Arches. Photo by Daniel Chia
Jun
29
2010

Mithramycin

control

Drug Summary: Mithramycin (also known as MIT and plicamycin) is an antibiotic that binds to DNA to regulate transcription. It attaches to specific regions of DNA that are rich in guanine and cytosine. While it is currently prescribed for the treatment of certain types of cancer and a few other conditions, recent research shows that it is helpful in treating motor symptoms and prolonging life in a mouse model of HD.

How could Mithramycin treat HD?^

Normally in the course of Huntington’s disease certain genes are prevented from being expressed in their normal protein products. This abnormal repression of genes is referred to as transcriptional dysregulation. Remember that “transcription” is the process by which the information of DNA is copied into messengers that are then used as templates for protein synthesis. Many of the genes that are prevented from being expressed are important for nerve cell health and survival. So, when these genes are blocked from producing their respective proteins, they cannot help to prevent the neurodegeneration that is typical of HD. This repression is caused by the mutant huntingtin protein, which interacts with molecules that would normally aid in the transcription of these helpful genes. The proteins that are encoded by the repressed genes have a wide variety of functions, which may explain why there are so many different symptoms associated with HD. One possible way to prevent many of these symptoms is to restore normal transcription, which is the proposed function of the drug mithramycin.

How does Mithramycin work?^

A number of hypotheses exist for how mithramycin acts in the body, but a group of researchers recently discovered what may be the key mechanism by which it prevents gene repression. One way that genes are repressed, or “silenced,” is through a process called methylation. Before explaining exactly what methylation is, we will first review how DNA is organized.

The entire DNA code is extremely long, and in order to be able to fit it into each cell, it needs to be very tightly compacted into chromosomes. (For more information on chromosomes, click here.) To accomplish this compression, the DNA is wound around structures called nucleosomes. You can think of a nucleosome as a spool and the DNA as the thread. Nucleosomes are made up of smaller proteins called histones. There are four different core histones (H2A, H2B, H3, and H4), and two of each are present in every nucleosome, along with one helper histone (H1). Histones play a very important role in the regulation of transcription.

By controlling access to DNA, histones determine if and when transcription occurs. When the histones keep the DNA tightly wound up, transcription factors cannot access the DNA, and it therefore cannot be transcribed. Different chemical groups (a chemical group can be a single atom or a small molecule) can be added to the histones in specific spots, causing the DNA to coil up to prevent transcription or causing the DNA to uncoil to allow transcription.

One way to control transcription is by methylation. In methylation, a chemical group called a methyl group is added to histone H3 or H4 at specific spots. When a histone is methylated at one of these spots, a protein called HP1 recognizes this signal and binds to the methyl group. These HP1 proteins also recognize each other and bind together, winding the DNA up into the coiled form of chromosomes in the process. Remember that in this coiled form, transcription factors cannot bind to the DNA so transcription cannot occur: the genes have been silenced. Methylation is a fairly permanent way of controlling transcription, so it can be devastating for an important section of the DNA to be wrongly methylated.

According to an important hypothesis, mithramycin helps restore normal transcription by regulating methylation. Researchers have discovered in a mouse model of HD that histone H3 was hypermethylated (methylated more that usual) at its ninth amino acid (recall that histones are proteins made up of amino acids in a long string or “chain”). It is already well known that increased methylation at this particular spot affects transcription. After conducting several experiments, the researchers concluded that mithramycin exerts its neuroprotective effects by preventing this hypermethylation and restoring normal transcription. Exactly how mithramycin prevents the hypermethylation has yet to be decisively determined, but the researchers have proposed a likely explanation.

Mithramycin is known to bind to areas of DNA that are rich in guanine (G) and cytosine (C), two DNA bases. (For more information on DNA bases, click here.) By binding to a GC-rich section of DNA, mithramycin likely prevents another molecule, which plays a role in methylation, from binding. Such molecules that aid in methylation could be either transcription factors or a type of enzyme called histone methyltransferase (HMT). HMT adds methyl groups to histones, causing the DNA to coil up as noted earlier and make it inaccessible for transcription. Transcription factors can also play a role in gene silencing by recruiting HMT. One type of transcription factor (TF) is already known to recruit an HMT that specifically binds to histone H3 at the ninth amino acid. These transcription factors bind to the DNA and recruit a specific HMT. The HMT then methylates the histone, preventing transcription. While a transcription factor that binds to GC-rich areas of DNA has yet to be found, researchers hypothesize that if there is such a molecule, mithramycin displaces it, preventing methylation. By preventing hypermethylation, mithramycin restores normal transcription, preventing much of the neurodegeneration typical of HD in the mouse model.

Research on Mithramycin^

Ferrante, et al. (2004) tested the effects of mithramycin on a transgenic mouse model of HD. The researchers daily injected one group of mice with one of five different doses of mithramycin and another group of mice with a placebo to serve as a comparison group. The mice were tested for body weight twice a week, motor performance once a week at first and later twice a week, and observed twice a day for survival. The researchers found that the dose of mithramycin influenced how long the mice lived. The benefits of mithramycin peaked at a dose of 150 micrograms per kilogram per day, since lower doses were less effective and higher doses were not well tolerated (and even resulted in death). The optimal dose of mithramycin led to the longest extension of survival ever seen in the HD transgenic mouse, extending survival by 29.1%. While this finding is very encouraging, we must remember that the experiment was done on mice and issues with drug safety and tolerability may prevent mithramycin from being so effective in humans.

In addition to their longer lives, the mithramycin-treated transgenic mice also performed better than the placebo-treated mice on the motor performance test each time they were tested. Motor performance was tested using the “rotarod” apparatus, which is a rotating rod on which the mice are placed and timed for how long they can stay on. The total motor improvement over placebo-treated mice was 42.6%. Mithramycin did not appear to affect the body weight of the transgenic mice.

When HD in the transgenic mice had become so advanced that they were no longer able to feed or move when prodded, they were euthanized and their brains examined. Amazingly, the researchers found that the mice that were treated with mithramycin had almost none of the typical brain deterioration seen in HD. The mithramycin-treated mice did not exhibit brain atrophy, enlarged ventricles, or loss of nerve cells in the striatum, which are typical symptoms of HD in both mice and humans. (For more information on HD and the brain, click here.) While the placebo-treated mice had a 21.3% reduction in brain weight, those treated with mithramycin experienced only a 2.8% reduction in brain weight. Also, the size of the nerve cells in placebo-treated mice decreased by 41.9%, while the mithramycin-treated mice did not have any significant decrease in nerve cell size compared to non-HD mice. These results show that mithramycin improves survival and is neuroprotective, since fewer nerve cells in the brain die and they remain at the normal size.

Once the researchers determined the effectiveness of mithramycin in treating HD mice, they tested several different hypotheses to find out how exactly the drug works. These experiments ruled out several mechanisms. They found out that mithramycin does not reduce the amount that the HD allele is transcribed (which would result in less of the harmful huntingtin protein); it does not change the amount of glutamate receptors or their activity (which would reduce the amount of excitotoxicity); and it does not change the permeability of mitochondria (which would reduce programmed cell death). The final hypothesis left was that mithramycin prevents the mutant huntingtin protein from interfering with transcription of specific genes that are important for nerve cell survival. There are several ways that mithramycin could restore normal transcription, and the researchers determined that it was by preventing methylation at a specific spot of histone H3 (as explained earlier). By preventing too much methylation, mithramycin allows genes to be expressed that promote survival in the presence of mutant huntingtin.

This initial study of mithramycin on the HD transgenic mouse shows very promising results. While the drug is already approved by the Food and Drug Administration (FDA) for the treatment of cancer and other diseases, it is too early to tell if mithramycin will be useful in treating people with HD. Since HD is a chronic condition, it important to determine whether a potential treatment can safely be used for long periods of time. Unfortunately, mithramycin is not well-tolerated in people at the typical dose for long-term use. In fact, the typical human dose is 25-30 micrograms per kilogram per day (recall that the optimal dose given to mice was much larger at 150 micrograms per kilogram per day) and is only given up to ten days. Mithramycin has been shown to cause unpleasant side effects, including nausea and vomiting, and long-term use has been shown to lead to excessive bleeding. Research on humans with HD will have to be conducted to test the efficacy of mithramycin given at non-continuous doses and/or smaller doses.

For further reading^

  1. “Antitumor Medicine Improves HD in Mice.” E-MOVE – Research News in Movement Disorders. 12 Jan 2005. Online.
    This is a short article of low difficulty that summarizes Ferrante et al.’s findings.
  2. Ferrante, et al. “Chemotherapy for the Brain: The Antitumor Antibiotic Mithramycin Prolongs Survival in a Mouse Model of Huntington’s Disease.” The Journal of Neuroscience. 2004; 24(46): 10335-10342.
    This is a very technical, scientific article. It details the experiment where mithramycin was given to HD mice and discusses the drug’s likely mechanism of action.
  3. “Mithramycin.” Online.
    This is an excellent summary of the drug mithramycin and Ferrante et al.’s findings. It is of medium to high difficulty.
  4. Zhang and Reinberg. “Transcription regulation by histone methylation: interplay between different covalent modifications of the core histone tails.” Genes & Development. 2001; 15(18): 2343-2360. Online.
    This is an extremely technical article that goes into detail about transcriptional regulation and histone methylation. It is only recommended to the most scientifically literate audiences.

-K. Taub, 8-10-05