onset of motor systems (Paulsen et al).  fMRI was also able to distinguish differences between patients closer to estimated onset and patients further away from estimated onset.  These results suggest that fMRI may be useful for tracking changes in neural function during the early stages of pre-HD, and could improve the prediction of when HD manifests in an individual.  The study also suggests that fMRI can be helpful in determining the optimal time frame of treatments aimed at slowing down the progression of HD.  However, further research is required to determine whether the potential of fMRI technology will be realized.

Positron Emission Tomography (PET)^

Positron emission tomography is a technique that uses radioactively labeled molecules (called tracers) that are injected into the bloodstream and taken up by active neurons.  PET studies blood flow and metabolic activity in the brain and helps visualize biochemical changes that take place.  Essentially, PET indicates how well the brain is functioning.

Public domain, Jens LangnerA PET scan of a human brain. (Public domain)

How PET works^

The scanner consists of a ring of detectors that surround that subject.  Detectors contain crystals that scintillate (give off light) in response to gamma rays, which are extremely high-energy rays of light.  Each time a crystal in a detector absorbs a gamma ray is called an event.  When two detectors exactly opposite from each other on the ring simultaneously detect a gamma ray, a computer hooked up to the scanner records this as a coincidence event.  A coincidence event represents a line in space connecting those two detectors, and it is assumed that the source of the two gamma rays lies somewhere along that line.  The computer records all of the coincidence events that occur during the imaging period and then reconstructs this data to produce cross-sectional images.

The tracer is usually a substance, such as a type of sugar like glucose, that can be broken down (metabolized) by cells in the body, and it is labeled with a radioactive isotope.  There is minimal risk involved since the dose of radiation is low and the isotope is quickly eliminated from the body through urination.  After it has been injected into the bloodstream, the isotope, which is very unstable, starts to decay, becoming less radioactive over time.  In the process it emits a positron (a positively charged electron).  When a positron collides with an electron, the two particles annihilate each other, producing two gamma rays with the same energy but traveling in opposite directions.  These gamma rays leave the subject’s body and are sensed by two detectors positioned 180 degrees from each other on the scanner, which gets recorded as a coincidence event.  A computer can determine where the gamma rays came from in the brain and generate a three-dimensional image.

As blood is more concentrated in activated brain areas than in inactivated ones, the scanner will detect more gamma rays coming from parts of the brain that are working harder.  On a PET scan regions of the brain show up as different colors depending on the degree of activity in those regions.  Yellow and red regions are “hot” and indicate high brain activity, while blue and black regions indicate little to no brain activity.

The brain function measured by a PET scan varies according to the type of radioisotope that is used.  For instance, oxygen-15 is used to study oxygen metabolism in the brain.  FDG, which is fluorine-18 attached to a glucose molecule, is used to study sugar metabolism in the brain.  Many more radioisotopes exist, and which one is chosen for a specific PET scan depends on what type of brain function a researcher desires to study.

Advantages and Disadvantages of PET^

PET, unlike other imaging tests, is able to detect irregularities in body function caused by disease, which often occur before anatomical changes become observable.  The quality of a PET scan is not affected by small movements, so the subject does not have to remain as still for a PET scan as they would for a fMRI or MRI scan, both of which can be ruined by small movements.

A main setback to PET is that it affords relatively poor spatial resolution, so the images may not be very clear.  Due to this, it is common for PET to be used together with CT or fMRI.  In addition, the use of radiation, even in a small dose, always involves a slight risk.

PET and Huntington’s disease^

PET scans are sometimes ordered by physicians as a follow-up to a CT or MRI scan in order to reveal any irregularities in brain processes.  They can be used as part of a more detailed diagnosis for HD, as well as in research related to abnormalities in metabolism and progression of the disease.   Nowadays PET scans are often combined with CT scans to provide images that pinpoint the location of abnormal activity within the brain; combined, these scans provide more accurate diagnoses than either performed alone.


CT, fMRI/MRI, and PET are three of the most popular techniques currently used for neuroimaging.  Each method comes with its own advantages as well as its own risks and disadvantages.  While neuroimaging is still a fairly new development in medicine and neuroscience, the discipline will likely continue expanding in the future, and will continue to provide valuable clinical applications and scientific insights about the brain.

Works Cited^

Paulsen JS, Zimbelman JL, Hinton SC, Langbehn DR, Leveroni CL, Benjamin ML, Reynolds NC, Rao SM. fMRI biomarker of early neuronal dysfunction in presymptomatic Huntington’s Disease. AJNR Am J Neuroradiol. 2004 Nov-Dec;25(10):1715-21.

Further Reading^

FAQ about getting CT scans

A more detailed explanation of how MRI works
Additional information about fMRI

FAQ about getting PET/CT scans

J. Nguyen 4.9.12


Weight Loss: Demystifying a Medical Mystery

While Huntington’s disease is traditionally thought of as a disease of the brain, its effects are much more widespread: many people with HD lose a dangerous amount of weight, complicating a disease that is already complicated enough. Although weight loss is one of the most serious non-neurological problems of HD, scientists don’t fully understand why it occurs. This medical mystery has driven scientists deep into the biology underlying weight loss in HD. Researchers have recently turned up a few potential explanations, and our increased understanding of this symptom is leading scientists to look at possible new ways of treating the disease.

Weight Loss in HD^

People with HD tend to weigh less than those without the disease. A group of researchers from the Huntington Study Group followed 927 people with early-stage HD. For a description of the stages of HD, please click here. The investigators found that people with early-stage HD weighed an average of 10 kilograms (22 pounds) less than age-matched controls, which are people of the same age who don’t have the disease. Another study found that people with HD lose an average of 0.9 pounds per year, which stands in stark contrast to the average American, who gains 0.4-2 pounds yearly.

Unfortunately, while 0.9 pounds doesn’t seem like much, that’s just an average; some people with HD lose so much weight that their health is impacted. Weight loss worsens other aspects of the disease as underweight patients become malnourished and weak. Underweight patients are more susceptible to infection, and take longer to recover from illness, operations, and wounds. Weight loss also increases the likelihood of developing pressure ulcers, commonly known as bedsores, as bedridden patients have less fat tissue to cushion them from pressure. Patients who lose the most weight report a lower quality of life, and are more likely to feel apathetic and depressed. In the late stages of the disease, some patients lose so much weight that they need a feeding tube to stay healthy, as described here. On the other hand, people who start out heavier fare better; people who have a high body-mass index (BMI) when symptoms begin progress more slowly through the disease. Visit this website for an explanation of BMI and a for BMI calculator.

A Medical Mystery^

While weight loss is one of the most serious non-neurological problems associated with HD, doctors don’t understand why it happens. Many suggestions have been put forth, but most of them have been disproved, forcing researchers to dig deeper to understand this phenomenon.


Doctors once believed that weight loss was due to chorea, the uncontrolled movements characteristic of HD. Doctors thought that people with HD lost weight because they burned extra energy as a result of the involuntary movements of chorea. However, three experiments indicate that chorea can’t be fully responsible for weight loss.

The first piece of evidence comes from looking at the early stages of the disease. People who have just been diagnosed with HD – and therefore have very mild symptoms – already weigh less than people without the disease. As mentioned earlier, people in early-stage HD weigh an average of 10 kg less than those who are not affected by the disease. Another group of researchers arrived at similar results; a study of 361 people with early-stage HD found that they have BMIs an average of 2 points lower than those without the disease, even if the patients had just been diagnosed with HD within that year and hadn’t yet begun to experience choreic movements. Researchers concluded that chorea alone could not explain why people with HD have lower BMIs, and that other factors are at play.

Other studies suggest that chorea may not have as much of an impact as doctors once thought. Pratley et al. measured how much movement chorea caused, in an attempt to quantify how much weight patients lose due to choreic movement. After measuring the movements of 17 people with mild to moderate HD for a week, they found that chorea caused people with HD to move more than people without the disease when sedentary: people with HD moved 14% more than people without HD while sitting or lying down. However, people with HD do less voluntary activity. Study participants with HD walked around and exercised less than people without the disease. In the end, Pratley et al. were surprised to discover that sedentary over-activity balanced out voluntary under-activity: people in the early and middle stages of HD don’t actually move more than people without the disease.

A similar study by the European Huntington’s Disease Initiative Study Group (EHDI) measured weight loss in 517 people with HD, and found no correlation between the amount of weight people lost and the severity of their motor symptoms; people with good scores on tests measuring motor symptoms (such as the UHDRS) were just as likely to lose weight as those with bad motor scores. For more information on diagnostic tests like the UHDRS, click here.

The final strike against the chorea theory comes from observations of people with late-stage HD. Weight loss is most drastic in the final stages of HD, despite the fact that chorea has usually ceased and patients are largely bedridden. So while chorea contributes to weight loss in HD, it cannot stand as the sole explanation.

Reduced Food Intake^

Others suggest that people with HD lose weight because they have trouble eating; as the disease progresses, it becomes increasingly difficult to perform the complicated series of movements needed to eat, chew, and swallow.

However, this theory is also not enough to fully explain the weight loss. Studies have shown that people with HD actually tend to eat more than people without the disease; a study of 25 people with HD found that they ate an average of almost 400 calories more each day than people without the disease. Others report that they’ve had patients who eat up to 5000 calories a day – over twice the average daily caloric intake – just to maintain their weight.

So two popular explanations for weight loss in HD – chorea and insufficient diet – cannot entirely explain why people with HD lose so much weight.

Possible Biological Causes^

Though the reasons for the mysterious weight loss are unclear, scientists are currently testing a few ideas.

Abnormalities in Energy Metabolism^

One leading idea has to do with metabolism, the way the body burns calories to produce energy. HD researchers have long suspected that the disease-causing form of huntingtin (hereafter described as mutant huntingtin) interferes with energy metabolism, as described here. Results from a recent study suggest that this interference might contribute to weight loss.

After discovering that weight loss is not correlated with motor symptoms, scientists from the EHDI Study Group looked for other factors that might be to blame. They found that weight loss could be partially predicted by the number of CAG repeats on a patient’s copy of the mutant huntingtin gene; for every additional CAG repeat a patient had, they lost on average an extra 0.136 BMI points (0.8 pounds) over the course of the three year period that the study was conducted. For an explanation of CAG repeats, please click here.

The same holds true in mouse models of HD. The EHDI Study Group found that the more CAG repeats an HD mouse had, the more it tended to eat. Yet paradoxically, the mice with the most CAG repeats lost the most weight. So people and mice with more CAG repeats lose more weight.

The EHDI investigators suspect that this is due to the long tail of the mutant huntingtin protein. People with more CAG repeats produce mutant huntingtin with a longer tail, as described here. The EHDI investigators suggest that the mutant huntingtin protein interferes with the way cells make energy, and that longer-tailed proteins cause more problems. Mutant huntingtin has been shown to disrupt proteins that are needed to make energy and can damage mitochondria, the “energy factory” of our cells, as described here. In support of the theory that proteins with longer tails are more problematic, scientists at the MacDonald lab in Boston studied cells engineered to express mutant huntingtin. They found that cells with more CAG repeats made less ATP, the energy currency of the cell. So it seems possible that the more CAG repeats individuals have, the less efficient their cells are at converting calories to energy.

Hormonal Shifts^

A second school of thought suggests that weight loss is due to hormonal disturbances in people with HD. Hormones are the body’s chemical messengers, and are important for regulating physiological processes, like hunger. The hypothalamus secretes many hormones, so when HD causes cells in the hypothalamus to malfunction and die, hormone production is disturbed.

Some of the hormonal signals that the hypothalamus sends out go to the gut and fat tissue, and direct processes like eating and burning energy – processes that are very important in maintaining a healthy weight. Therefore, some scientists think that cell death in the hypothalamus causes hormonal changes that might contribute to weight loss and other problems such as sleep disturbances, as described here.

Dysfunctional Digestion^

Further insights have come from studying the way mutant huntingtin interacts with the digestive system. Certain symptoms of HD have hinted that the disease might affect the gut; apart from weight loss, people with HD often experience nutritional deficiencies, cramps, and wasting of skeletal muscles. People with HD are also prone to gastritis, a disease where the stomach lining becomes irritated or swollen.

Despite these symptoms, many HD researchers have traditionally thought that mutant huntingtin only affected the brain – a belief that struck some as strange because the protein is made and found throughout the body. However, results from a recent study suggest that mutant huntingtin in the gut might interfere with important digestive processes, thus contributing to weight loss.

In the study, van der Burg and colleagues looked at R6/2 mice, which are mouse models of HD described in greater detail here. They noticed several physiological changes that could all impact digestion. First, they noticed that the small intestines of HD mice were 10-15% shorter than those of normal mice, and that they had smaller villi, the tiny finger-like projections in the gut that take up nutrients. On top of that, scientists noticed that the mucus lining of the gut of the HD mice was 20-30% thinner. Since all of these structures are needed for nutrient absorption, these findings suggest that HD mice can’t take up nutrients as efficiently as normal mice.

Furthermore, the group found that the HD mice were missing a few key hormones that control the speed at which food passes through the body. This caused an increase in ‘transit time’: the food passed more slowly through the gut. Longer transit time might foster bacterial growth; if food takes longer to pass through the gut, harmful bacterial have more time and a better opportunity to flourish. This could make the small intestine irritated and inflamed, which could cause malabsorption of nutrients, chronic diarrhea, nausea, bloating, flatus, and weight loss. Those bacteria might also use up nutrients that the body would have otherwise taken up.

To see whether these physiological differences actually have an impact on digestion, researchers then compared the feces of HD mice to those of normal mice. They found that HD mice excreted more of what they ate, suggesting that they absorbed fewer calories and nutrients from their food. Notably, the mice that were the worst at absorbing nutrients from their food lost the most weight.

Van der Burg et al. had a few ideas as to what mutant huntingtin might be doing to interfere with digestion. Since the protein is present in gut cells, it could interfere with cell function and nutrient absorption. They also thought that mutant huntingtin might affect transcription, the process by which DNA is converted into protein as described here. If mutant huntingtin affects transcription in gut cells, it could cause a decrease in levels of important proteins needed for cells to survive and function properly.

While findings in HD mice don’t always translate to humans, these results indicate that scientists might benefit from studying the way HD affects digestion in people. Van der Burg et al. suggest that such research might help doctors improve their understanding of nutritional supplements for HD, and might even change the way we think about how people with HD metabolize and react to medicine.


Weight loss in HD has long puzzled doctors, patients, and caretakers alike. Two popular explanations of the phenomenon – chorea and reduced food intake – have been debunked as major contributors to weight loss. However, scientists have made new in-roads in recent years. By discovering that mutant huntingtin might disrupt energy metabolism, digestion, and hormones in HD mice, scientists have enhanced our understanding of HD, which may pave the way to new treatments and therapies. For example, the hypothesis that weight loss is linked to abnormalities in energy metabolism suggests that energy-boosting drugs – namely creatine and Coenzyme Q10 – are strong candidates to fight HD, as described in these articles here. Each further discovery about HD leads to a greater understanding of the disease, and brings hope for patients and families.


1.     Aziz NA, van der Burg JM, Landwehrmeyer GB, Brundin P, Stijnen T; EHDI Study Group, Roos RA. Weight loss in Huntington disease increases with higher CAG repeat number. Neurology. 2008 Nov 4;71(19):1506-13.

This medium-difficulty study describes how people with more CAG repeats lose more weight – and provides some theories as to why that might be the case.

2.     Djoussé L, Knowlton B, Cupples LA, Marder K, Shoulson I, Myers RH. Weight loss in early stage of Huntington’s disease. Neurology. 2002 Nov 12;59(9):1325-30.

This medium-difficulty article describes weight loss in people with early-stage HD

3.     Hamilton JM, Wolfson T, Peavy GM, Jacobson MW, Corey-Bloom J; Huntington Study Group. Rate and correlates of weight change in Huntington’s disease. J Neurol Neurosurg Psychiatry. 2004 Feb;75(2):209-12.

This medium-difficulty article describes weight loss in people with early-stage HD

4.     Kremer HP, Roos RA. Weight loss in Huntington’s disease. Arch Neurol. 1992 Apr;49(4):349.

This short, medium-difficulty column suggests that cell death in the hypothalamus could contribute to weight loss

5.     Petersén A, Björkqvist M. Hypothalamic-endocrine aspects in Huntington’s disease. Eur J Neurosci. 2006 Aug;24(4):961-7. Epub 2006 Aug 21. Review

This technical article describes how hormonal changes in people with HD might lead to weight loss

6.     Pollard J, Best R, Imbrigilo S, Klasner E, Rublin A, Sanders G, Simpson W. A Caregiver’s Guide for Advanced-Stage Huntington’s Disease. Huntington’s Disease Society of America, 1999.

This easy-to-read handbook is a very helpful resource for caregivers taking care of people in late-stage HD

7.     Pratley RE, Salbe AD, Ravussin E, Caviness JN. Higher sedentary energy expenditure in patients with Huntington’s disease. Ann Neurol. 2000 Jan;47(1):64-70

This study measured movements of people with HD, and found that their total energy expenditure was the same as that of people without the disease, and is somewhat technical

8.     Seong IS, Ivanova E, Lee JM, Choo YS, Fossale E, Anderson M, Gusella JF, Laramie JM, Myers RH, Lesort M, MacDonald ME. HD CAG repeat implicates a dominant property of huntingtin in mitochondrial energy metabolism. Hum Mol Genet. 2005 Oct 1;14(19):2871-80. Epub 2005 Aug 22.

This technical article describes how huntingtin interferes with energy metabolism in a CAG-dependent fashion

9.     Trejo A, Tarrats RM, Alonso ME, Boll MC, Ochoa A, Velásquez L. Assessment of the nutrition status of patients with Huntington’s disease. Nutrition. 2004 Feb;20(2):192-6.

This medium-difficulty paper discusses the result of the study on 25 HD patients that ate an average of 400 calories more than controls each day.

10. van der Burg JM, Winqvist A, Aziz NA, Maat-Schieman ML, Roos RA, Bates GP, Brundin P, Björkqvist M, Wierup N. Gastrointestinal dysfunction contributes to weight loss in Huntington’s disease mice. Neurobiol Dis. 2011 Oct;44(1):1-8. Epub 2011 May 23.

This technical article describes the impact of huntingtin on digestion in HD mice

M. Hedlin 11.16.11


University of California at San Francisco 2010 HD Research Symposium

HOPES summary of the talks from scientists and clinicians

Note: This article includes references to Dimebon, which is no longer being considered as a potential treatment for HD after the HORIZON clinical trial showed that Dimebon was not better than a placebo. For more information, click here

Lisa Ellerby, PhD, Buck Institute for Age Research^

Target Validation in Huntington’s Disease

Dr. Ellerby’s talk focused on the question: “What are possible biological targets for drugs designed to treat HD?” Therapeutic drugs work by targeting specific processes in the human body that have gone awry in disease. For example, the common symptom chorea in HD is thought to be a result of the increased activity of the neurotransmitter dopamine. Tetrabenazine is used to treat chorea because the drug reduces the amount of dopamine in the brain.

At present, the pharmaceutical industry is focused on developing therapeutics for approximately 200 to 300 different targets in the human body that may be related to HD. However, those few hundred targets represent a very small subset of all possible biological targets and it is possible that drugs that have been created for other diseases could have therapeutic benefits for individuals with HD. One example of this is Dimebon, a drug that was originally used as an anti-histamine and is now being studied in clinical trials for HD because of its neuroprotective effects.

Dr. Ellerby is interested in identifying targets that play specific roles in the death of neurons in HD. While it is well known that the mutated Huntingtin protein (Htt) results in the neurodegeneration characteristic of HD, Dr. Ellerby’s research is important because it can provide insight into how this neuronal death occurs. Her lab used small interfering RNA (siRNA) to block the production of different proteins and then assessed the effects of these knockdowns on neuronal death. If shutting off a particular target results in decreased neuronal death, it is possible that this target plays a role in neurodegeneration due to mutant Htt. These experiments found that blocking the activity of proteases, enzymes that break down other proteins, reduces the death of neurons in a cellular model of HD. Specifically, Dr. Ellerby mentioned that decreasing the level of a family of proteases known as matrix metalloproteases (MMP) reduces the toxicity of mutant Htt. By using siRNA to identify targets that play a role in HD, Dr. Ellerby’s research is laying a foundation for the discovery and development of drugs that can prevent, treat, and reverse the devastating effects of HD.

Jill Larimore, BSc, 4th year graduate student in neurobiology at UCSF^

Immune System Dysfunction in Huntington’s Disease

Ms. Larimore, as a member of Dr. Muchowski’s lab, researches the effect of HD on immune system function. Using yeast and animal models, her lab explores the HD on the molecular level in order to find new therapeutic targets. Ms. Larimore began her talk by explaining the immune system of the brain and the role of microglia cells. These specialized cells differ from those found in the peripheral immune system (i.e. the immune system which operates in all parts of the body besides the brain and spinal cord). Although the blood-brain barrier normally keeps these immune systems apart, Larimore’s research interestingly showed that there was peripheral immune system activation in HD patients. This indicates that the neurological symptoms of HD are either paralleled in the peripheral immune system or communication between cells traverses the blood-brain barrier to connect the two immune systems.

The Muchowski lab’s research also showed that HD increased inflammatory response in both the neural and peripheral immune system, even before manifestations of HD symptoms. Inflammation is an acute immune response that counters tissue injury by releasing chemical signals in the area of injury. Physical inflammation acts as a barrier against the spread of infection while immune cells repair the damaged tissue. Although normally beneficial, inflammation can be harmful when it becomes chronic and remains after healing. In HD patients, a key immune protein, interleukin-6 (IL-6) was found at higher concentrations both in the brain and body. IL-6 helps activate and regulate inflammatory response in the immune system, and indicates immune activity when found in heightened concentrations. In the brain, microglia activation increased as well, indicating microglia were responding to tissue damage in the brain. While immune activation could potentially be a natural healing response, Muchowski’s lab hypothesized that it was chronic inflammation that contributes to HD progression.

Similar inflammatory symptoms found in other neurodegenerative diseases have been extensively researched. Treatments have been found to regulate the heightened inflammatory response that occurs when certain immune proteins are activated. In mouse models of Alzheimer’s disease, the cannabinoid type 2 receptor (CB2) in the brain was found to decrease IL-6 and other proteins involved in the immune response in both the peripheral and neural immune systems. Inhibition of the CB2 receptor in a mouse model of HD worsened symptoms, as shown in behavioral assays (testing the mouse for motor functions and balance). Activating the CB2 receptor resulted in improved coordination and motor function, and slowed the onset of HD symptoms. Because CB2 therapeutics are already in clinical trials for autoimmune diseases, if CB2 is found to be beneficial in HD models by decreasing inflammation in the brain and the peripheral immune system these drugs could potentially be clinically tested as a therapy for HD.

Jan Nolta, PhD, Stem Cell Program, UC Davis^

Working toward mesenchymal stem cell-based therapies for HD

Dr. Jan Nolta, director of stem cell research at UC Davis, presented on recent developments in her work on therapies for HD using mesenchymal stem cells (MSCs).  MSCs have been found to deliver protein products throughout tissue for 18 months at a time. MSCs can potentially be engineered to deliver proteins that help prevent neurodegeneration to the brain. MSCs themselves exhibit neuroprotective activities. They restore synaptic connections, decrease inflammation, decrease neuron death and increase vascularization. Using vessels in the brain as train tracks, they are able to travel throughout the brain to assist other cells. Videos taken under a microscope show that MSCs are “social cells,” meaning they are constantly communicating with other cells around them. By interacting with another cell, an MSC can sense the needs of that particular cell and initiate a flow of appropriate nutrients directly into the other cell. In this way, MSCs act as cellular “paramedics” of the body.

One possibility for an HD therapy involves injecting MSCs into the brain where the cells could help reduce neurodegeneration by saving damaged neurons. Scientists at UC Davis conducted tests on non-human primates to ensure that injecting MSCs into the brain is safe for humans. Safety testing was recently completed with MSCs being injected through the skull into the brains of fetal non-human primates. Fortunately, results showed that after 5 months, the MSCs were still present. This means MSCs will be able to stay in the brain for a good length of time, theoretically assisting neurons and preventing additional cell death. Also, no tumors or tissue abnormalities were detected, indicating that MSC injection is largely safe. More studies about the intracranial transplantation and long-term MSC safety are needed.

Research on MSCs in Dr. Nolta’s lab currently involves three main goals: test bone marrow-derived MSCs to see if they restore neurons in non-human primates, test MSCs for the ability to secrete factors like brain-derived neurotrophic factor (BDNF) that help brain cell function, and to investigate MSC production of siRNA. Dr. Nolta was happy to announce that the FDA recently approved injection of MSCs into the central nervous system of individuals with another disease called amyotrophic lateral sclerosis. This sets an important precedent that will increase the likelihood that Dr. Nolta’s eventual therapy will work in other diseases.

Michael Geschwind, MD PhD, UC San Francisco Memory and Aging Center^

Update on Clinical Studies and Trials in Huntington’s Disease

Dr. Michael Geschwind, a neurologist at the UCSF Memory and Aging Center, provided updates about clinical trials in HD. First, he reviewed the two types of clinical research: observational and clinical trials. An observational study is a type of study in which individuals are observed and certain outcomes are measured (such as motor control or mental function) but no attempt is made to affect the outcome in the form of treatment or therapy. In contrast, a randomized double-blind clinical trial is a study in which each individual is assigned randomly to a treatment group (experimental therapy) or a control group (placebo or standard therapy) and the outcomes are compared. Currently, there is important and promising HD-related clinical research being conducted both within and outside of the United States. The following paragraphs summarize the significant points regarding recent or ongoing studies in the HD field.

The PREDICT-HD study is an observational study that began in 2001, was expanded in 2008, and is still underway. The ultimate goal of the PREDICT-HD study is to define the earliest biological and clinical features of HD before at-risk individuals have diagnosable symptoms of the disease. While the current approach is to treat HD at the beginning of the onset of symptoms, this study aims to help design future studies of experimental drugs aimed at slowing or postponing the onset of HD in the at-risk population prior to observable symptoms. The PREDICT-HD study has identified markers that were shown to appear long before an individual would expect to be diagnosed. One marker is CAG repeat length: CAG stands for the nucleotides (DNA building blocks), cytosine, adenine, and guanine. The HD mutation consists of multiple repeats of CAG in the DNA.  This study validated the CAG repeat length-age formula, in which the CAG repeat length for an individual could estimate the average age of HD onset.  In general, fewer than 30 repeats is considered normal, whereas more than 39 repeats means the person will likely develop HD in a normal lifespan. To read more about CAG repeat lengths click here. Other markers such as the size of ventricles in the brain and the volume of other specific brain areas (i.e. striatum) were also found.  In short, the PREDICT study has validated models for predicting motor onset of HD, which will ultimately increase the likelihood of treating HD before patients become symptomatic.

The DIMOND-HD study was a phase II clinical trial investigating the efficacy of the drug Dimebon, which is a small molecule that inhibits nerve cell death. This drug has been shown to decrease cognitive impairment in Alzheimer’s patients, and has been shown to improve cognition and memory in rats. Dimebon is often referred to as latrepirdine. The DIMOND-HD study evaluated the safety of administering Dimebon for 3 months and the efficacy of Dimebon in improving cognitive, motor, and overall function in subjects with Huntington’s Disease. The study was completed in the summer of 2008, and showed that Dimebon is a well-tolerated drug that generally improves cognition in HD. The researchers concluded that the drug should be tested in phase III clinical trials, which has resulted in the HORIZON trial described below. To read more about Dimebon click here.

The HORIZON study is a randomized, double-blind, placebo-controlled study that is ongoing at 37 sites, spanning 7 countries. The study is in phase III of clinical trials, and also aims to expand upon the results of the DIMOND-HD study and determine if Dimebon (latrepiridine) safely improves cognition in patients with Huntington’s disease.

The HART study is also a randomized, double-blind, placebo controlled study that is ongoing in both Europe and North America. The purpose of the study is to determine if ACR-16, also known as pridopidine and Huntexil, is effective and safe in the symptomatic treatment of HD. ACR-16 is a dopamine stabilizer, which means that it works to help regulate the many functions of dopamine in the striatum and other areas of the brain. ACR-16 has passed phase II of the clinical trials in Europe, and has been allowed to be tested in stage III in North America (US and Canada). Initial results of the clinical trials are promising, and have shown that ACR-16 can improve motor control and may translate to 0.5-1.5 years of disease improvement in voluntary and involuntary movements. To read more about ACR-16, click

The following table outlines the types of characteristics researchers are looking for in each of the ongoing HD Clinical Trials described above. For more information, click on the links provided.

Inclusion Criteria for HD Clinical Trials

PREDICT-HD StudyFor more information on the PREDICT-HD study click here Gene negative and gene positive individuals: specifically, men and women at risk for HD, who have been tested for the HD gene mutation, and who have not been diagnosed with symptoms of HD (CAG > or equal to 36 for CAG-expanded group or CAG < 36 for CAG-norm group).
• 18 years of age or older
• Able to commit to a minimum of 5 yearly evaluations
• Commitment of a companion to attend visits or complete surveys via mail
• Able to undergo a MRI
HORIZON StudyFor more information on the HORIZON study click here • Have clinical features of HD and a CAG polyglutamine repeat expansion ≥ 36• Have cognitive impairment as noted by the following:

1. A screening MMSE and a baseline (pre-dose) MMSE score between 10 and 26 (inclusive); and

2. A subjective assessment of cognitive impairment with decline from pre-HD levels by the Investigator after interviewing the subject and caregiver;

•  Are willing and able to give informed consent

•  Aged 30 years or older

• Have a caregiver who assists/spends time with the subject at least five days per week for at least three hours per day and has intimate knowledge of the subject’s cognitive, functional, and emotional states, and of the subject’s personal care.

HART StudyFor more information on the HART study click here • Able to provide written Informed Consent prior to any study related procedure, including consent to genotyping of the CYP2D6 gene.• Clinical features of HD, and a positive family history and/or the presence of ≥ 36 CAG repeats in the Huntington gene.

• Male or female age ≥ 30 years.

• Willing and able to take oral medication and to comply with the study specific procedures.

• Ambulatory, being able to travel to the assessment center, and judged by the Investigator as likely to be able to continue to travel for the duration of the study.

• Availability of a caregiver or family member to accompany the subject to two visits.

• A sum of ≥ 10 points on the mMS at the screening visit.

• For subjects taking allowed antidepressants or other psychotropic medication, the dosing of medication must have been kept constant for at least 6 weeks before enrollment.

Works Cited

F. Clum, C. Garnett, T. Wang and A, Lanctot, 2010


Induced Pluripotent Stem Cells: The Future of Tissue Generation

As viable human brain tissue is not available for use in studying disease development and creating therapies for neurological disorders like Huntington’s disease (HD), researchers desperately needed an alternative cell source for this purpose.  Embryonic stem cells fit this role but have many disadvantages, especially for treatments, including immune rejection by the recipient. Some of these drawbacks have been overcome by a recent discovery that revolutionized the face of stem cell biology.  In 2006, Shinya Yamanaka’s research group at Kyoto University made a groundbreaking announcement: they had discovered that adult cells could be genetically engineered to revert back to apluripotent, stem cell-like state.  As iPSC (induced pluripotent stem cell) production rapidly improved, the cells were soon able to compete with traditional fetal, embryonic, and adult stem cells. The primary advantages of iPSCs compared to other stem cells are: a) iPSCs can be created from the tissue of the same patient that will receive the transplantation, thus avoiding immune rejection, and b) the lack of ethical implications because cells are harvested from a willing adult without harming them.  These patient-specific cells can be used to study diseases in vitro, to test drugs on a human model without endangering anyone, and to hopefully act as tissue replacement for diseased and damaged cells.

Like other stem cells, iPSCs have the ability to proliferate indefinitely in vitro, creating a theoretically unlimited source of cells. Like embryonic stem cells,  iPSCs can also differentiate into any cell of the body, regardless of the original tissue from which they are created.  Scientists have found how to direct the differentiation of pluripotent stem cells into many types of target tissue, including neural tissue.  iPSCs demonstrate that by the introduction of just four genes into somatic cells that normally cannot differentiate at all, cells can be created that can differentiate into every cell type in the body.  The early results of iPSC differentiation studies look promising. For example, human fibroblasts have been successfully turned into iPSCs that then are differentiated into insulin-producing cells, a result that holds much potential for the treatment of diabetes.  Mouse iPSCs have been differentiated into cardiovascular (heart muscle) cells, that actually show the contractile beating expected of heart tissue.

Although there are many problems that still must be addressed for iPS technology, such as the tendency for tumors to evolve after iPSC transplantation and the low efficiency of the technology, iPSCs could completely change how diseases are approached in biomedical research.  For HD and other neurological disorders, iPSCs could create perfect models for the cells of the central nervous system that are harmed in the diseases.

Stem cell biology is a very hot topic in modern medicine, yet much is still unknown about the mechanisms underlying pluripotency and differentiation. In order for safe, controllable, and efficient cellular reprogramming to be achieved, there must be more knowledge on the regulation of stem cell states and transitions. iPSCs show that specialized cells and tissue can be transformed into other types of cells, proving cells are much more flexible than previously thought.  As the study of HD will greatly benefit from this new, unlimited source of neural cells for research and cell therapy, iPSCs may be able to provide new and innovative treatments for HD.

The Discovery of iPSCs^

The creation of pluripotent cells has been widely studied for decades. In 1976, the first method of fusion of an adult somatic cell with embryonic cells to create pluripotent stem cells was reported. However, fusion with embryonic cells created unstable cells that were rejected by immune systems after transplantation. If the genes that induced pluripotency could be isolated from their parent embryos and injected into somatic cells, these problems could be avoided.

Yamanaka’s research team studied twenty-four genes expressed by embryonic stem cells in an effort to track down these essential genes that induce pluripotency. To detect pluripotency, they looked for cells expressing genes that were traditionally expressed only in embryos.  They discovered that the addition of four genes induced a cell into a pluripotent state capable of then becoming many different cell types.

The Four Factors^

Subsequent studies showed that other gene combinations were also successful in reengineering cells into iPSCs, but none were as efficient as the first four. Adding other genes that are expressed in early development was shown to increase reprogramming efficiency, and the specific genes needed varied depending on the cell type that was being forced back to its pluripotent state.  As the four factors and their alternatives were largely discovered by trial and error, it is not known how the genes induce pluripotency. Discovering how genes work may point to ways of improving the efficiency of the process and assessing the quality of iPSCs.

The specific genes that induce iPSCs tell scientists a lot about the characteristics of the cells themselves. Pluripotent stem cells are very closely related to tumor cells. Both can survive and proliferate indefinitely, and a test of pluripotency is whether a cell can create a tumor.  It is therefore no surprise that two tumor-related genes, c-Myc and Klf4, are needed to create iPSCs.  Another requirement of pluripotent stem cells is open and active chromatin structures (for more information on chromosomes, click here and DNA transcription click here). The c-Myc gene codes for proteins that loosen the chromatic structure, stimulating differentiation.  Klf4 impedes proliferation.  c-Myc and Klf4 in this way  regulate the balance between proliferation and differentiation.  If only c-Myc and Klf4 are used in the engineering of iPSCs, tumor cells will arise—instead of pluripotent stem cells. Oct3/4 and Sox2 are required to direct cell fate towards a more embryonic stem cells (ESC)-like phenotype.  Oct3/4 directs specific differentiation, such as neural and cardiac differentiation, while Sox2 maintains pluripotency.  Oct3/4 and Sox2 together ensure that iPSCs are indeed pluripotent stem cells and not tumor cells.

Amounts and Timing of Reprogramming Factors^

The programming of iPSCs depends both on the original cell type being transformed and the levels of each reprogramming factor that is expressed.  Expressing Oct3/4 more than the other genes increases efficiency.  Increasing the expression of any of the other three genes decreases the efficiency.  There is clearly a correlation between gene expression ratio and reprogramming efficiency, but the optimal ratio is likely to vary depending on the cell type being reprogrammed. For instance, when neural progrenitor cells are reprogrammed, they do not require Sox2 as they express this gene sufficiently already.  The level of expression of other important genes for maintaining pluripotency also can affect the reprogramming process and the quality of the resulting cells.

The effect gene expression ratio has on reprogramming may explain why efficiency is typically so low (less than 1% of cells are reprogrammed successfully).  Reprogramming is a slow process, and so the timing of various events may also exert a great influence over thecell’s success. The minimum time for the full reprogramming of a mouse somatic cell into an iPSC is between eight and twelve days. The timing of the mechanism for cellular reprogramming may also be a reason for low efficiency, as the cells can only proceed if the right molecular events happen in the correct order.

Creation of a Germline Competent Model^

In the first studies of iPSCs, the cells were shown to be similar to ESCs in morphology and proliferation.  But the cells were not germline-competent, in other words they were unable to differentiate into cells that expressed genes of the parent cells, and so they could not give rise to adult chimeras when transplanted into blastocysts.  As chimeras play key roles in biomedical research, scientists identified iPSCs through a stricter gene marker that only identified iPSCS that were germline competent. It was found that cells that expressed Nanog, a gene closely tied to pluripotency, were germline competent. These cells also were virtually indistinguishable from ESCs in gene expression, and were more stable.  The transgenes were better silenced in the Nanog identified cells although 20% of the iPSCs still developed tumors due to the reactivation of c-Myc.  Unfortunately this stricter criterion also decreased efficiency to only 0.001-0.03%. While subsequent studies improved this efficiency by varying methods, the fact remains that iPSCs are generated with incredibly low efficiency.

Characteristics of iPSCs^

iPSCs exhibit many characteristics that are related to their pluripotency. They lose proteins that are common to somatic cells and gain proteins common to embryonic cells. They also lose the G1 checkpoint in their cell cycle control mechanism, which embryonic stem cells lack as well. During the reprogramming of somatic cells in the iPS mechanism, the cell cycle structure of stem cells must be reestablished. Another distinguishing characteristic of pluripotent stem cells is their open chromatin structure, as this is needed to maintain pluripotency and to access genes rapidly for differentiation. iPSCs have the open chromatin structure associated with ESCs and other pluripotent cells.  Finally, female iPSCs show reactivation of the somatically silenced X chromosome.  A very early step of stem cell differentiation is the inactivation of one of the two X chromosomes in female mammals, a random process. By the reactivation of this X chromosome, iPSCs show that they are truly pluripotent and identical to ESCs.

Non-retroviral Methods of iPSC Production^

A huge barrier to the eventual use of iPSC-derived treatments is the use of retroviruses to force the expression of the four key genes, discussed above, and activating their transcription factors. Retroviruses can carry target DNA that is inserted into a host cell’s genome upon injection, making them ideal for incorporating the four genes into target cells. However, this DNA and the rest of the viruses’ genomes remain in the host genome, which can lead to transcription of unwanted genes and greatly increases the risk of tumors. The expression of the four transgenes must be silenced after reprogramming to avoid harmful gene expression.  c-Myc, a tumor-promoting gene, especially must be silenced after cellular reprogramming or the risk of tumor development becomes too great for clinical use. These retroviral methods in which the transgenes are still present in the pluripotent cells pose a danger to safety, and also are less closely related to ESCs in gene expression than their non-retroviral alternatives.  Methods of reprogramming iPSCs without transgene expression in the reprogrammed cell is therefore essential not only for potential therapies and clinical applications, but also for reliable and accurate in vitro models of diseases.  Yet, the low efficiency of alternatives remains a worry. Whether these methods will be viable for human clinical use remains to be seen.


The excision strategy (transient transfection) of iPSC generation allows the transgenes to briefly integrate into the genome but then removes them once reprogramming is achieved. An example of this site/enzyme combination is the loxP site and the Cre enzyme.  In a study of Parkinson’s disease (PD), specific iPSCs, this loxP/Cre combination was used to generate the iPSCs. Neural differentiation was then induced on the iPSCs to test whether they could differentiate into dopaminergic neurons, the cells harmed in PD.  The differentiation was successful, indicating the transgenes had been excised. However, a loxP site remains in iPSC genome as does some residual viral DNA, so there is still a small potential for insertional mutagenesis. The piggyBac site/enzyme system on the other hand is capable of excising itself completely, not leaving any remnants of external DNA in the iPSC genome.  The piggyBac system also was much more efficient than other non-retroviral methods, with comparable efficiency to retroviral methods, but with the added benefits of safety and ease of application.

Adenoviral Methods^

Adenoviral methods do not pose the same threats as retroviral methods of generating iPSCs. Adenoviruses work like all viruses by hijacking their hosts’ cellular machinery to replicate their own genome and reproduce, but unlike retroviruses they do not incorporate their genome into the host DNA. Because the transgenes are never even incorporated into the host’s genome they do not have to be excised. Instead, the genes are expressed directly from the virusgenome. iPSCs created by adenoviral methods demonstrated pluripotency, but have extremely low reprogramming efficiency.   Viralgenomic material could not be detected in any of the iPSCs, and no tumor formation was reported. This suggests that the use of non-integrating adenoviral methods substantially lowers the threat of tumorgenesis. The successful creation of iPSCs from adenoviral methods proves definitively that safer, non-retroviral methods can also successfully reengineer cells.

Non-DNA methods of iPSC generation^

Recent studies have implied that perhaps genetic material is not required for iPS cellular reprogramming.  The substitution of transgenes with small molecules that promote iPSC generation would be a safe, clinically appropriate way of creating iPSCs, though it remains to be seen if small molecules will be able to completelyreplace genetic methods of iPSC generation or are just useful as supplementary aids to the process.  Protein transduction is a different method shown to entirely replace gene delivery. In this method fusion proteins are created, which fuse each of the transgenes to a cell-penetrating peptide sequence that allows it to cross the cellular membrane. Reprogramming without DNA intermediates should eliminate the risk of tumorgenesis and distorted gene expression due to the reactivation of the transgenes.

Issues Facing the Use of iPSCs^

With iPSC research being a hotspot for several years now, many of the problems the technology first faced have been studied and resolved.  iPSCs are now germline competent,  can be generated from many different types of human and animal somatic tissue, and can be generated in a  variety of retrovirus-free methods. This lack of retroviruses ended worries about transgene reactivation and subsequent tumorgenesis. The nature of the transgenes in question made the risk of tumor development particularly prevalent, as two of the genes, c-Myc and Klf4, directly inducing tumorgenesis.  Retroviral delivery posed a threat to safety in its increased risk of tumorgenesis and in its tendency to alter gene expression. When other methods were established that did not require retroviruses, these concerns were put to rest, yet these new methods’ efficiencies must be improved and some issues still remain concerning the safety of iPSCs and their abilities to act on par with any other pluripotent cell.


Even without the use of retrovirsues, tumorgenesis is still a large concern for iPSCs, especially if they are ever to be used as cell replacement therapies. Using retroviral methods, twenty percent of iPSCs developed tumors in one study, and though this number has significantly lowered, it must become negligible for iPSCs to be considered for clinical use.  It is telling that the assay for pluripotency in stem cells is the ability to form teratomas, or tumors. This test of “stemness” illustrates the precariously close link between stem cells and tumor cells.  There are several proposals on how to prevent this tumor formation.  The idea to sort cells before transplantation and after differentiation, so that only well-differentiated neural progenitors will be transplanted, is one such proposal.  Another proposal is to genetically modify iPSCs so that they will have a suicide gene to self-destruct when tumors are created. Finally, some antioxidants, such as Resveratrol, have been shown to have tumor-suppressing qualities, and could potentially aid in any treatment proposed to prevent tumors (for an article about the potential of Resveratrol for the treatment of HD, click here).


Directed differentiation has been a perennial problem in stem cell biology, and iPSCs bring their own unique characteristics to the dilemma.  As with ESCs, iPSCs sometimes have the tendency to not fully differentiate. Also, as with all stem cell research with neurodegenerative diseases, a more efficient and comprehensive method to differentiate cells into neural progenitors and specific neuronal tissue must be discovered, as current methods are imperfect and slow.

Quality Assessment and Variability^

In iPSC research there is a need to establish methods to evaluate the reprogramming process and the final quality of the cells.  To create human iPSCs suitable for cell replacement therapies, there must be tests to ensure that all pluripotent cells have differentiated, and that the cells have not been genetically altered during reprogramming or during differentiation.  With cells derived from diseased individuals for an autologous treatment, there is naturally the concern that the underlying genetic cause of the disease remains in the iPSCs and will manifest itself in the same way. Some studies have indicated that iPSC lines differ drastically, which makes the reproducibility of any particular phenotype difficult.  Analyzing this variability may help discover which somatic tissue is best for generating iPSCs.


A problem that has not been significantly improved upon since the beginnings of iPSC research is the technology’s low efficiency. Some hypothesize that the addition of other factors would greatly aid the reprogramming process, and that reprogramming success depends on specific amounts and ratios of the four factors, which are only achieved by chance in a small percentage of the cells. Modifying the culture conditions is another area of study for increasing efficiency and rate of iPSC production. For cellular transplantation therapies, other questions must also be considered, such as the optimal cell dose and source tissue, and the best way to deliver the cells. There are potential solutions to this problem, though. Induction efficiencies have been improved up to a hundred times initial values by use of different somatic starting cells and the aid of small molecules. Although there are barriers to iPSC production, research in this field is still in its infancy and has made impressive gains for the short time it has been going on. As more studies are conducted on iPSCs, these issues may be resolved and iPSCs may enter a state capable of clinical use.

Origin Cells Used^

Another potential way to improve iPSC generation efficiency is to establish the best somatic cells type to reprogram for the cleanest, easiest reprogramming. Many different tissue types have been reprogrammed, including fibroblasts, neural progenitor cell, and stomach epithelial (stomach lining) cells. Certain cell types are much more efficient and rapid than others. There is also the probability that subtly varying iPSCs are generated from different types of starting tissue, some of which may prove to be useful for research or replacement purposes.

An interesting “type” of somatic cell was used in studies of secondary iPSCs.  iPSCS were initially generated and then implanted into blastocysts to create chimeric animals. Somatic cells from these chimeras were then removed and iPSCs were generated from these cells, creating secondary iPSCs.  These secondary iPSCs were generated more efficiently. The differentiation status of thecells to be reprogrammed also affects efficiency, as adult progenitor cells are reprogrammed at three hundred times the efficiency of completely differentiated somatic cells.


An interesting possibility for the reprogramming methods of iPSCs is the potential for transdifferentiation. It may not always be necessary to reprogram cells all the way back to their most primitive pluripotent stem cell state, and instead reprogram one type of adult somatic tissue directly into a different type, bypassing the lengthy processes of complete reprogramming and subsequent differentiation.  For example, in theory fibroblasts that can be easily and safely obtained from a patient’s skin could be converted into neurons or heart muscle cells without ever passing through a pluripotent stage. This would have advantages not only in the conservation of time and resources but also for safety, as transdifferentiation does not pose the risk of tumorgenesis as the cells never are pluripotent. Unfortunately, the technology for such processes is very difficult. To reprogram cells directly into a different cell type, the qualities and characteristics of the desired cell type must be comprehensively understood. For iPSCs the desired cell type was embryonic stem cells, which were very well researched and characterized, but for many types of cell of interest, including cells of the central nervous system, there are still many unanswered questions about the target cell population. Excitingly, the Wernig lab at Stanford has recently created induced neurons (iN) directly from mouse fibroblasts.

Paramedic functions^

A potential use of iPSCs for cellular therapy that can be applied much more quickly than actual replacement of damaged tissue is the transplant of pluripotent cells as support cells rather than replacement neurons. These cells offer neuroprotection by preventing inflammation and producing neurotrophic factors (for the therapeutic use of neurotrophic factors in HD, click here).  In various studies, the transplantation of iPSCs has significantly improved host neuronal survival and function.  This bystander mechanism of therapy is of huge immediate potential in iPSCs, and Dr. Nolta’s lab recently submitted a request for a clinical study of the same mechanism using mesenchymal stem cells to the FDA. For a detailed study of the use of iPSCs for this purpose click here.


Stem cell biology has been an area of great interest and intense debate since its inception, and iPSC technology has furthered this research and created hope for potential therapeutic applications. While there are still many barriers to the clinical use of stem cells, iPSCs may help elucidate the nature of both pluripotent stem cells and of many disease pathologies to reach an eventual concrete connection between the two. With their potential for autologous cell replacement and disease modeling in vitro iPSCs are the future of stem cell research, and as such they are key players in the battle against HD.

For Further Reading^

Abeliovich, Asa and Claudia A. Doege. “Reprogramming Therapeutics: iPS Cell Prospects for Neurodegenerative Disease.” Neuron. 12 Feb, 2009, 61 (3): 337-39.

Short, approachable article reviewing two studies deriving iPSCs from patients with neurological disorders.

Cox, Jesse L. and Angie Rizzino. “Induced pluripotent stem cells: what lies beyond the paradigm shift.” Experimental Biology and Medicine. Feb 2010, 235 (2): 148-58.

Very detailed, mostly accessible review of the state of iPS research and the discoveries to date, as well as what iPS cells mean for stem cell biology and modern medical approaches. Perfect thorough introduction to iPS technology.

Crook, Jeremy Micah, and Nao Rei Kobayashi. “Human stem cells for modeling neurological disorders: Accelerating the drug discovery pipeline.” Journal of Cellular Biochemistry. 105 (6): 1361-66.

Accessible, interesting article that argues the greatest potential for iPSCs is to test potential drugs for neurological diseases in vitro and find problems early on in the drug development, saving time and resources.

Gunaseeli, I., et al. “Induced Pluripotent Stem Cells as a Model for Accelerated Patient- and Disease-specific Drug Discovery.” Current Medicinal Chemistry. 2010, 17: 759-766.

Readable review on the future of iPS cells, comparing them with other stem cells and elucidating their pontential drawbacks. Good summary of the landmark discoveries in iPS technology to date.

Haruhisa, Inoue. “Neurodegenerative disease-specific induced pluripotent stem cell research.” Experimental Cell Research. 2010.

General overview of use of iPS cells specific to neurological diseases for modeling diseases in vitro and eventually using as a cellular replacement therapy. Good, non-technical overview of the various potential pathways of iPS technology.

Hung, Chia-Wei, et al.  “Stem Cell-Based Neuroprotective and Neurorestorative Strategies.” International Journal of Molecular Science. 2010, 11(5): 2039–2055.

Overview of various neurological diseases and the potential of stem cell therapeutics, either using adult neural stem cells or iPS stem cells. Experiment descriptions are fairly technical, but the review’s reflections and discussion are accessible and interesting.

Laowtammathron, Chuti, et al. “Monkey hybrid stem cells develop cellular features of Huntington’s disease.” BioMed Center Cell Biology. 2010, 11 (12).

Detailed article on the establishment of pluripotent HD monkey model cell line and its use in the study of Huntington’s.

Marchetto, Maria C.N., et al. “Pluripotent stem cells in neurodegenerative and neurodevelopmental diseases.” Human Molecular Genetics. 2010, 19 (1).

Fairly technical review describing the use of iPSCs for modeling neurological disorders.

Niclis, J.C., et al. “Human embryonic stem cell models of Huntington’s Disease.” Reproductive Biomedicine Online. July 2009, 19 (1): 106-13.

Detailed, technical article on the use of human embryonic stem cell lines for HD.

O’Malley, James. “New strategies to generate induced pluripotent stem cells”. Current Opinions in Biotechnology. Oct. 2009: 20 (5): 516-21.

Longer technical article on the various strategies to generate iPS cells without using potentially dangerous viral vectors.

Okita, Keisuke, et al. “Generation of germline-competent induced pluripotent stem cells.” Nature. 19 Jul, 2007, 448(7151):313-17.

Fairly technical article about an early study in iPS research, where cells were selected for Nanog expression rather than the less pertinent gene Fbx15. This higher caliber of selected cells were germline-competent.

Okita, Keisuke, et al. Generation of Mouse Induced Pluripotent Stem Cells Without Viral Vectors.” Science. 7 Nov, 2008, 322 (5903): 949-53.

Technical article about the advancements in finding non-viral, clinically applicable methods of creating iPS cells.

Orlacchio, A., et al. “Stem Cells: An Overview of the Current Status of Therapies for Central and Peripheral Nervous System Diseases.” Current Medicinal Chemistry. 2010, 17: 595-608.

Technical review on the various types of stem cells used in the studies of neurological diseases and the progress made to date with these cells.

Park, In-Hyun, et al. “Disease-Specific Induced Pluripotent Stem Cells.” Cell. 2008, 134 (5): 877-86.

Fairly accessible article on the creation of iPS cells with genetic defects, as tools for studying the symptoms and experimenting with treatments of various diseases.

Robbins, Reisha D., et al. “Inducible pluripotent stem cells: not quite ready for prime time?” Current Opinion in Organ Transplantation. 15 (1): 61-57.

Clear review of the barriers facing clinical use of iPSCs, accessible and realistic.

Soldner, Frank, et al.Parkinson’s Disease Patient-Derived Induced Pluripotent Stem Cells Free of Viral Reprogramming Factors.” Cell. 6 Mar, 2009, 136 (5): 964-77.

Technical article about first successful derivation of iPS cells from a patient with a neurodegenerative disease without using viral vectors. Relevant to HD research as a protocol that will likely be followed for subsequent creation of neurodegenerative iPSC lines for in vitro study.

Stradtfeld, Matthias, et al. “Induced Pluripotent Stem Cells Generated Without Viral Integration.” Science. 7 Nov, 2008, 322 (5903): 945-49.

Technical article outlining a method for creating iPS cells using excisable adenoviruses, rather. than retroviruses that have the potential to harm the cells.

Takahashi, Kazutoshi, et al. Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors” Cell. 30 Nov, 2007, 131(5): 861-72.

Landmark article in the discovery of induced pluripotent stem cells and the factors that create them. Short, but fairly technical.

Yamanaka, Shinya. “Induction of pluripotent stem cells from mouse fibroblasts by four transcription factors.” Cell Proliferation. Feb, 2008, 41 (Suppl. 1):51-6

Short review, less technical summary of first iPS discovery by Yamanaka. Perfect for quick overview of the basics of iPS cell generation.

Yamanaka, Shinya. “Strategies and New Developments in the Generation of Patient-Specific Pluripotent Stem Cells.” Cell: Stem Cell. 7 June 2007, 1(1): 39-49.

Comprehensive review of various methods for creating pluripotent stem cells with a detailed introduction to iPSC methods. Fairly accessible, and very thorough.

A. Lanctot 2011


Adipose Derived Stem Cells and HD: Real Applications of iPSCs

While stem cells have always been heralded as the future of cellular replacement therapies, recent stem cell research has explored the potential “bystander” or “paramedic” effects of stem cells, which use stem cells to repair damaged cells rather than replacing them. Bystander therapies do not require the stem cells to become the type of cell that is damaged (to differentiate into this cell type),  but rather can help damaged neurons by changing the host environment. The phenotype of the undifferentiated, stem cell that are still pluripotent, able to differentiate into many different cell types, may provide therapeutic benefits in Huntington’s Disease (HD) by releasing neurotrophic factors that promote neuron growth and survival and arrest the mutant huntingtin protein’s negative influence on key cellular survival and energetic pathways. This paramedic function of stem cells might be harnessed to prevent mechanisms of HD that cause harmful symptoms rather than replacing damaged cells, as an alternative approach to traditional drug therapy for the treatment of neurodegenerative diseases.

Researchers in South Korea have recently found that adipose-derived stem cells (ASCs) can serve the same “paramedic” function in HD as is observed in the mesenchymal stem cells that Dr. Nolta is currently researching at the University of California, Davis. Similar to mesenchymal stem cells, adipose-derived stem cells do not create the ethical debates that embryonic stem cells do, as they are removed from adults during elective surgery, not from embryos in vitro. But in contrast to mesenchymal stem cells, ASCs are not found naturally in the body, but are rather multipotent stem cells created by iPS, or induced pluripotent stem cells (for more information about the new technology of iPS, click here). Derived from fat tissue taken from consenting patients, ASCs have the double advantage of easy access and minimal ethical implications. Like all stem cells, ASCs have the ability to differentiate into different somatic cells, though the mechanisms of differentiation are still unclear and scientists do not know how to direct differentiation of ASCs into certain types of tissue.  This study was not concerned with the differentiation of ASCs, but rather their neuroprotective abilities, such as the release of growth factors that are essential in combating many of the symptoms of HD.

How the Bystander Mechanism Works^

Current research using HD mouse models indicates that the use of fetal striatal tissue to replace the damaged striatal tissue in HD mouse models is not possible at this time, as the replacement tissue does not alter the toxicity of the mutant huntingtin protein.  This is like replacing a wall that has been eaten away by termites without doing anything to remove the termites: the new wall will soon also be harmed by the pests. Even if this barrier was overcome, the use of stem cells to replace damaged tissue is hampered by many other problems, such as a lack of donor tissue and rejection by the immune system, identical problems to organ replacement.  Rather than replacing damaged tissue, a different approach could use stem cells to preemptively prevent HD from harming neuronal tissue.  Scientists have noted that with other diseases, stem cells often act through a bystander mechanism, preventing the symptoms of the disease from manifesting, rather than directly replacing damaged cells. This novel approach to stem cell use does not require the extensive technology of differentiation, transplantation, and incorporation with host tissue that cell replacement needs. Instead, the bystander mechanism approach of stem cells takes advantage of the cells’ ability to release factors in their pluripotent state that combat the symptoms of HD. The researchers in this study wished to test this mechanism using ASCs, knowing that the cells could not only differentiate into cells such as neurons and glial cells that could be useful in combating HD, but could also release growth factors that may slow the symptoms of HD (to learn about growth factors and their role in fighting HD, click here.

HD symptoms may be caused by the alterations the mutant huntingtin protein makes to the transcription of DNA by interfering with transcription factors (for more information about the HD’s affect on transcription, click here) Transcriptional interference and mitochondrial dysfunction are key aspects of HD pathology. Mutant huntingtin protein aggregates impede important transcription factors such as the CREB-binding protein which is essential for transcription of pathways key to cell survival.  In HD, one way neural cell death is induced is by inhibiting this transcription factor.  ASCs could slow neurodegeneration and neural death by releasing neurotrophic factors that help prevent premature cell death. Neurotrophic factors encourage the maintenance, growth, and survival of neurons and so serve as a counterforce to mutant huntingtin’s stimulation of cell death. Another transcription factor, PGC-1, controls the creation of mitochondria, and when it is repressed by mutant huntingtin, reduced numbers of mitochondria causes the cell to receive insufficient energy. This makes the cell susceptible to glutamate, an excitatory neurotransmitter that can harm neurons and may be involved in many of the symptoms of HD. ASCs increase PGC-1 expression by preventing the huntingtin protein from impeding its transcriptional regulation, and so prevent glutamate levels from becoming harmfully high in the neuron.

Effects of ASCs in Three Different Models^

The effect of ASC transplantation was tested in three models, a knockout rat model, a transgenic mouse model and an in vitro cell model (for more information about different types of experimental models, click here). All models showed the positive effects of ASCs. The rat model showed that the ASCs were neuroprotective, reducing neuronal death. More specifically, this was observed by comparing the size of the rats’ ventricles of the brain, which are enlarged by the loss of neural tissue in HD. The ventricles of the ASC-treated rats were smaller than those of the control HD model rats, indicating less tissue loss. Ventricle volume is closely correlated with HD progression, as larger ventricles indicate greater loss of neural tissue. By showing a decreased ventricle volume, ASC rats seem to exhibit less cell death, resulting in less loss of neural tissue. In addition, compared to the HD control mice, the mouse model with ASC transplantation had a delayed decline in motor function, had less mutant huntingtin aggregates, and  lived longer.

Potential Problems with ASCs for Clinical Use^

The promising results of the three models hint at the potential of ASCs to delay the onset of HD symptoms, but there is much that still needs to be researched.  It was found that the transplanted ASCs were not evenly distributed across the striatum as expected, but rather most ASCs remained at the initial site of transplantation. This may have impeded the effectiveness of the ASCs and may even be harmful in long-term models. The proliferation of the ASCs was slow once transplanted, so the documented ability of stem cells to proliferate in actual organisms does not seem to apply to ASC transplantation in HD models.  This study did not test whether the same results could be achieved in model organisms that are longer lived. Whether ASCs would be effective in humans, for extended periods of time, has not yet been determined.  Furthermore, the ASCs must be prevented from differentiating into cells that would be harmful in the brain. Currently, the research for directed differentiation of pluripotent cells is rudimentary, and there is particular risk associated with the spontaneous differentiation of ASCs in vivo. The potential risk of the cells to differentiate into other tissue like heart or skin cells should be tested. Another problem of stem cells is that they may be rejected by the patient’s immune system, though this problem is greatly reduced with ASCs. In this study, human ASCs were successfully injected into rat and mouse models without the aid of immunosuppressants, which is encouraging. To further prevent the risk of rejection, the patient’s own adipose cells could be used to create the ASCs. But ASCs derived from patients with the mutant huntingtin protein have yet to be tested and it is possible that these cells may be damaged or not fully effective.


Researchers have shown that ASCs may have the potential to protect mechanisms of transcription and rescue degenerating neurons by combating the detrimental actions of huntingtin aggregates through the release of growth factors. This bystander mechanism is a novel approach to using stem cells, as they have been traditionally thought of as replacement cells for damaged tissue. The value of stem cells to replace damaged tissue with healthy, fully-differentiated replacement cells cannot be dismissed, especially with new iPS technology and the ability to engineer replacement cells from the patient who is to receive them, reducing the risk of immune rejection. Unfortunately, much more research must be done before stem cells will be used in clinical therapies for cellular replacement, but a more immediate potential for stem cells is in a paramedic capacity, where differentiation and incorporation with the host tissue is not required. By influencing key events in the pathogenesis of HD, ASCs may delay the onset of harmful symptoms. Current research has shown that ASC transplantation may allow for the expression of transcription pathways that HD suppresses, reduce the number of toxic huntingtin aggregates, and decrease the extent of neuron death in mouse models. Its therapeutic use still requires much more research and exploration, and then must make the leap from animal models to human trials, but ASCs have the potential to rescue degenerating neurons and prevent HD symptoms. This ability of stem cells to not only replace damaged tissue, but also prevent tissue damage, holds promise for the treatment of HD.

For Further Reading

Lee, et al. “Slowed Progression in Models of Huntington Disease by Adipose Stem Cell Transplantation.Annals of Neurology. 2009, 66(5): 671-681.

Technical but well-explained article on a specific study of the use iPS cells derived from adipose cells in three HD models: induced rat, transgenic mouse, and in vitro.

A. Lanctot 2011

While stem cells have always been heralded as the future of cellular replacement therapies, recent stem cell research has explored the potential “bystander” or “paramedic” effects of stem cells, which use stem cells to repair damaged cells rather than replacing them. Bystander therapies do not require the stem cells to become the type of cell that is damaged (to differentiate into this cell type), but rather can help damaged neurons by changing the host environment. The phenotype of the undifferentiated, stem cell that are still pluripotent, able to differentiate into many different cell types, may provide therapeutic benefits in Huntington’s Disease (HD) by releasing neurotrophic factors that promote neuron growth and survival and arrest the mutant huntingtin protein’s negative influence on key cellular survival and energetic pathways. This paramedic function of stem cells might be harnessed to prevent mechanisms of HD that cause harmful symptoms rather than replacing damaged cells, as an alternative approach to traditional drug therapy for the treatment of neurodegenerative diseases.

Researchers in South Korea have recently found that adipose-derived stem cells (ASCs) can serve the same “paramedic” function in HD as is observed in the mesenchymal stem cells that Dr. Nolta is currently researching at the University of California, Davis. Similar to mesenchymal stem cells, adipose-derived stem cells do not create the ethical debates that embryonic stem cells do, as they are removed from adults during elective surgery, not from embryos in vitro. But in contrast to mesenchymal stem cells, ASCs are not found naturally in the body, but are rather multipotent stem cells created by iPS, or induced pluripotent stem cells (for more information about the new technology of iPS, click Derived from fat tissue taken from consenting patients, ASCs have the double advantage of easy access and minimal ethical implications. Like all stem cells, ASCs have the ability to differentiate into different somatic cells, though the mechanisms of differentiation are still unclear and scientists do not know how to direct differentiation of ASCs into certain types of tissue. This study was not concerned with the differentiation of ASCs, but rather their neuroprotective abilities, such as the release of growth factors that are essential in combating many of the symptoms of HD.

How the Bystander Mechanism Works

Current research using HD mouse models indicates that the use of fetal striatal tissue to replace the damaged striatal tissue in HD mouse models is not possible at this time, as the replacement tissue does not alter the toxicity of the mutant huntingtin protein. This is like replacing a wall that has been eaten away by termites without doing anything to remove the termites: the new wall will soon also be harmed by the pests. Even if this barrier was overcome, the use of stem cells to replace damaged tissue is hampered by many other problems, such as a lack of donor tissue and rejection by the immune system, identical problems to organ replacement. Rather than replacing damaged tissue, a different approach could use stem cells to preemptively prevent HD from harming neuronal tissue. Scientists have noted that with other diseases, stem cells often act through a bystander mechanism, preventing the symptoms of the disease from manifesting, rather than directly replacing damaged cells. This novel approach to stem cell use does not require the extensive technology of differentiation, transplantation, and incorporation with host tissue that cell replacement needs. Instead, the bystander mechanism approach of stem cells takes advantage of the cells’ ability to release factors in their pluripotent state that combat the symptoms of HD. The researchers in this study wished to test this mechanism using ASCs, knowing that the cells could not only differentiate into cells such as neurons and glial cells that could be useful in combating HD, but could also release growth factors that may slow the symptoms of HD (to learn about growth factors and their role in fighting HD, click (|here).

HD symptoms may be caused by the alterations the mutant huntingtin protein makes to the transcription of DNA by interfering with transcription factors (for more information about the HD’s affect on transcription, click |here) Transcriptional interference and mitochondrial dysfunction are key aspects of HD pathology. Mutant huntingtin protein aggregates impede important transcription factors such as the CREB-binding protein which is essential for transcription of pathways key to cell survival. In HD, one way neural cell death is induced is by inhibiting this transcription factor. ASCs could slow neurodegeneration and neural death by releasing neurotrophic factors that help prevent premature cell death. Neurotrophic factors encourage the maintenance, growth, and survival of neurons and so serve as a counterforce to mutant huntingtin’s stimulation of cell death. Another transcription factor, PGC-1, controls the creation of mitochondria, and when it is repressed by mutant huntingtin, reduced numbers of mitochondria causes the cell to receive insufficient energy. This makes the cell susceptible to glutamate, an excitatory neurotransmitter that can harm neurons and may be involved in many of the symptoms of HD. ASCs increase PGC-1 expression by preventing the huntingtin protein from impeding its transcriptional regulation, and so prevent glutamate levels from becoming harmfully high in the neuron.

Effects of ASCs in Three Different Models

The effect of ASC transplantation was tested in three models, a knockout rat model, a transgenic mouse model and an in vitro cell model (for more information about different types of experimental models, click|here). All models showed the positive effects of ASCs. The rat model showed that the ASCs were neuroprotective, reducing neuronal death. More specifically, this was observed by comparing the size of the rats’ ventricles of the brain, which are enlarged by the loss of neural tissue in HD. The ventricles of the ASC-treated rats were smaller than those of the control HD model rats, indicating less tissue loss. Ventricle volume is closely correlated with HD progression, as larger ventricles indicate greater loss of neural tissue. By showing a decreased ventricle volume, ASC rats seem to exhibit less cell death, resulting in less loss of neural tissue. In addition, compared to the HD control mice, the mouse model with ASC transplantation had a delayed decline in motor function, had less mutant huntingtin aggregates, and lived longer.

Potential Problems with ASCs for Clinical Use

The promising results of the three models hint at the potential of ASCs to delay the onset of HD symptoms, but there is much that still needs to be researched. It was found that the transplanted ASCs were not evenly distributed across the striatum as expected, but rather most ASCs remained at the initial site of transplantation. This may have impeded the effectiveness of the ASCs and may even be harmful in long-term models. The proliferation of the ASCs was slow once transplanted, so the documented ability of stem cells to proliferate in actual organisms does not seem to apply to ASC transplantation in HD models. This study did not test whether the same results could be achieved in model organisms that are longer lived. Whether ASCs would be effective in humans, for extended periods of time, has not yet been determined. Furthermore, the ASCs must be prevented from differentiating into cells that would be harmful in the brain. Currently, the research for directed differentiation of pluripotent cells is rudimentary, and there is particular risk associated with the spontaneous differentiation of ASCs in vivo. The potential risk of the cells to differentiate into other tissue like heart or skin cells should be tested. Another problem of stem cells is that they may be rejected by the patient’s immune system, though this problem is greatly reduced with ASCs. In this study, human ASCs were successfully injected into rat and mouse models without the aid of immunosuppressants, which is encouraging. To further prevent the risk of rejection, the patient’s own adipose cells could be used to create the ASCs. But ASCs derived from patients with the mutant huntingtin protein have yet to be tested and it is possible that these cells may be damaged or not fully effective.


Researchers have shown that ASCs may have the potential to protect mechanisms of transcription and rescue degenerating neurons by combating the detrimental actions of huntingtin aggregates through the release of growth factors. This bystander mechanism is a novel approach to using stem cells, as they have been traditionally thought of as replacement cells for damaged tissue. The value of stem cells to replace damaged tissue with healthy, fully-differentiated replacement cells cannot be dismissed, especially with new iPS technology and the ability to engineer replacement cells from the patient who is to receive them, reducing the risk of immune rejection. Unfortunately, much more research must be done before stem cells will be used in clinical therapies for cellular replacement, but a more immediate potential for stem cells is in a paramedic capacity, where differentiation and incorporation with the host tissue is not required. By influencing key events in the pathogenesis of HD, ASCs may delay the onset of harmful symptoms. Current research has shown that ASC transplantation may allow for the expression of transcription pathways that HD suppresses, reduce the number of toxic huntingtin aggregates, and decrease the extent of neuron death in mouse models. Its therapeutic use still requires much more research and exploration, and then must make the leap from animal models to human trials, but ASCs have the potential to rescue degenerating neurons and prevent HD symptoms. This ability of stem cells to not only replace damaged tissue, but also prevent tissue damage, holds promise for the treatment of HD.


Induced Neurons

Despite predictions of transdifferentiation being a technology of the future, Dr. Marius Wernig’s lab at Stanford has recently discovered a method of reengineering neurons directly from fibroblasts by the the forced expression of transgenes. This is the same method by which induced pluriptent stem cells (iPSCs) are produced, and transdifferentiation, the engineering of cells so they change their type without preceding through an intermediate stage, has been suggested as the logical progression from iPSCs. Its success suggests that, along with iPSC technology, cellular differentiation and cell fate are far more flexible than previously thought.  The new discovery that somatic cells can be turned into completely different cell types by a cocktail of a few genes revolutionizes previous thought about the unchangeable nature of fully differentiated cells, and adds new theories as to how cell fate is determined. The research also importantly shows that reengineering cells with transcription factors can create the complex structures and functions of somatic cells, not simply undifferentiated stem cells.  A combination of three transcription factors that are specifically found in the brain were shown to be able to convert both embryonic and postnatal fibroblasts directly into neurons, in much the same way iPSCs are created, but without preceding through any pluripotent stem cell intermediate. The lab tested nineteen potential genes to see what combination could efficiently convert mouse fibroblasts into neurons in vitro. The gene Asc11 alone could generate immature neurons with undeveloped properties. Another eighteen genes were then tested in combination with Ascl1, and five genes (Brn2, Brn4, Myt1l, Zic1, and Olig2) substantially improved neuron development. While none of these five genes generated induced neuronal (iN) cells when tested individually, it was found that the three factor combination of Ascl1, Brn2, and Myt1l produced the highest quantity of neurons with the most mature action potentials. As a result this combination was concluded to be the ideal grouping of factors for inducing neuron phenotypes.

Induced Neurons: Are they just like normal neurons?^

A difficulty in evaluating whether transdifferentation is successful is that unlike the relatively simple phenotypes of undifferentiated cells, somatic cells have far too many characteristics that must be tested to ensure that complete reengineering occurred. Induced neuronal cells (iNs) express neuron-specific proteins, generate action potentials and form functional synapses but it is still not known how, if at all, the iNs differ from normal neurons. The cells expressed three neuron marker proteins, MAP2, NeuN, and synapsin, and produced spontaneous action potentials, which are the distribution of charged ions across a neuron membrane that causes a signal to be propelled down the neuron’s axon and to the next neuron. Action potentials promote neural communication as they allow a message to cross a cell before neurotransmitters become involved in its movement between two cells. These action potentials were blocked when a sodium ion inhibitor, tetradotoxin, was introduced, as would occur in normal brain tissue. In normal brains, the concentration of sodium across the membrane of the neuron is essential for the cells to generate action potential and relay electrochemical messages through the neuron and onto the connecting cells. The cells’ expression of functional membrane channel proteins that allow sodium ions to flow in and out of the neuron supports the claim that iN cells and normal neurons exhibit identical membrane properties.

Problems in iN Generation^

The iN cells had various phenotypes, but did not exhibit all types of nervous tissue. Most iNs were excitatory neurons, while almost none contained periperin, a protein characteristic of neurons in the peripheral nervous system. The majority of iN cells were excitatory cells that expressed markers indicative of cortical identity, i.e. specific to cells typically found in the cortex. Further research may focus on the generation of iN cells of other specific neuronal subtypes, not to mention the generation of iNs from human cells (currently only mouse iNs have been successfully engineered).  Additional neural transcription factors may aid in creating neurons of more specific phenotypes.

Like induced pluripotent stem cells, iNs go through a gradual process of reengineering. While immature neuron-like cells can been seen as early as three days after infection, it takes five days for branching neuronal cells to form. Physical maturation continues over several weeks.  The efficiency of converting cells into iNs ranged from 1.8–7.7%, which is substantially better than efficiency of iPSC production. However, iNs cannot proliferate like iPSCs, so creating a larger quantity of cells is crucial.

An important question concerning iNs involves their ability to function as neurons by forming functional communication with other cells. This is a crucial requirement if they are to be used as tissue replacement in the future. The researchers tested whether iN cells have the capacity to form functional synapses with other iN cells and whether iN cells were capable of integrating into preexisting neural networks. When iN cells were grown with actual neurons, spontaneous and rhythmic neural activity was observed. The cells could receive synaptic inputs from the normal neurons, demonstrating their ability to integrate into preexisting tissue. It was also shown that iN cells are capable of forming functional synapses with each other.

Transgenes that Engineer Neurons^

Like iPSCs, the gene combination for creating iN cells allows some leniency and variation, but a certain “recipe” seems the most effective. It remains to be seen if different genes or different ratios of the genes that Dr. Wernig’s lab identified will even further improve efficiency of iN production. While Ascl1 alone is sufficient to induce some neuronal traits, such as expression of proteins that generate action potentials, the addition of Brn2 and Myt1l creates more mature cells with increased efficiency up to 19.5%. The highly efficient production of iNs makes it unlikely that they are merely formed from rare stem or precursor cells in the starting cell population, as great care was taken to exclude neural tissue in the isolation of the initial cell population, and no neurons or neural progenitor cells were detected in the culture. Future studies are nonetheless needed to unequivocally demonstrate that cells that have their own unique morphologies can be directly converted into neurons, and that the iNs are not mere derivatives of stem cells.

Why certain neuronal subpopulations (such as the cortical neurons) are more favored than others is another aspect of iN technology that remains to be researched. It may be that high expression of neural cell-fate determining factors directs certain cell types to form, so they are reengineered more often. Different cell types are produced during development by sets of transcription factors that cause cell type specific proteins to be produced. Each cell can be thought of as a person who walks by a lot of doors, but only has one key (specific transcription factors) that allows him to open one door, i.e. the cell can only become the type specified by its transcription factors.

iN cells are a possible alternative to iPSCs for generating patient-specific neurons. The generation of iN cells is fast, efficient, and has the major advantage over iPSCs that it does not go through a stage of pluripotent stem cells that are susceptible to tumor production.  The iN cells could also provide new methods for studying cellular identity and neural development. They have potential uses in neurological disease-modeling, drug discovery, and regenerative medicine. Formerly, transdifferentiation was never thought of as anything besides a futuristic version of cell engineering that would take many years to accomplish. The Wernig lab has shown that cells with more complex morphologies can indeed be generated directly from other cell types using much the same method as iPSCs, and although much remains to be tested, this new technology may revolutionize cell therapies as more cell types are derived.

For Further Reading^

Vierbuchen, et al. “Direct conversion of fibroblasts to functional neurons by defined factors.” Nature. 25 Feb. 2010, 463 (7284):1035-1041.

Well written, fairly accessible article. Some parts a bit technical but very nice section on the next steps for iN.

A. Lanctot 2011.


Mesenchymal Stem Cells

Mesenchymal stem cells (MSCs) are a type of multipotent stem cell, meaning that they can give rise to many but not all types of cells in the body. MSCs secrete substances, including cytokines and growth factors, that are essential to cell growth and help repair damaged tissue. Researchers are still exploring the functions of human MSCs in the body, but current knowledge about the stem cells suggests that they play an important role in cell repair, acting as a sort of “cellular paramedic.”


Mesenchymal Stem Cells: The Cellular Paramedic^

MSCs can be thought of as “cellular paramedics,” helping to restore damaged cells and tissue. As mentioned before, MSCs are able to secrete substances like cytokines and growth factors that can promote tissue repair. MSCs have even been shown to transfer products as large as mitochondria to damaged cells that need help. Specifically, MSCs stimulate angiogenesis, the process of new blood vessel formation, which has been linked to neurogenesis, the process by which new nerve cells are produced. The factors secreted by MSCs also reduce the harmful effects of oxidative damage and apoptosis.

When researchers discovered the “paramedic” quality of MSCs, they conducted several experiments to see how MSCs could potentially be used to help treat human diseases. In one experiment, MSCs obtained from humans were injected into mice that had some type of tissue damage and did not have a functional immune system. The MSCs were labeled so that scientists could track where they migrated after being injected into the mice. The researchers observed that the cells migrated throughout the damaged tissues apparently evenly and continued to be present in the tissue for a substantial period of time. The continued presence of MSCs is important to therapeutic development because it indicates that potential positive long-term effects of a treatment might be capable of persisting.

In additional experiments, scientists found that MSCs function differently in chronic disease models than in more temporary conditions like injury and trauma. In mouse models of acute injury, injected MSCs responded by helping to repair the tissue but were not present in the tissue for a substantial period of time, as they were in mouse models of chronic disease. For more information on mouse models, click here. When the experiment was repeated in injured mice without functional immune systems, the MSCs were again only temporarily present in the tissue. This suggests that temporary presence of MSCs is not a result of the host immune system. It is important to establish what types of environment foster the sustained presence of MSCs in the tissue so that clinicians can increase the effectiveness of future treatments using MSCs.

Research Using Mesenchymal Stem Cells^

The unique ability of MSCs to secrete their own growth factors enables scientists to culture them in a laboratory with relatively little maintenance. Furthermore, MSCs multiply rapidly in cell cultures. As a result, compared to other cells, it is easier to grow many MSCs after obtaining a limited number of the cells from a patient. Cells that are grown in vitro often develop different characteristics or stop multiplying after a period of time. However, MSCs have been found to maintain their characteristics and the ability to multiply even after many cycles of replication. Most other cells require expensive cytokines and growth factors to grow in vitro, so the low maintenance of MSCs increases their appeal as a source of stem cells to investigate potential treatments.

In the body, MSCs are found in the bone marrow, umbilical cord tissue, and fat pads.  MSCs are relatively rare in the bone marrow, comprising only 1 out of every 10,000 cells. On the other hand, umbilical cord blood and fat are rich sources of MSCs. Human MSCs can be harvested with minimal patient discomfort by tapping into an individual’s marrow space or fat pads.

Unlike most other cells, MSCs can be transferred between organisms with little immune rejection, in which the immune system of the organism receiving the transplant attacks the foreign tissue being transplanted. Scientists have found that MSCs suppress the immune system and reduce inflammation, making them good candidates for transplantation or injection into a host because they can avoid rejection by the host’s immune system.

The ease with which MSCs can be obtained, cultured, and transferred into a host without immune rejection is one reason why researchers are hopeful that MSCs may offer a promising way for scientists to develop treatments for neurodegeneration.

MSCs and the Brain^

In the brain, MSCs can help repair neurodegeneration by providing neurotrophic factors, proteins in the nervous system that promote the growth of nerve cells. For example, detailed experiments have shown that human MSCs express the neurotrophic factor BDNF (brain-derived neurotrophic factor) but do not express certain types of neurotrophins. You can read more about BDNF by visiting the HOPES article here. Interestingly, MSCs still exhibited their “cellular paramedic” effects when BDNF activity is blocked by an antibody, suggesting that MSCs secrete factors other than BDNF that help with cellular growth in the brain.  The effects of these factors allow nerve cells to carry out several processes that support survival: axon extension, growth, and cell attachment. In essence, MSCs change the tissue environment to enhance cell growth and regeneration in the brain.

Experiments done on mice with nerve cell injury have shown that MSCs injected into the brain promote recovery by secreting neurotrophic factors that facilitate nerve cell survival and regeneration. More relevant to HD are animal experiments showing that MSCs have the potential to repair striatal degeneration.

Bantubungi et al conducted experiments using rat MSCs to help treat rat models of HD with parts of their striatum removed. The research showed that MSCs proliferated more rapidly in the rat brains with striatal lesions than in healthy rat brains, suggesting that MSCs selectively respond to areas needing repair.  Furthermore, the scientists identified a protein called stem cell factor that encouraged proliferation and directed migration of the MSCs to damaged tissue. Stem cell factor is a naturally occurring protein that plays an important role in communication between cells. The experiment by the scientists suggests that MSCs do in fact play an important role in the brain and have potential to become a cellular therapy for neural repair.

Amin et al. conducted an experiment in which rat models of Huntington’s disease with damage to one side of the brain responded positively to MSC implantation into the brain. Specifically, damage within the striatum, the region of the brain drastically affected in HD, was significantly reduced in rats that received MSC implantations.

It is important to note that although MSCs promote cell growth and repair in the brain, scientists have not yet confirmed that MSCs can become mature nerve cells with the ability to signal, or communicate with, other nerve cells. Other types of stem cells, such as neural stem cells have been found to generate mature nerve cells. MSCs may not be able to become mature nerve cells themselves.

Genetically Engineered MSCs^

In addition to exploiting the natural ability of MSCs to help repair damaged nerve cells, scientists have found ways to genetically engineer MSCs to enhance their reparative properties in the brain. Scientists can introduce genes into MSCs that cause them to produce a greater quantity of factors such as cytokines and neurotrophins. Even after genetically engineered MSCs are allowed to multiply through several generations, they retain these genetic characteristics that boost production of helpful factors.  Furthermore, MSCs have proven to be robust cells: genetic engineering does not hinder the cells’ ability to multiply or grow. It is important however to continue these types of studies to ensure there are no unintended side effects of enhanced neutrophin and cytokine productions in MSCs or other cells and tissues in the body.

An experiment conducted by Dey et al. showed that mouse models of HD responded positively to treatment by MSCs. When the MSCs were genetically engineered to produce greater quantities of BDNF, the delay in disease progression was even more drastic in the mice.

The potential to genetically engineer MSCs to deliver factors such as BDNF is important because directly injecting some of these factors is not effective. Transplanted MSCs, as indicated in the studies mentioned above, have been shown to disperse throughout damaged tissue for a sustained period of time. The characteristics of the compounds themselves often prevent them from having a sustained physiological effect on their own. Therefore, the genetically engineered MSCs serve as a vehicle to enable effective delivery of helpful factors into the brain.

Another exciting possibility is to have MSCs themselves become vehicles for delivering genetic material that can help with diseases like HD. Dr. Jan Nolta’s research group at University of California Davis, for example, hopes to have MSCs deliver molecules for RNA interference, a type of gene therapy, into the cells of HD patients. You can read more about RNAi in the HOPES article here. This area of research is still in its preliminary stages and may take several years to obtain approval from government agencies such as the Food and Drug Administration (FDA). Nevertheless, it holds promise as a potential future treatment for HD.

A Potential Treatment for HD Using MSCs^

Future cellular therapies using MSCs would involve delivering MSCs into the brain, which has been approached in a number of different ways. Scientists have proposed delivering MSCs through an injection directly into the brain, an injection into the space surrounding the spinal cord, or a route through the nose (e.g. a nasal spray).

Although clinical trials using MSCs in humans have not yet been approved in the United States, one human cellular therapy trial has been conducted in France. In the trial, neural stem cells rather than MSCs were used. Five patients with HD received transplants from human fetal neural stem cells. After two years, three out of the five patients demonstrated motor and cognitive improvements. While this experiment provides hopeful evidence that stem cell therapies may provide a treatment for HD, the results should be interpreted with caution. First, two of the patients did not show significant improvements. Second, as noted before, neural stem cells and MSCs have different characteristics. Therefore, the results from this experiment do not indicate whether MSCs would provide an effective treatment. Finally, after four to six years, the patients showed clinical decline once again, suggesting that additional research is required before an effective long-term treatment is developed.

In addition to showing that stem cells are in fact an effective treatment for HD, researchers must also show that implanting MSCs into the brain is a safe procedure before treatments can continue to be developed. One of the main concerns with MSCs is that they could cause abnormal cell growth. Abnormal growth could result in extra bone or tumor formation. In particular, MSCs have been found to migrate to areas in the body that contain tumors. This could be dangerous if the MSCs excrete factors that encourage angiogenesis, cell growth, and cell proliferation within the tumor. For safety reasons, proposed clinical trials for cellular therapies exclude anyone who has had a brain tumor or other cancer within the past 5 years. Before treatment, an MRI will be administered to ensure the absence of any brain tumors.

Extensive biological safety trials have been conducted with MSCs by Dr. Gerhard Bauer and Dr. Jan Nolta at University of California at Davis. They have performed numerous experiments over the past decade on different animal models including mice, rats, and primates, to test if MSCs can be safely injected or grafted without tumorous growths.  Additionally, a successful clinical trial in France with five HD patients suggests that transplantation of stem cells into the brain can be done without negative health consequences. However, more evidence for the biological safety of injecting MSCs into the brain is needed to meet the rigorous safety standards of the FDA in the United States

MSCs have potential to be a safe and effective therapy for HD. While there is promising evidence from animal research that MSCs can slow neurodegeneration, specifically in the striatum, there are still many aspects of the potential therapy that require additional experimentation.

Further reading:^

1.     Aggarwal, S. and M. F. Pittenger (2005). “Human mesenchymal stem cells modulate allogeneic immune cell responses.” Blood 105(4): 1815-1822.

A technical paper that discusses how MSCs interact with the immune system.

2.     Aizman, I., C. C. Tate, et al. (2009). “Extracellular matrix produced by bone marrow stromal cells and by their derivative, SB623 cells, supports neural cell growth.” J Neurosci Res 87(14): 3198-3206.

A technical paper that talks about the various compounds secreted by MSCs

3.     Amin, E. M., B. A. Reza, et al. (2008). “Microanatomical evidences for potential of mesenchymal stem cells in amelioration of striatal degeneration.” Neurol Res 30(10): 1086-1090.

This paper discusses how MSCs might help counter nerve cell damage in the striatum, and is difficult

4.     Bachoud-Levi, A. C., V. Gaura, et al. (2006). “Effect of fetal neural transplants in patients with Huntington’s disease 6 years after surgery: a long-term follow-up study.” Lancet Neurol 5(4): 303-309.

This is the study in which human fetal neural stem cells were transplanted into HD patients

5.     Bantubungi, K., D. Blum, et al. (2008). “Stem cell factor and mesenchymal and neural stem cell transplantation in a rat model of Huntington’s disease.” Mol Cell Neurosci 37(3): 454-470.

A technical paper that discusses transplantation of MSCs in rats

6.     Crigler, L., R. C. Robey, et al. (2006). “Human mesenchymal stem cell subpopulations express a variety of neuro-regulatory molecules and promote neuronal cell survival and neuritogenesis.”

This paper discusses some of the compounds that MSCs secrete that play a role in nerve cell health, and is quite difficult

7.     Danielyan, L., R. Schafer, et al. (2009). “Intranasal delivery of cells to the brain.” Eur J Cell Biol 88(6): 315-324.

This paper discusses one of the several ways MSCs might be delivered through the brain.

8.     Dey, N. D., M. C. Bombard, et al. (2010). “Genetically engineered mesenchymal stem cells reduce behavioral deficits in the YAC 128 mouse model of Huntington’s disease.” Behav Brain Res 214(2): 193-200.

This technical paper discusses how MSCs helped behavior problems in a mouse model of HD.

9.     Joyce, N., G. Annett, et al. (2010). “Mesenchymal stem cells for the treatment of neurodegenerative disease.” Regen Med 5(6): 933-946.

This paper discusses potential applications of MSCs in medicine, and is of medium difficulty

10.  Meyerrose, T. E., M. Roberts, et al. (2008). “Lentiviral-transduced human mesenchymal stem cells persistently express therapeutic levels of enzyme in a xenotransplantation model of human disease.” Stem Cells 26(7): 1713-1722.

This technical paper discusses how MSCs migrate through the nervous system when introduced into a mouse’s brain

11.  Spees, J. L., S. D. Olson, et al. (2006). “Mitochondrial transfer between cells can rescue aerobic respiration.” Proc Natl Acad Sci U S A 103(5): 1283-1288.

This technical paper describes how transfer of mitochondria between cells can help the cell that receives the mitochonfrion

12.  Wineman, J., K. Moore, et al. (1996). “Functional heterogeneity of the hematopoietic microenvironment: rare stromal elements maintain long-term repopulating stem cells.” Blood 87(10): 4082-4090.

This technical paper discusses the conditions needed to grow MSCs

T. Wang, 7-25-11


Clinical Trials on Huntington's disease

What are clinical trials?

In order for any drug or treatment to be approved for human use by the FDA, it must first successfully pass clinical trials. A clinical trial is a medical or health-related research study in humans that follows a strict protocol in a carefully monitored, scientifically controlled setting. Clinical trials are generally conducted after a drug or treatment has shown promise in research studies using animal models.

What are the different types of clinical trials?^

There are four main types of clinical trials: treatment trials, prevention trials, diagnostic trials, and screening trials.

  • Treatment trials test the effects of new drugs, new combinations of drugs, or new procedures used to treat an illness or condition. Participants in this type of trial would already experience symptoms of HD and could be in any stage of HD.
  • Prevention trials aim to prevent or delay onset of a disease in people who are at risk and test the effects of treatments that do so. Participants in this type of trial would be pre-symptomatic HD patients, usually people who have tested positive for the HD gene but have not yet exhibited any symptoms.
  • Diagnostic trials are conducted to discover better procedures to diagnose an illness or disease.
  • Screening trials are conducted to discover better ways to detect an illness or disease.

Diagnostic and screening trials are not needed to diagnose HD since current genetic tests can reliably and accurately identify HD. However, these types of trial design may be useful to research presymptomatic measures of HD disease progression and/or develop ways to better assess disease risk in the intermediate range where definitive genetic diagnosis is not currently possible. For more on genetic testing of HD, click here.

Clinical trials are conducted in phases.

  • In Phase I trials, researchers first test a new treatment on a small group of individuals, typically 20-80 people, to evaluate its safety, determine a safe dosage range, and to identify side effects.
  • Once the treatment passes Phase I trials, Phase II trials are conducted on more people, around 100-300 people, to see if it is effective and to further evaluate its safety and side effects.
  • Once Phase II trials are completed successfully, the drug moves onto Phase III trials, in which researchers confirm the drug’s effectiveness, monitor any side effects, compare it to standard treatments, and collect information that will allow the experimental drug or treatment to be used safely long-term.
  • Only after the drug or treatment has passed all phases will it be approved by the government.

For more information on the different phases of clinical trials, click here.

All clinical trials have criteria specifying who can or cannot participate. There are many risks and benefits to participating in a clinical trial. For example, participants contribute to medical research, have access to medical care, and if assigned to the treatment group, are given new potential treatments throughout the trial. However, participants may also experience negative side effects as a result of participating, or they may receive a placebo. Clinical trials must follow strict ethical codes and are highly regulated to ensure the safety of participants as much as possible.

What is the Huntington Study Group?^

The Huntington Study Group (HSG) is an international non-profit group whose aim is to support clinical research of Huntington’s disease (HD). It was formed in 1993 and has members and research sites in the US, Canada, Europe, Australia, New Zealand and South America. The HSG often partners with pharmaceutical companies, private foundations, and government agencies to fund research investigating the effects and safety of HD interventions. (For more information on the HSG, click here).

Ongoing Studies that are Currently Enrolling Participants^


2CARE is a phase III trial that aims to study coenzyme-Q10 as a potential treatment for HD. For more on coenzyme-q10, click here. The study aims to measure the effectiveness of coenzyme q-10 in slowing the symptoms of HD and to study the long-term safety of administering the compound to people with HD. Previous studies have shown that coenzyme q-10 slightly slowed the progression of HD, but not enough to yield significant results. Compared to these previous studies, 2CARE uses a much higher dosage for a longer time period. To date, it will be the largest therapeutic clinical trial of HD, with expected enrollment of over 600 in the United States, Canada, and Australia. The study began in March of 2008 and is expected to be completed by April 2014. 2CARE is a double blind placebo study, in which participants are randomly assigned to one of two groups. The experimental treatment group will receive oral administration of coenzyme q10 in chewable form twice a day, for a total of 2400 mg/day. In a preliminary study called Pre2CARE, dosages ranging from 1200 to 3600 mg/day were tested; 2400 mg/day appeared to be the most effective dosage, as smaller dosages were not as effective, and larger dosages resulted in the mildly unpleasant side effect of upset stomach. Researchers will compare total function capacity (TFC) scores, tolerability, adverse events, vital signs, and laboratory test results between the two groups.


The Huntington Study Group (HSG), Massachusetts General Hospital, and the University of Rochester are currently conducting a phase III clinical trial to assess the effects of creatine supplements on slowing the progression of symptoms in HD patients. Creatine is a molecule naturally produced in the body and consumed in the diet, found mostly in meat. Previous studies conducted on transgenic mouse models have shown that mice supplemented with creatine displayed improved motor performance, diminished loss of brain mass, reduced huntingtin aggregates, and delayed neuronal death. The study is called the Creatine, Safety, Tolerability, & Efficacy in Huntington’s disease (CREST-E). Participants are randomly selected to receive either 40g per day of powdered creatine monohydrate or 40g per day of a placebo. The study is a fairly large clinical trial. It will involve 44 research centers from around the world and enroll up to 650 participants. The study will last 37 months and is estimated to be completed in December of 2014. (For the most updated information on this study, click here).


Earlier in 2009, the HSG received funding from the NIH to test the safety and tolerability of coenzyme-Q10 in individuals who have tested positive for HD but do not yet show any motor symptoms. The study is called PREQUEL (Study in PRE-manifest Huntington’s disease of coenzyme Q10 (UbiquinonELeading to preventive trials) and is a phase II trial. The study will be conducted at 10 clinical sites throughout the nation and is the first therapeutic research study in pre-manifest HD patients. Participants will be randomly assigned to experimental groups receiving 600, 1200, and 2400 mg/day of coenzyme q10. The principal investigators hope that this initial trial will lead to later trials that study the delay of HD.

Ongoing Studies that are No Longer Enrolling Participants^


ACR16 is a dopamine stabilizer and can enhance or inhibit dopamine controlled functions. For more information on the role dopamine plays in HD, click here. The HSG is conducting a phase II clinical trial testing different doses of ACR16 on HD patients age 30 and older. HART is sponsored by NeuroSearch Sweden AB, a biopharmaceutical company, and is being conducted in 35 research sites across North America. Previous studies showed that ACR16 is safe and tolerable in patients with HD and Parkinson’s disease. Additionally, it has been shown to significantly improve patients’ voluntary and involuntary movements. Participants are randomly assigned to one of four groups-three experimental groups given different doses of ACR16 and a placebo group. The study occurs over a course of 12 weeks. As of October 2010, results have been promising, as patients in the highest dosage group (90 mg/day) displayed significant improvement in motor function as measured by the modified Motor Score (mMS).

Recently Completed Clinical Trials^


HORIZON was a phase III clinical trial conducted by the HSG and the European Huntington’s Disease Network that investigated whether dimebon is safe and effective in improving cognitive abilities in patients with HD. Dimebon is an experimental drug that has been shown to prevent the death of brain cells in animal models and is currently being tested to treat HD and Alzheimer’s Disease. It is thought to work by stabilizing and improving function of the mitochondria. For more information on dimebon, click here. The study was conducted in various centers in the United States, Canada, Europe, and Australia. It was a double-blind, placebo study in which participants were either given 60 mg of Dimebon daily or a placebo. Results showed that dimebon is not effective in treating HD. There was no statistically significant difference in symptoms between the experimental and placebo groups. According to the president and chief executive officer of Medivation, development of dimebon in HD will be discontinued.


Minocycline is an antibiotic that is primarily used to treat acne and other skin disorders. For more on minocycline, click here. The goal of DOMINO was to assess whether minocycline was safe and effective in slowing HD progression and whether further studies should be conducted. The study is a phase II clinical trial that was started in 2006 and completed in November 2008 by the HSG with funding by the FDA Office of Orphan Products Development. It was a double-blind, placebo experiment in which participants were randomly assigned to a treatment group that received 100mg of minocycline twice daily or a placebo. The TFC scores of the two groups were then compared. Results showed that while minocycline was safely tolerated, it did not produce a significant effect in terms of delaying HD symptoms, and thus, further study of minocycline in treating HD is not warranted.


TREND-HD was a large phase III trial that began in September 2005 and was completed in August 2007. The goal of the study was to determine whether ethyl-EPA (Miraxion) slowed the progression of motor decline in HD patients. Ethyl-EPA is an omega-3 fatty acid commonly found in fish oil. Study participants had mild to moderate HD, meaning they displayed early signs of HD but were self-sufficient in daily living activities. For the first 6 months, the treatment group received ethyl-EPA while the placebo group received a placebo. For the next 6 months of the study, the placebo group was also given the active drug. Interestingly, there were no significant differences in Total Motor Scores between the two groups after the first 6 months of the placebo study. However, after the next 6 months in which all participants received the drug, the experimental group showed improvement as compared to the group that had initially received the placebo for the first 6 months. Further studies will need to be conducted to determine the efficacy of ethyl-EPA. There did not appear to be any safety concerns. After the study was completed, the investigators and sponsor of the study, Amarin Neuroscience Ltd., took the unprecedented step of telling the study participants about the results of the study by calling them and inviting them to a telephone conference regarding the study results. Study participants are typically not informed of the results of the clinical trials they participated in.


Huntington Study Group (HSG) and Prestwick Pharmaceuticals collaborated on a clinical trial. Called TETRA-HD involving tetrabenazine, a dopamine depletor. Led by Dr. Frederick J. Marshall from the University of Rochester Medical Center, TETRA-HD was a phase III clinical trial with the goal of determining the optimal dosage of tetrabenazine in treating chorea in people with HD. The trial was carried out at 16 different HSG sites in the United States, involving a total of 84 participants with HD. Fifty-four of the participants were randomly assigned to receive tetrabenazine for 12 weeks with increasing dosages over the first 7 weeks. The other 30 served as the comparison group and received a placebo. The results of the study indicated that tetrabenazine is effective in treating chorea with side effects that are less severe than those associated with other anti-choreic drugs. On the CGI Global Improvement Scale, 6.9% of the patients receiving placebo had more than minimal improvement compared to 45.1% of the patients receiving tetrabenazine. Clinical assessments showed that tetrabenazine was associated with drowsiness and insomnia in four patients, depressed mood in two, parkinsonism in two and akathisia in two. Most cases of adverse effects improved after adjusting dosage levels, but the risk of side effects should be considered. These results confirm the benefits of tetrabenazine usage in ameliorating the symptoms of chorea. In August 2008, the FDA approved tetrabenazine as the first drug for treatment of chorea, and it is used worldwide today.

For more information:^


-A. Zhang, 7-5-11 More

Studying Huntington's Disease

This chapter explains some of the many different types of research that scientists use to study Huntington’s Disease.

Fig Y-1: Transgenic Mouse Paw

The above figure shows a rather strange mouse paw photographed under fluorescent light. Why on earth is this paw green? Despite its appearance, the mouse is not an alien nor has it taken a bath in nuclear waste. Instead—and this also sounds crazy, but it’s true—the greenness comes from a special “fluorescence gene” that belongs to a jellyfish! When the mouse was just an embryo, scientists inserted this special gene into it and the gene became incorporated into the mouse’s DNA. When the gene then had its effects in the mouse, the resulting fluorescent protein caused the mouse’s whole body to light up, just as if it were still in the jellyfish.


You might be asking yourself: what could this green mouse possibly have to do with Huntington’s Disease (HD)? Although it is not clear from the picture, the mouse may actually have a lot to do with HD. Since a foreign gene is incorporated in its DNA, the green mouse is called a transgenic mouse. The fluorescence gene is just one example of a multitude of genes from many different animals that researchers are now adding into mouse DNA. One particular gene of interest is the human HD gene, which has been inserted into transgenic mice for research purposes for a number of years. These mice are part of the large field of animal research, a field that is teaching scientists important new things about HD.

Animal research is just one piece of the immense HD research puzzle. Other forms of HD research include family tree studies, epidemiological studies, genetic studies, postmortem studies, and clinical studies. Each of these areas of research has contributed a great deal to our current understanding of HD. More importantly, each of these areas will help contribute to the exciting breakthroughs in future HD research. These various areas of research, and some of the facts that they have already uncovered, are the subject of this chapter.

Animal Research^

  • Click here to learn about the basics of HD mouse models
  • Click here to view an article on the ethics of HD testing

Epidemiological Studies^

Epidemiology is the study of the spread of diseases within and between human populations. Similar to family tree studies, epidemiological studies involve finding which individuals show the symptoms of a disease and which do not. However, epidemiological studies differ from family tree studies in that epidemiologists generally study a greater number of people at one time. In fact, many epidemiological studies deal with the populations of entire countries, or even continents. In the case of HD epidemiology, researchers might, for example, dig through a country’s health statistics and find out how many individuals have been diagnosed with HD. These data can then be combined with other records—or perhaps with personal interviews if the individuals are willing—in order to reveal aggregate trends about HD in populations. For instance, one epidemiological study showed that five people per million get HD in Finland, as opposed to between 30 and 70 people per million in most other Western countries. Another study showed that the prevalence of HD among African Americans in South Carolina is only 9.7 per million, a strikingly low prevalence, and five times lower than the prevalence for Caucasians in the same area. These are just a few examples of the interesting findings of HD epidemiology. Since they tell us information about HD in populations throughout the world, epidemiological studies will be a vital piece in the HD research puzzle for many years to come.

Genetic Studies^

Fig Y-2: The Human Genome

Genetic studies seek to find a link between a particular gene (or genes) and a certain disease. If a genetic basis has already been established for a given disease, the studies seek to find the location of the gene(s) within the DNA. To do this research, geneticists depend heavily on data from family tree studies. They look at patterns of disease inheritance across generations of a given family, and, if possible, they supplement these data by studying the blood samples from living members. With regard to HD research, the most important question since HD was shown to have a genetic basis has been this: On which of the 23 human chromosomes is the Huntington gene located? (The 23 human chromosomes are shown in Figure Y-2). The most effective research to date to determine the location of the Huntington gene involves so-called “linkage studies”, as described below.

Linkage Studies^

Linkage studies use data from very large families with a history of HD. The principle behind linkage studies is that if nearly every person with HD in a single family shares the same version of a particular “marker trait,” such as the same color eyes or the same blood type, then the genes that code for that marker trait must be located close to the Huntington gene on the same chromosome. The marker’s gene and the Huntington gene are then said to be “linked” to one another. The logic behind such studies is straightforward—genes that lie close together on the same chromosome will tend to be inherited together over the generations. (Genes lying farther apart on the same chromosome are often not inherited together due to a complex process called recombination.)

In reality, eye color and blood type were not themselves useful as markers in HD studies, for their genes lie on chromosomes different from that of the Huntington gene. However, other markers were a tremendous help in locating the Huntington gene, including markers in a region of the Huntington-bearing chromosome called the “non-coding region.” The DNA in these regions does not code for proteins, but it still consists of a linear sequence of the chemical components called nucleotides. Using laboratory techniques, researchers found a few regions or “loci” of this DNA where family members with HD all had the very same nucleotide sequence. Since the researchers knew the locations of these particular loci, they were able to determine that the Huntington gene lies close by on the same chromosome. For example, a locus called “D4S90″ had the same nucleotide sequence in every person with HD in a particular family. Since D4S90 was known to reside on chromosome #4, researchers concluded that the Huntington gene must lie along chromosome #4, very near D4S90. Data from other loci have confirmed this fact.

Once the location of the Huntington gene was identified through linkage studies, further genetic research (including linkage studies and other types of genetic investigation) revealed more details about the Huntington gene. With the cutting-edge technologies available today, it is likely that genetic research will continue to tell us a great deal about HD.

Human Postmortem Studies^

Human postmortem studies are carried out using the donated bodies of people who have died. In the case of HD, postmortem studies have been very important in locating the specific parts of the brain that HD affects. In comparing postmortem HD brains with non-HD brains, doctors have found that HD brains often show damage or decay in the basal ganglia, whereas no damage or decay is seen in the non-HD brains. (For more information about the basal ganglia and the affects of HD on the brain, click here). Postmortem studies also offer insight into some of the cellular events that take place in the brains of people with HD. For instance, by using very special staining techniques, postmortem studies have investigated the presence of nuclear inclusions (NIs) (For more information about NIs and huntingtin protein aggregation, click here). Understanding NIs and other cellular phenomena will be tremendously helpful in developing future treatments for HD. For this reason, postmortem studies are a very important type of HD research.

Family Tree Studies^

Family tree studies (also known as “pedigree studies”) have been a tremendously successful form of HD research throughout the years. This type of research involves looking at a large number of related individuals through several generations and searching for any disease-related similarities between them. Such research, combined with blood samples taken from living family members, allowed scientists to establish that HD is a genetic disease in the first place. For more information about the most famous family tree study on HD (conducted in the vicinity of Lake Maracaibo, Venezuela), click here.

Clinical Studies^

Clinical studies (also called clinical trials) are studies that involve human subjects with informed consent. In the case of HD, these studies have been critical in identifying the various symptoms that doctors now use to diagnose the disease (For more information on symptoms of HD, click here). In fact, the very name “Huntington’s Disease” comes from Doctor George Huntington, who was the first to notice that many of his patients’ symptoms were part of the same disease.

Now that the symptoms and typical age of onset of HD are clearly defined, clinical studies today are more geared toward judging the effectiveness of particular drug treatments. After a new drug passes the test in animal studies, the U.S. Food and Drug Administration requires that clinical studies be performed to ensure that the drug is safe for humans. Once approved by the Food and Drug Administration, drugs may be sold either as prescription drugs or over-the-counter drugs.

For more information on clinical trials, click here.

For further reading^

  1. Freeman, T. B.; Cicchetti, F.; Hauser, R. A.; Deacon, T. W.; Li, X.-J.; Hersch, S. M.; Nauert, G. M.; Sanberg, P. R.; Kordower, J. H.; Saporta, S.; Isacson, O. : Transplanted fetal striatum in Huntington’s disease: phenotypic development and lack of pathology. Proc. Nat. Acad. Sci. 97: 13877-13882, 2000.
    A technical paper regarding the transplantation of fetal neurons.
  2. Robbins, C.; Theilmann, J.; Youngman, S.; Haines, J.; Altherr, M. J.; Harper, P. S.; Payne, C.; Junker, A.; Wasmuth, J.; Hayden, M. R. : Evidence from family studies that the gene causing Huntington disease is telomeric to D4S95 and D4S90. Am. J. Hum. Genet. 44: 422-425, 1989.
    A technical paper regarding a particular linkage study that showed the Huntington gene to be located on chromosome 4.
  3. Mouse paw picture obtained from National Geographic web site (

Updated by T. Wang, November 2010


Animal Research: The Ethics of Animal Experimentation

Many medical research institutions make use of non-human animals as test subjects. Animals may be subject to experimentation or modified into conditions useful for gaining knowledge about human disease or for testing potential human treatments. Because animals as distant from humans as mice and rats share many physiological and genetic similarities with humans, animal experimentation can be tremendously helpful for furthering medical science.

However, there is an ongoing debate about the ethics of animal experimentation.


An Introduction to Animal Models of Huntington's Disease

Huntington’s disease research and drug testing frequently involve the use of mouse models. However, when different scientists refer to an HD mouse model they may not be referencing the same model. A variety of HD mouse models exist and are used regularly. This chapter will describe the most common HD mouse models, how they differ, and the ways in which they mimic the disease.

This article makes reference to both the mouse and the human huntington gene. The mouse huntington gene is a homolog of the human huntington gene, but there are subtle differences between them. For example, the mouse huntington gene has fewer CAG repeats than the normal human huntington gene, explaining why spontaneous mouse models of the disease do not exist. Despite this difference, the mouse huntington gene is 81% similar to the human huntington gene at the DNA level, showing that the two are true homologs. Different mouse models have been generated since the isolation of the HD gene in 1993. There are three general types of mouse model: knockout, transgenic, and knock-in.

Knockout models were the first models to be generated. These are models where the gene coding for the huntingtin protein (the mouse huntington gene) is removed or interrupted so that the DNA can not be transcribed. Although these models provide valuable scientific insights, they do not actually represent the disease because mice that are nullizygous for the huntington gene die during embryogenesis.

Transgenic models are made when the mutant human huntington gene, or a fragment of that gene, is inserted into the nuclei of a model organism. In transgenic models the gene insertion is not targeted to a specific location in the animal genome, meaning that where the gene inserts itself cannot be predicted. For a mouse model, this means that the mouse will express both the two normal copies of the mouse huntington gene as well as the human mutant gene or fragment that was inserted into its genome. It also means that the expression of the inserted mutant human huntington gene will not be controlled by the homologous promoter region of the mouse huntington gene. This frequently leads to much higher protein expression than normal endogenous levels.

Knock-in mice have either part of or the entire human mutant huntington gene inserted in place of part of or the entire endogenous mouse gene. Knock-in mice, therefore, carry the expanded CAG repeat mutation in the same place in the genome that it would appear if it were to develop naturally. This is the most faithful model in the sense that the mutant gene is located in the appropriate genomic context and will have the normal promoter region associated with the huntingtin protein. Knock-in mouse models can be either homozygous or heterozygous for the huntingtin mutation.

To learn more about the process of creating genetically modified model animals click here.

The remainder of this chapter considers the following topics:

More about knock-out mice^

Although knock-out mouse models are not a viable model of HD, studying these early models did increase understanding of the disease. Showing that a complete absence of the huntington gene, and consequently the huntingtin protein, was embryonic lethal revealed that Huntington’s disease is not a loss-of-function disease. The huntingtin protein is necessary for successful embryogenesis and when it is entirely absent the organism will die before birth. This suggests that the mutant huntingtin protein in individuals with HD must fulfill some of the same functions as the normal huntingtin protein.

Since most individuals with HD are heterozygous for the disease, they do still have one copy of the normal huntington gene and can express some levels of the normal huntingtin protein. However, HD homozygotes do develop normally and have an age of disease onset comparable to HD heterozygotes, confirming that the mutant protein fulfills some of the normal huntingtin protein functions. This suggests that HD is a dominant “gain of function” disease as HD patients are able to live with either one or two mutant HD alleles.

Some scientists suggest that despite the finding that HD is a gain-of-function disease, it is still possible that some loss of normal huntingtin protein function contributes to the disease. One study supporting this idea found postnatal brain degeneration in mice in which huntingtin was selectively inactivated in the brain and testes after early embryogenesis. Furthermore, another study found that normal huntingtin, but not mutant huntingtin, increases the production of brain-derived neurotrophic factor, which is necessary for the survival of striatal neurons in the brain. Although both of these studies support the conclusion that loss of normal huntingtin function contributes to the pathogenesis of HD, most researchers believe that HD should be considered a gain-of-function disease.

More about transgenic mice^

The earliest HD transgenic mouse models and still some of the most frequently used are from the line designated R6. In this model, a fragment encoding exon 1 of the human mutant huntingtin gene is inserted into the mouse genome. Four different R6 lines were established, with CAG repeat lengths ranging from 115-156. The R6/2 mouse, one of this line, is probably still the most commonly used HD mouse model. All four lines use the IT15 promoter. In humans it only takes a CAG repeat length of 36 or more to develop HD, but mouse models seem to need much larger repeat lengths to develop an HD-like phenotype, possibly due to their shorter lifespan.

Three of the R6 lines ubiquitously expressed the mutant huntingtin protein and exhibited an abnormal phenotype. The level of mutant huntingtin protein expression varied among these three R6 lines. The level of protein expression is usually represented as a percentage of the normal endogenous mouse huntingtin protein level. The R6/1 line had the lowest expression, measured at about 31% of endogenous mouse huntingtin levels. The R6/2 and R6/5 lines had an average expression of 75% and 77% endogenous mouse huntingtin levels. These lines displayed motor abnormalities such as chorea-like movements, weight loss, seizures, frequent urination, unusual vocalizations, and sudden deaths of unknown cause.

Newer transgenic mouse models now express longer fragments or the full length human huntington gene. One example is the YAC line. Mice from this line express the full length human huntington gene with a CAG repeat length of up to 128. These newer transgenic mouse models tend to show progressive motor abnormalities as well as neuronal loss in the striatum. These symptoms all closely parallel human symptoms in HD, and provide evidence that these mice might accurately model many aspects of the disease.

More about knock-in mice^

The first HD knock-in models did not display overt motor deficits like those observed in the first transgenic mouse models. Instead, they displayed some behavioral abnormalities such as increased aggression. The earliest models usually had between 70-80 CAG repeats. One reason that the early models did not reveal a phenotype resembling HD may be that they had a low number of CAG repeats for mouse models, which tend to have very high CAG repeat lengths.

Later knock-in models were more promising as they did show abnormal motor abnormalities, as well as cellular, molecular and neuropathological abnormalities suggestive of an HD phenotype. They also tended to reveal behavior abnormalities at earlier ages. These later knock-in models tended to have many more CAG repeats than the earlier models (from 94-140 CAG repeats).

Knock-in models are not all the same. Besides differences in CAG repeat length, there are also differences in how much of the mutant huntington gene is inserted and in the background strain of the mouse. In some models, only the sequence coding for the polyglutamine tract in exon 1 was replaced, while in others the entire first exon was replaced. The models that replace all of exon 1 with the mutant version should be more accurate representations of the human condition, because many of the proteins that interact with mutant huntingtin do so in the region immediately surrounding the polyglutamine tract. In other words, when a larger portion of the gene is inserted and it is expressed at an endogenous level, the mouse imitates the genomic context of the huntingtin mutation as well as the mutation itself, and consequently then phenotype may be more consistent with human HD. Most significantly, this means that any results found with these mice, either about relevant proteins, molecular pathways involved, or pre-clinical drug studies, are much more likely to be relevant to human HD.

Why do transgenic mice frequently display a more obvious disease phenotype?^

Transgenic mouse models sometimes seem to display a phenotype more similar to human HD than the phenotypes displayed by knock-in models. This might seem confusing since the knock-in models are theoretically more accurate representations of the disease genotype.

One reason for this unexpected result is that transgenic mice usually only express a fragment of the human mutant huntington gene. The mutation is then expressed as a truncated, or shortened, protein fragment. In vitro studies have founded the truncated protein to be more toxic than the full length protein, which could explain the more severe disease phenotype of these models.

In some transgenic models, this result could also occur because of unnaturally high expression levels of mutant huntingtin protein. Recall from above, for example, that HD line mice sometimes express as much as five times more mutant huntingtin protein than endogenous huntingtin protein. This overexpression could exaggerate the disease phenotype.

How are animal models used?^

Mouse models allow researchers to study aspects of the disease that would be impossible to do with human subjects. For instance, researchers can focus on changes in the brain or in cell-to-cell interactions during early stages of the disease with animal brain tissue. Mouse models are also a crucial part of early testing of potential therapeutics. Because mice have a shorter lifespan than humans, testing can be carried out much faster. Additionally, in the earliest stages of drug development it would be unethical to subject humans to the risks associated with the new drugs being tested.

Unfortunately, the use of mouse models in drug development does have some drawbacks. Mouse models of HD are not exactly the same as the human disease and there are also differences in how a drug will affect one species compared to another. Hopefully, as scientists develop more sophisticated models that better represent the genetics of human HD, model organisms will become more relevant for drug development and therapeutic testing. But it is always vitally important to keep in mind that drugs that look promising in animal models may not prove successful when applied to humans.

Further Reading^

– Adam Hepworth, 11-21-08


The HD Measuring Stick: Assessment Standards for Huntington's Disease




There are a number of well–established methods used to measure the severity and progression of Huntington’s disease (HD). These can evaluate a patient’s mental and physical capabilities and track any changes over time. Having standardized methods for measurement is important because it allows for the comparison of patients in clinical trials and the quantification of symptoms to guide treatment and therapy options.

Test Definitions^

Each of the tests measures the subject’s abilities to perform various mental and physical functions in different ways. The tests are often used together, providing a more complete picture of the patient’s physical and cognitive well–being.

It is important to recognize that the pages in this article are intended only to give general information about some of the different tests used clinically and in research. We do not recommend the self–administration of these tests. Accurate administration of these tests requires qualified personnel such as doctors, therapists, and other trained professionals.

Fahn Rating Scale (Physical and Mental)^

The Shoulson–Fahn functional capacity rating scale was first proposed in 1979. It measures independence in daily activities such as dressing, eating, managing personal finances, and engagement in occupation. Functional capacity in each category is ranked from Stage 1 to Stage 5, with Stage 1 representing the most independent level of function. The table below summarizes the scale as it was originally proposed:

Shoulson-Fahn Functional Capacity Rating Scale as Proposed in 1979
Engagement in occupation Capacity to handle financial affairs Capacity to manage domestic responsibilities Capacity to perform activities of daily living Care can be provided at
Stage 1 Usual level Full Full Full Home
Stage 2 Lower level Requires slight assistance Full Full Home
Stage 3 Marginal Requires major assistance Impaired Mildly impaired Home
Stage 4 Unable Unable Unable Moderately impaired Home or extended care facility
Stage 5 Unable Unable Unable Severely impaired Total care facility only

This table was adapted from Shoulson and Fahn, 1979. See further reading.

Unified Huntington’s Disease Rating Scale (UHDRS) (Physical and Mental)^

The UHDRS is a standardized rating system used to quantify the severity of HD. Used clinically and in research, it measures the patient’s abilities in four general areas: motor, cognitive, behavioral, and functional. The different portions of the test may be administered separately.

The following table summarizes the individual categories tested in the motor section of the UHDRS:

Skill Category Description
Ocular Pursuit the ability of the patient to follow a finger with the eyes in both the horizontal and vertical directions
Saccade Initiation the ability of the patient to turn the head in both the horizontal and vertical directions
Saccade Velocity the speed at which the patient is able to turn the head both horizontally and vertically
Dysarthria the presence of speech that is slurred, slow, and difficult to understand
Tongue Protrusion the ability to stick out the tongue and the speed to which the task is completed
Finger Taps the ability to tap the fingers of both hands (15 repetitions in 5 seconds is considered normal)
Pronation/Supination the ability to rotate the forearm and hand such that the palm is down (pronation) and to rotate the forearm and hand such that the palm is up (supination) on both sides of the body
Fist-Hand-Palm Sequence the ability to complete the sequence (making a fist, opening the hand palm down, and then rotating the hand palm up) more than 4 times in 10 seconds without cues is considered normal
Rigidity in arms the severity to which the range of motion of the arms is limited
Bradykinesia slowness in initiation and continuation of movements
Maximal Dystonia abnormal muscle tone (measured separately in the extremities, face, and trunk)
Maximal Chorea involuntary jerky movements of the body (measured separately in the extremities, face, and trunk)
Gait walking with normal posture
Tandem Walking the ability to walk in a straight line from heel to toe. The ability to do so regularly for 10 steps is considered normal
Retropulsion the ability to stand after being pushed back

In each category, patients are scored from 0 to 4, with 0 representing normal function, and 4 being the most severe dysfunction. The total score is the sum of the scores in the individual sub–categories. A higher UHDRS score indicates a more severe disease progression.

Zung Depression Scale (Mental)^

Patients with Huntington’s disease are significantly more likely to display signs of depression than people in the general population. Up to half of patients with HD demonstrate symptoms of depression. To learn more about the relationship between HD and depression, click here.

The Zung Depression Scale is a simple 20 item questionnaire. Patients judge statements about how they have been feeling on a qualitative scale ranging from “a little of the time” to “most of the time”. Each of the patient’s answers is then given a score from 1–4 and the sum of these scores is the total score. The range for total scores is between 20 and 80; patients with depression usually score between 50 and 69, while those with severe depression score above 70.

The scores and what they imply are summarized in the table below:

Score Indication
20-49 Normal
50-69 Depression
70+ Severe Depression

Mini–Mental State Examination (Mental)^

The Mini Mental State Examination (MMSE) assesses the overall cognitive status of patients. Its use is not limited to measurement of the progression of HD symptoms. For example, MMSE can also be used in the assessment of patients with other neurological diseases, such as Alzheimer’s disease.

It analyzes the patient’s abilities in 5 different areas of mental status: orientation, attention and calculation, recall, and language. Created in 1975, it is an effective 11–question test that only takes 5–10 minutes to administer and score. It has been used widely in both clinical practice and in research to measure the cognitive abilities of patients and subjects.


To test the patient’s orientation, he or she is asked what year, season, date, day, and month it is. He or she is then asked what state, country, town, hospital, and floor he or she is currently on.


Testing registration next, 3 objects are named, and the patient is given a chance to name all 3 of them. Assessing calculation abilities, patients are asked to count by 7’s.


Recall is tested by asking the patients to repeat the 3 objects he or she learned before.


Finally, language skills are tested in multiple parts. The patient is asked to name a pencil and watch, then is asked to repeat the phrase “No ifs, ands, or buts”. Next, he or she is asked to follow a verbal 3–stage command, and then a written command. Lastly, the patient is asked to write a sentence and copy the following drawing of two interlocking pentagons:


The successes and shortcomings of the patient are added up, and a total score is calculated. The maximum score on the MMSE is 30. Scores of 23 or lower are indicative of cognitive impairment.

Barthel Index (Physical and Mental)^

The Barthel Index (BI) is a commonly used scale to help assess the patient’s independence and his or her need for supervision or assistance. The test scores the patient’s ability to perform 10 basic daily living activities. Full credit for each criterion is not given if the subject needs even minimal help or supervision. The activities considered on this index include:

Scores for each individual item are given in increments of 5. The score for the items ranges from 5 to 15. The maximum total score is 100, and the higher the score, the more independent the patient.

Tinetti Scale (Physical)^

The Tinetti scale, also known as the Tinetti performance Oriented Mobility Assessment (POMA), is an easily administered test that measure’s a patient’s gait and balance abilities. The test takes approximately 10–15 minutes to complete and score.

The test is divided into two main parts, a balance portion and a gait portion. The patient’s balance in both the sitting and standing positions are measured. Additionally, the ability to stand from the sitting position and to sit down from standing up are quantified.

In the gait portion of the test, the subject is asked to walk across the room at a “normal” pace, and then back at a “rapid, but safe” pace. Various parts of the subject’s walk are noted, such as hesitation after being prompted to go, swing of the feet (height and path), step symmetry, step continuity, trunk sway, heel position, and smoothness of gait.

The test is scored on a 28 point scale. The indications for each score range are summarized in the table below. Scores ranging from 25–28 indicate a low fall risk, scores between 19 and 24 indicate a medium fall risk, and scores below 19 indicate a high risk for falls.

Score Indication
0-18 High risk for falls
19-24 Medium risk for falls
25-28 Low risk for falls

Physical Performance Test (Physical)^

The Physical Performance Test quantifies the subject’s performance in physical tasks. It is a standardized 9–item test that measures the subject’s performance on functional tasks:

Subjects are given two chances to complete each of the 9 items, and assistive devices are permitted for the tasks that require a standing position (items 6–9). Both the speed and accuracy at which the subjects complete the items are taken into account during scoring. The maximum score of the test is 36, with higher scores indicating better performance.

Symbol Digit Modalities Test (SDMT) (Mental)^

The Symbol Digit Modalities Test (SDMT) is a brief and simple mental test that takes less than 5 minutes to completely administer and score. The test measures the subject’s information processing speed and attention.

It involves a simple test in which numbers are randomly substituted for letters or geometric symbols. The subject is given a translation key, and is asked to translate them within 90 seconds. The task is easy for normal subjects to complete, but is more difficult for those patients with cognitive dysfunction.

The translation can be given in either a written or oral format. This flexibility in format allows for the testing of almost all subjects, including patients with speech or motor disorders. Additionally, the written format allows for the test to be administered to patients in a group setting.

Thurstone Word Fluency Test (Mental)^

The Thurstone Word Fluency Test (TWFT) is a simple test that measures the subject’s communication abilities. Given in either a written or oral form, the TWFT is commonly used to detect the presence of and define the nature of any cerebral dysfunctions.

First, the subject is given five minutes to write down or say as many words as possible that begin with the letter “s”. Next, he or she is given four minutes to list as many four–letter words as possible that begin with the letter “c”.

Several studies have shown that the TWFT is very accurate in identifying subjects with reduced cerebral function. However, the test is unable to identify which specific areas of the brain have been damaged. For example, the test can determine whether or not a patient has brain damage, but it cannot be used to detect whether the damage is on the left or the right side of the brain.

Despite its shortcomings, the TWFT is informative and is commonly used in combination with other tests to help gauge the presence and extent of brain damage in patients.

Stroop Test (Mental)^

The Stroop Test is a simple mental test commonly used to measure the subject’s attention and mental flexibility. It takes advantage of the Stroop effect, the interference that arises when the brain is presented with conflicting signals.

Patients are presented with a list of colors, like in the image below, each printed in a different color:

stroop test

Next, the subject is asked to name the color of each word rather than what the word is. For example, the correct response to the first word in the third column would be “red” not “blue”.

The subject’s accuracy and speed at the Stroop Test can be recorded and used to track the progression of cognitive disabilities.

Neuropathological Scales^

In addition to the tests discussed in this article, there are also various neuropathological grading systems which measure physical change in the brain as a result of the disease. One such scale that has been developed is the five–tiered pathological grading system, which rates damage done from Grade 0 to Grade 4, with Grade 4 having the most severe damage.

For further reading^

-A. Pipathsouk, 1-15-10


UC Davis Center of Excellence

HOPES team members Adam Hepworth and Amy Frohnmayer visited the UC Davis Center of Excellence team on August 2, 2007. The visit took place at the Veteran Affairs (VA) Hospital in Rancho Cordova, a facility which the Center uses for running clinical trials. During the visit the HOPES members had the opportunity to see exactly what takes place during a clinical visit for a participant enrolled in the COHORT trial. HOPES would like to offer thanks to all of the clinic staff who made the visit a positive experience with a special thank you to Teresa Tempkin for offering so much of her time and expertise.


Left to Right: Amy Frohnmayer, Teresa Tempkin, Adam Hepworth


Although the UC Davis Center of Excellence (COE) main office is located at the Davis Medical Center in Sacramento, our visit took place at the VA Hospital in Rancho Cordova. On the fourth floor of the main hospital building, an entire wing is devoted to clinical trials run by UC Davis doctors. The UC Davis COE had reserved space on this wing to run the COHORT clinical trial.

Tempkin explained that access to these clinical research facilities was crucial for successfully running clinical trials. At the VA Hospital, she has access to space and equipment unavailable at the Davis Medical Center. Without these resources, it would not be possible for the UC Davis COE to participate in clinical trials like COHORT.

COHORT is an acronym for Cooperative Huntington’s Observational Research Trial. It is currently the largest Huntington Study Group HD clinical trial and it takes place at 40 sites in North America and Australia. It is a long term observational study with no official end date. The goal of the study is to collect a huge database of information from subjects diagnosed with HD and make that information available to researchers around the world. To that end, trial enrollees commit to annual study visits for as long as they are willing and able. The study’s control group consists of adults who are part of an HD family. This particular study does not enroll individuals who are at risk of HD but do not know their gene status. The UC Davis COE entered the trial in January of 2007 and has already enrolled over 30 participants. For more information about COHORT click here.

Behind the Scenes of a COHORT Visit^

During our visit, we were fortunate enough to observe first hand most of what happens during an annual COHORT visit. In addition to a standard physical exam, the participant undergoes a few short cognitive and neurological assessments. A variety of questionnaires are also administered to the participant. These help assess the participant’s emotional and behavioral status and help to gauge the affect of disease progression on daily living and basic skills. The participant gives a detailed medical history and reports on any medications he or she currently takes. And finally, the visit includes a blood draw. With participant consent, the blood samples collected in COHORT are placed in a research facility and made available to HD researchers around the world. This is one unique aspect of the study that offers an amazing opportunity for HD researchers. Because all of the information collected during a participant’s annual visit is matched by code (all personal identification information is kept strictly confidential) to his or her blood sample, researchers using the samples can also have the benefit of knowing a huge variety of information about the status and progression of HD in the individual to whom the sample belongs. This additional body of information linked to every blood sample allows for much more sophisticated research and analysis.

Some of the procedures during the visit are fairly generic and could easily take place in any clinical research trial, while others are tailored specifically to HD participants. For instance, one of the cognitive tests administered was a short mental exam. One part of this test includes questions designed to measure a subject’s orientation in time and space, ability to maintain focus and attention, and basic communication skills. Although these are all important cognitive abilities, they are not usually the functions impaired by HD, which tends to negatively affect executive function. A later cognitive test looked specifically at executive function through tests of concentration and multitasking ability. That test tends to be more sensitive to cognitive changes in an HD patient.

The neurological examinations also include both more general tests and specific tests best suited for an HD population. Although it is common to think of chorea as the definitive neurological sign of HD, the COHORT neurological exam looks for many other HD-specific symptoms. Tempkin explained that although chorea is one of the most visually dramatic signs of HD, it is not usually the most debilitating aspect of the disease. Problems with balance and fine motor coordination are more likely to lead to serious complications through injuries like falls. Chorea is also extremely variable, and progresses at a different rate in different patients. This makes it especially important to test a variety of motor changes. The COHORT exam includes tests for characteristically abnormal eye movements, motor impersistence, speech changes, muscle stiffness or tightness, and balance impairment.

Administering the questionnaires about emotion and behavior may be one of the more difficult parts of the evaluation. The information from these questionnaires comes through the filter of the subject’s own perceptions. Frequently, the goal is to take the subject’s answers and turn them into numeric values along some scale, thus quantifying the measurement. However, assigning values objectively to subject responses can be difficult and some of the responsibility ultimately rests with the judgment of the investigator asking the questions and recording the results. In an effort to make the process as scientifically objective as possible, numeric values along the scales are defined very specifically. Investigators must constantly struggle to ensure that results from these questionnaires are consistent across subjects and across testing sites.

Participant Rights^

Tempkin emphasized that careful consideration of participant rights is an integral part of running a study like COHORT. No potential participant is ever pressured into entering the study or enters without the most comprehensive knowledge of what the study entails. Before enrolling in the study, every potential participant must sign a consent form. The seventeen page long document can seem a little intimidating, but every effort is made to ensure that the potential participant reads and understands the entire form.

Tempkin explained that when the patient first expresses interest in participating, the form is mailed to his or her home address. This ensures that there is time to read over and think about the form in a no-pressure environment. If the potential participant still wants to enroll in the study, he or she will come in for the first visit. At this point, Tempkin will verbally review the consent form with the potential participant, making sure that there are no misunderstandings. The participant will only be enrolled once Tempkin satisfies herself that he or she fully understands and agrees to all of the conditions.

Consent forms are a key part of protecting participant rights in clinical trials. There are some aspects of the consent form that remain consistent across all UC Davis studies. For instance, the form starts out with the Experimental Subject’s Bill of Rights, which states explicitly many of the participant’s most important rights. For instance, every subject is entitled to know what the study is trying to discover, what procedures will happen to him or her during the course of the study, and what the risks and benefits of the study will be. Additionally, the bill of rights guarantees that the subject can stop participating at any time and that failure to participate will not impact future medical care.

The seventeen page length of this particular consent form is due largely to the complexity of the COHORT trial. Different aspects of the trial, such as the medical history and the tissue sampling, must be consented to separately from the trial as a whole. This ensures that participants are fully aware of everything to which they are consenting and grants more participant flexibility. If a subject wants to participate in the trial, but doesn’t want to share his or her medical history, this is possible. Tempkin mentioned, however, that patients in the HD community tend to want to participate as fully as possible in the trials, because they know the huge importance and potential benefits of research studies like COHORT.

More about Clinical Trials in the HD World and the Davis Center of Excellence^

Although our visit focused on the COHORT trial, the Davis COE is currently involved with several other HD clinical research trials. The Predict HD trial enrolls subjects who are gene positive for HD but have not yet developed symptoms. One defining feature of this trial is very lengthy neuropsychological testing administered by a trained physician as well as volumetric brain scanning. The goal is to search prospectively for early neuropsychological changes that occur before any motor changes.

Another study in which the Davis COE participates is PHAROS. This study enrolls subjects who are at risk for HD but have never been tested. The study is generally looking for precursors to HD phenotype expression and includes a heightened emphasis on mood evaluation.

For more about these studies, as well as other currently active HD trials, visit the Huntington Study Group here.

The UC Davis Center of Excellence typically conducts clinical research trials like COHORT and the ones described above once or twice a week at the VA Hospital in Rancho Cordova. The exact amount of time spent at the VA Hospital depends on how many trials are ongoing. During the rest of the week, the center is active in Sacramento at the UC Davis Medical Center, where they run an HD clinic and a Parkinson’s clinic.

The Collaborative Nature of HD Research^

During our visit, one point that came up again and again was the collaborative nature of HD research. From scientists conducting basic research, to doctors running clinical trials, the HD community is engaged in a cooperative venture. Information is shared freely between researchers and frequent communication allows for a coordinated research effort. This is a unique atmosphere in the world of academic research and it can be partly attributed to the strong, dedicated presence of the HD community. Researchers understand the stakes in the battle against HD and realize that the importance of their work demands the best possible effort. Cooperation is a key part of that effort.

A. Hepworth and A. Frohnmayer, 8-1-08

The Huntington's Disease Pipeline

Have you ever wondered how new medicines are discovered? The process of going from an important discovery in a science laboratory to a successful drug available to the public is complex, costly, and time-consuming. This process is often referred to as Research and Development, or the R&D pipeline. It has many stages that interact and feedback into one another, takes many years, and costs millions of dollars.

In a way, R&D is like searching for two separate needles in two separate haystacks. The process begins with a thorough search and identification of all the different biological pathways that are involved in the disease process. The goal is to identify one or more pathways that contain possible biological targets for treating the disease. After finding one or more targets, the next step is to find potential drugs that block or change how the target causes or contributes to disease; this can be done by either designing a drug based on knowledge of the target’s molecular structure and function, or by screening large libraries of chemicals and molecular compounds to find one.

A potential drug treatment may emerge from these extensive searches. However, this “candidate” drug is not yet available for treating patients. First, it must journey through more research, tests, and clinical trials before it is finally approved by the Federal government’s Food and Drug Administration (FDA). Only then, many years later (and after spending a lot of money!), might a new medicine be available to help patients with debilitating diseases. For every attempt, the probability of success is less than 1%. But thousands of scientists around the world are working hard to beat those odds every day, discovering new biological pathways to target and new chemicals to try for all sorts of diseases.


Let’s take a closer look at the steps of the R&D pipeline. Below is a brief overview of each stage, and you can read other sections of this article to get a more comprehensive look at that stage of the process and how it applies to HD research.

Basic Research:
Before scientists can even begin to think about finding treatments for a disease, they must have a good understanding of the biology and chemistry involved in the disease’s molecular pathways. Basic research tries to understand how certain biological and chemical imbalances cause disease. Scientists must investigate everything, from the genes and proteins involved, to clinical symptoms and disease progression.

Target Identification and Validation:
While gaining a thorough understanding of a disease through basic research, scientists also attempt to identify biological targets- usually genes or proteins – that trigger or contribute to a disease. A biological target is a good place to start looking for a treatment. There are numerous genes and proteins involved in most diseases- so how can scientists know if they have picked the right one to target? Researchers generally perform experiments on animal models to assess whether the biological target they have identified is crucial to the disease pathway.

Drug Optimization:
The next step is to find one or more drugs that might make a useful treatment. One way to find candidates is to search through, or screen, thousands of chemical compounds to identify potential drugs. Alternatively, if enough is known about the shape and function of the biological target, scientists may be able to design new molecules or drugs that change the way the target behaves in living cells and patients. As opposed to screening, this approach is often called “rational drug design.” A potential drug must interact with the chosen biological target, and modify it in a way that will cure the disease or decrease its effects.

Drug Development/ Pre-Clinical Animal Studies:
When a potential drug is discovered from a screen or rational drug design, it has much promise as a therapeutic drug. However, little is known about the safety and effectiveness of this so-called lead compound in animals or humans. All lead compounds must be thoroughly tested in at least two animal models to determine safe doses, understand side effects, and discover more about long-term toxicity.

Investigational New Drug (IND) Application:
After animal model studies have been completed, pharmaceutical companies must submit an IND application to the Food and Drug Administration (FDA) to continue drug development. If approved, it gives the pharmaceutical company permission to begin clinical trials which involve testing the potential drug in human participants. The IND application must include data from animal studies, information about the drug’s production, and a detailed proposal for human clinical trials. Unless the FDA specifically objects, an IND application is automatically accepted after 30 days and clinical trials can begin.

Clinical Trials: Phase I
Phase I clinical trials are also called “First-In-Man” studies. These studies are mostly concerned with safety, determining the maximum tolerated dose (MTD) of the drug in healthy volunteers that do not have the disease of interest. They also look for how the drug should be delivered (for example, by pill, injection, or IV fluid), how it is absorbed into different organs, how it is excreted from the body, and if it has any side effects. Approximately, 70% of potential drugs make it through this initial stage of testing.

Clinical Trials: Phase II
Once a safe dose of the potential drug has been found in healthy human participants, phase II clinical trials test it in participants with the disease of interest. Like phase I trials, these trials look for evidence of toxicity, and side effects, but they also look for evidence that the drug helps patients with the disease feel better in some way. These trials help to further refine the optimal dosage, in order to maximize beneficial effects and minimize harmful side effects.

Clinical Trials: Phase III
Phase III clinical studies are the most expensive, time consuming, and complex trials to design and run. They use a very large participant group, all of whom have the relevant disease condition. These studies determine if the drug’s benefits outweigh the risks (like side-effects and long term toxicity) in a larger patient group, and also compares the new potential drug with older, more commonly used treatments if any are already on the market.

New Drug Application (NDA):
The FDA must review results at the end of each phase to approve the drug before it can move into the next phase of trials. Once phase I, II, and III trials have all been successfully completed, the pharmaceutical company submits a New Drug Application (NDA) to the FDA. The results of all of the trials are given, as well as the results of animal studies, manufacturing procedures, formulation details, and shelf life. In short, they include everything that would be used to label the drug. If the FDA approves a company’s NDA, they can begin to mass produce and market the drug in the US. Pharmaceutical companies submit similar applications to authorities in Europe, Australia and elsewhere to market successful treatments there.

Clinical Trials: Phase IV:
Phase IV trials are done after a drug has been approved, officially manufactured, and put on the market. They are done to determine the absolute optimal dosages in case it needs to be refined, to look at the very long term safety and efficacy of a drug, and to discover any rare side-effects. Phase IV trials are also used to identify the potential for new indication studies, meaning that the same drug may be able to treat diseases other than the one for which it was initially tested.

Basic Research^

Basic research is concerned with understanding the background biology and chemistry underlying the mechanism of a disease. Scientists identify the genes, proteins, and specific types of cells involved in a disease and investigate how they contribute to the disease state. This type of science research is usually conducted in academic laboratories and research institutes around the world, and is less likely to take place in pharmaceutical companies. It is important to understand that basic science researchers focus their efforts on understanding the pathways and molecules involved in the disease- they don’t concentrate on finding a treatment. This information is then used by scientists in other research fields, such as drug discovery and therapeutics, to aid in the identification of drug targets and possible disease cures.

Usually, a research laboratory will concentrate on a few molecules or proteins in a disease pathway and try to discover as much as they can about those molecules. They may study the molecular structure of the molecule, how it usually functions in normal cells, and figure out what other proteins and genes it is related to or interacts with. Scientists will look at how their molecule (or molecules) of choice changes in diseased cells – if it works too much, stops working, or starts interacting with parts of the cell in a way that causes damage. All of this basic research helps to determine whether this molecule should be investigated as a biological target- the next step in the R&D pipeline. Although not all basic research directly contributes to a drug treatment, it all has the potential to, and so it is important to continue to strongly support basic science research programs to provide a solid foundation for drug development research.

Scientists often use model systems to study the genes and proteins involved in a disease. These can be in vitro, which is a useful way to isolate specific molecules and determine if they interact with one another in any way. Model systems can also be in vivo, which are more useful for studying specific molecules in the context of a disease, to see how they change a cell. In vivo model systems are also used to study the progression and development of a disease throughout an animal’s lifetime. The most common kinds of models include mice, yeast cells, worms, flies, rats, and human tissue culture.

HD and Basic Research

The HD community is doing a lot of basic research. We have much of this information located elsewhere on the HOPES site.

  • For more information on the kinds of institutions and programs that do HD research, click here.
  • For information on HD research going on here at Stanford University, click here.
  • To find out about the techniques that scientists use in basic HD research, click here.
  • To get an in-depth look at the day-to-day workings of a few research laboratories that conduct basic HD research click here.

Target Identification and Validation^

Basic research on the genes, proteins, and molecular pathways involved in a disease, may assist in the discovery of a biological target. Since there are many genes and proteins involved in most disease pathways, there needs to be a way to identify which ones would be worth targeting for creating a drug therapy. A biological target is a molecule that may hold the key to a disease- it may greatly contribute to, or possibly be the direct cause of a disease.

Validation of the molecule as a biological target usually requires answering two questions. The first asks if the target is relevant to the disease, by examining if a change in the biological target results in a change in the disease. If a certain molecule is produced in a mutated form, in abnormally high quantities, or abnormally low quantities in a disease, it is usually a good biological target. Secondly, if a biological target is proven relevant to a disease, it is then important to determine whether it is drugable – that is, can it be targeted or changed by treatment with a drug.

It is important to remember that validation occurs throughout many stages of the R&D pipeline, including the basic science research, the drug discovery, and the development processes. What may appear effective in a tissue culture model may not work in a mouse model. In each stage of testing or