Neurturin and its receptor can be found in the striatum, the region of the brain that is greatly affected by HD (Click here to see our section on the effects of HD on striatal neurons). One in vivo study compared the protective effects of neurturin and GDNF (the namesake NTF of the GDNF family) by engineering cells to serve as NTF production factories and grafting, or transplanting, these cells into mice. These mice were then given injections of chemicals intended to mimic the excitotoxic model of HD (Click here to see our section on the excitotoxic model). Neurturin was not only more effective than GDNF at rescuing a specific type of striatal neurons, but the former NTF also reduced the extent of neuronal damage caused by excitotoxic damage. Interestingly, the study found that neurturin and GDNF interacted with striatal neurons in different ways, suggesting that these factors may work together to protect these neurons. Indeed, GDNF has been found to be more effective than neurturin at protecting certain populations of striatal interneurons, nerve cells that connect afferent neurons (those that carry sensory information to the brain) and efferent neurons (those that carry nerve impulses away from the brain). Future research may look at ways of combining different NTFs to more effectively preserve damaged neurons.
Can neurturin one day be used to treat human patients with neurodegenerative diseases?^
The therapeutic application of neurturin is currently being investigated in a series of clinical trials run by the drug company Ceregene. A major challenge to the therapeutic use of neurturin and other NTFs is figuring out how to sustainably deliver these compounds into the brain. Because NTFs do not cross the blood-brain barrier, they cannot be administered orally. One proposed method has been the use of viral vectors to deliver a gene engineered to over-express neurturin into the striatum (For more information on viral vectors, click here). These genes can be thought of as neuturin factories, designed to increase the levels of neuturin produced by these cells. Once introduced, viral vectors with these genes have been shown to consistently and selectively deliver neurturin to dying neurons in cultures. Scientists at Ceregene have demonstrated that the viral vector delivery of neuturin (trade name: CERE-120) protected damaged neurons in mice and monkey models of Parkinson’s disease. Based on these results, CERE-120 for Parkinson’s disease is currently being evaluated in Phase II clinical trials. However, recent results have not been encouraging—patients treated with CERE-120 failed to show significant improvements over those who did not receive treatment. As a result, Ceregene is currently evaluating their future plans for CERE-120.
CERE-120 has also been proposed as a potential treatment for HD. The administration of CERE-120 to mouse models of HD showed evidence of both structural and functional protection of nerve cells—the mice not only showed decreased rates of neuron death, but also exhibited improved motor control. Positive results have been observed both in transgenic HD rodents, as well as rodents chemically induced to show symptoms of HD. The use of CERE-120 in humans to treat HD is currently being evaluated in pre-clinical development. Updates on the progress of CERE-120 will be added to this page as necessary.
Gasmi, M., Brandon, E.P., Hergoz, C.D. et al. (2007) AAV2-mediated delivery of human neurturin to the rat nigrostriatal system: Long-term efficacy and tolerability of CERE-120 for Parkinson’s disease. Neurobiology of Disease 27: 67-70.
Heuckeroth, R., Enomoto, H., Grider, J., et al. (1999) Gene targeting reveals a critical role of neurturin in the development and maintenance of enteric, sensory, and parasympathetic neurons.Neuron 22(2): 253-263.
This article seeks to characterize the normal functions of neurturin by examining mice incapable of producing this NTF. Although the language is technical at times, the article is pretty easy to understand.
Kordower, J.H., Hergoz, C.D., Dass, Biplob, et al. (2006) Delivery of neurturin by AAV2 (CERE-120)-mediated gene transfer provides structural and functional neuroprotection and neurorestoration in MPTP-treated monkeys.Annals of neurology 60: 706-715.
This technical article examines the effectiveness of CERE-120 in monkey models of PD. New treatments are usually tested on monkeys before they go into clinical trials.
Within this web site, the term "cause" is synonymous with "explanation". That is, a cause is an explanation, or a partial explanation, of why something (such as HD) occurs.
Diseases are complex processes, and individuals with HD have many diverse experiences with the disease. Just as there is no single "HD experience" (exact age of onset, severity of symptoms, etc.), there is no single cause of HD. Despite the fact that a single gene is the major factor determining whether or not an individual will get HD, other factors play a role in an individual's particular disease experience. Although genes predispose individuals to diseases, it is the combination of genes and environment (including the internal environment of an individual's body) that leads to the particular disease experience of an individual. Thus, there is not one cause of HD; instead, each person's experience with the disease has many explanations.
The cerebellum is known to be the place in the brain where learned movements are stored. Because of this, it has a large amount of control over the coordination of movements. But exactly how is this control exerted? First, the cerebellum receives input from various other parts of the brain (and spinal cord). One such brain part is called the inferior olive, which itself receives sensory information from many parts of the brain and spinal cord. It then relays this info to the cerebellum. In the cerebellum, the data are analyzed and a course of action is quickly decided. To put this action into play requires the output of information from the cerebellum and this is where specialized nerve cells called Purkinje cells become very important. Each and every piece of information that leaves the cerebellum does so through the Purkinje cells. Hence, these cells have a great deal of control over the refinement of motor activities. (See Figure F-8.)
Given this mechanism for the cerebellum’s control over movement, it’s no wonder that when there is severe damage to Purkinje cells and nerve cells in the inferior olive, the symptoms of ataxia (including the aforementioned loss of coordination and difficulty with speech) begin to show.
In all people, the three-letter codon sequence C-A-G is repeated several times at one end of the Huntington gene. In people with HD, the Huntington gene has an increased number of CAG repeats. Thus, there are different versions or alleles of the Huntington gene, one for each different number of CAG repeats. For simplicity within this website, the term "non-HD allele" is used to refer to any allele of the Huntington gene containing a number of CAG repeats within the normal range (10-35). The term "HD allele" is used to refer to any allele of the Huntington gene with an extended number of CAG repeats.
Correlation and causality are ways to describe the relationship between two events. If two events are correlated, then they usually occur together. For instance, people with 40 or more CAG repeats usually develop HD. People with 35 or fewer repeat numbers usually do not develop HD. These instances are examples of correlated events. Correlation also implies that two events change in a systematic way. For example, a negative correlation between two things implies that as one increases the other usually decreases. Hence, as the number of CAG repeats increases, the age of onset decreases. In contrast, a positive correlation between two things implies that as one increases the other increases as well. The age and height of children are positively correlated. Hence, older children are usually taller than younger children.
On the other hand, causality describes the cause and effect relationship between two events. The observation that two events are correlated is not enough to conclude that one causes the other to happen, nor is it enough to conclude that one doesn't cause the other to happen. In other words, correlation does not imply anything about causality. Correlation shows that two events are related, but it does not determine their cause and effect relationship.
For example, many psychological studies have shown that children who watch violent television shows are more likely to exhibit violent behavior. The general trend shows that a child's "level" of violence is positively correlated to the amount of violent television programs a child has seen. That is, the more violent TV shows a child watches, the more violent behavior he or she is likely to exhibit. This information does not prove that watching the television shows actually causes the children to become violent.
In this example the two events, viewing violent television and exhibiting violent behavior, are correlated. Hence, if a child is violent, it is very likely that he or she has also watched violent television. However, the violent television itself is not necessarily what caused the violent behavior. The child could exhibit violent behavior for any number of reasons. For example, it is possible that children who behave violently for other reasons are especially fond of watching violent television. The correlation between the two events is just not enough information to conclude anything about cause and effect. Thus, violent behavior is correlated to viewing violent television, but not necessarily caused by it.
In the case of HD, we know that the number of CAG repeats is negatively correlated to the age of onset. Usually, people with more repeats have an earlier age of onset. The important point to remember is that we do not know whether the additional CAG repeats are the actual cause of the earlier appearance of symptoms.
This chapter explains some of the many different types of research that scientists use to study Huntington’s Disease.
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.
Click here to learn about the basics of HD mouse models
Click here to view an article on the ethics of HD testing
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 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 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 “markertrait,” such as the same color eyes or the same blood type, then the genes that code for that markertrait 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 (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.
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.
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.
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.
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.
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 huntingtingene 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 huntingtinprotein and exhibited an abnormal phenotype. The level of mutant huntingtinprotein 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-inmodels 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-inmodels 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-inmodels tended to have many more CAG repeats than the earlier models (from 94-140 CAG repeats).
Knock-inmodels 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?
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.
Transgenic Animals This webpage maintained by a retired biology professor provides a quick introduction to the methods of creating transgenic animals.
On June 8th, 2008, the Huntington’s Disease Society of American (HDSA) honored HOPES with the first annual “Giving a Voice to HD” award. According to the HDSA, the award recognizes “an individual or group whose efforts have significantly helped to raise awareness about Huntington’s disease in their local community and beyond.”
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.
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
Requires slight assistance
Requires major assistance
Home or extended care facility
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:
the ability of the patient to follow a finger with the eyes in both the horizontal and vertical directions
the ability of the patient to turn the head in both the horizontal and vertical directions
the speed at which the patient is able to turn the head both horizontally and vertically
the presence of speech that is slurred, slow, and difficult to understand
the ability to stick out the tongue and the speed to which the task is completed
the ability to tap the fingers of both hands (15 repetitions in 5 seconds is considered normal)
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
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
slowness in initiation and continuation of movements
abnormal muscle tone (measured separately in the extremities, face, and trunk)
involuntary jerky movements of the body (measured separately in the extremities, face, and trunk)
walking with normal posture
the ability to walk in a straight line from heel to toe. The ability to do so regularly for 10 steps is considered normal
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:
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.
High risk for falls
Medium risk for falls
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:
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.
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.
When Bryan was younger, he used to love playing baseball with his dad. Every day after school, Bryan’s dad pitched to him and helped him become a very good hitter.
Almost every time Bryan’s friends came over to play, his dad would go outside and play with them. He would throw passes to them when they were playing football and he would shoot baskets with them when they were playing basketball. All of Bryan’s friends liked Bryan’s dad because he was so much fun to play with!
One summer, Bryan’s dad started to act a little strangely. When he walked, his hips moved around almost like he was dancing. When they played baseball, he began to throw pitches that missed the plate a lot (he used to throw perfect strikes). When Bryan’s friends came over, his dad could not throw passes or shoot baskets as well as he had before. Soon, his dad stopped playing with them.
Bryan was sad and confused about why his dad was having so much trouble moving around. He became even more concerned when he noticed that his dad was treating his mom differently. Bryan’s dad had always been very nice, but now he began to start arguments with Bryan’s mom and he sometimes said mean things.
Soon, Bryan returned to school. One day when his dad came to take Bryan home, Bryan’s friend Lisa said, “Your dad walks funny!” This comment made Bryan sad and angry because nobody had ever said anything bad about his dad before. Bryan was not just upset at Lisa, but also at his dad for becoming so different.
On the way home, Bryan’s dad noticed that his son was upset. He asked Bryan what was wrong and Bryan replied, “What’s going on with you, Dad? You don’t play sports as well as you used to, you walk really weird, and you haven’t been very nice to Mom lately. Why are you being like this?”
Bryan’s dad had noticed these changes in himself, too, and he told Bryan that he did not know why he was acting this way. “Actually,” he said, “your mom and I are going to see the doctor today to find out what’s going on. You are welcome to come along if you want.” Bryan never liked going to the doctor’s office, but he decided to go with his parents this time because he really wanted to know why his dad was acting so strangely.
When they got to the doctor’s office, the doctor asked, “So what seems to be the problem?”
Before his dad could speak, Bryan’s mother said to the doctor, “Well, my husband has been acting strangely the past few months. His body seems to shake when he walks and he has also been kind of grouchy.”
Bryan added, “We used to play sports together all the time, but lately he hasn’t been throwing as well as he used to and he even stopped playing with me and my friends.” Bryan’s dad looked sad when he heard his family say these things, but he knew there was truth in what they said.
The doctor replied, “There are a lot of different things that can cause these kinds of problems, so I will need to find out some more information.” She went on to ask Bryan’s dad about many different things, including how old he was and if any of his family members ever had a disease.
Bryan’s dad replied, “Well, I am 35 years old and my mom passed away when I was 27 from something called Huntington’s disease.”
Then Bryan heard his dad say this, he asked the doctor, “What is Huntington’s disease?”
The doctor replied, “Well Bryan, Huntington’s disease is a disease that can cause a lot of changes in the way someone moves, thinks, and feels emotions. These changes start to show up usually when the person is between 30 and 50 years old. The disease is generally passed down through families. Just like when you flip a coin and you have a 50% chance of the coin landing on ‘tails,’ a parent who has Huntington’s disease has a 50% chance of passing it on to his or her child. Both men and women can get the disease and it occurs all around the world, although it is generally very rare.”
Bryan’s dad asked, “So how do these changes come about?”
The doctor replied, “Well, the brain is made up of millions of small living cells called nerve cells and these basically allow us to do everything we do, from talking and reading to walking and even throwing a baseball. In people with Huntington’s disease, many nerve cells are damaged in certain parts of their brain called the basal ganglia. Healthy nerve cells in these parts of the brain help with making the smooth movements that we normally make when we do things like walking or playing catch. So when these nerve cells are damaged, this can lead to a lot of problems with movement.”
Bryan asked, “What is wrong with the nerve cells? Why are they damaged?”
The doctor replied, “The problem actually exists down within the nerve cells in tiny, microscopic structures called genes. The word sounds the same as the jeans you wear on your legs, but these genes inside the cells work in a completely different way. They contain a special kind of information in the form of DNA. Have you heard about DNA?”
“Only a little, in the news and at school.” said Bryan. “I don’t really understand it. What does it do?”
“Well,” said the doctor, “DNA is chemical stuff in the body’s cells that carries information for sustaining life. Each cell in the body has DNA that contains many thousands of genes, but the problem in Huntington’s disease has to do with just one of these genes, one small piece of the DNA that is called the Huntington gene.”
“Like every other gene, the Huntington gene contains many small sections called codons, each made of DNA. You can think of it like a chain: the entire Huntington gene is like a long chain and each codon is like a different link in the chain. In people who do not get Huntington’s disease, there are 35 or fewer identical links in a key part of the chain. But in people who do get Huntington’s disease, there are 40 or more of these identical links in the key part. Huntington’s disease results from having too many identical codons in the key part of the Huntington gene.”
“So the chain is too long?” asked Bryan. “Does it take up too much space or something?”
The doctor replied, “Well, not exactly. The system works like this: each gene makes another chemical called a protein and proteins are the things that do much of the work in cells. The Huntington gene makes a protein that we call huntingtin. In people who do not get Huntington’s disease, the huntingtin protein comes from a version of the Huntington gene with 35 or fewer identical codons in the key part. This huntingtin protein is able to do its normal work in cells. But in people who do get Huntington’s disease, the huntingtin protein comes from a version of the Huntington gene with 40 or more identical codons in the key part. This causes the huntingtin protein to have a different shape than normal and because of this altered shape, the protein cannot do its normal job.”
“After enough time, huntingtin proteins with the altered shape form into clumps or aggregations down inside cells, and we think these clumps clog up cells. We also think that the huntingtin proteins in these clumps may grab onto other proteins and keep them from doing their jobs in cells. So, these altered huntingtin proteins create a big problem, especially for nerve cells in the basal ganglia regions of the brain.”
“Eventually, these altered huntingtin proteins cause enough damage that the nerve cells containing the clumps begin to die. When enough nerve cells die in the basal ganglia regions of the brain, people start to have problems doing the movements that they always did before. This can be followed by changes in the person’s personality. For instance, the person might become more sad at times or more grouchy.”
After the doctor finished, Bryan’s dad asked, “So do I definitely have this disease?”
The doctor replied, “I don’t know for sure because your attitude changes and problems with moving could be caused by something other than Huntington’s disease. But since you said that your mother had Huntington’s disease, this means that there is a 50% chance that you might have it as well, just like a flip of a coin.”
In order to see for sure if Bryan’s dad had Huntington’s disease or not, the doctor suggested that Bryan’s dad go to a genetic testing center where he could give some of his blood to have it tested for the disease. Bryan’s dad went the next week.
At the dinner table one night, Bryan’s dad told him that the results of the test had come back from the genetic testing center. He said, “Bryan, I don’t know how to tell you this, but the test that I took showed that I do have Huntington’s disease. That is the reason I have been acting so strangely.”
Bryan could tell that his parents were upset, but they were trying hard to be brave about this news. As his dad finished talking, Bryan saw that his mom was beginning to cry. Bryan was not used to seeing his mom cry. He was also scared to hear that his dad had Huntington’s disease. Bryan felt overwhelmed and this made him cry too.
Later that night when Bryan was going to bed, his parents came into his bedroom to tuck him in. Bryan’s dad said, “You know Bryan, this disease may cause a lot of changes in my behavior from now on. The doctor even says that these changes will get worse over time.”
Bryan’s mom added, “But even though some things your dad does might start to change, he will never stop loving you. We both love you very, very much and nothing about this disease is going to change that. There may be a lot of challenges for us in the future, but the most important thing is that we keep a positive attitude and keep loving each other.”
Although Bryan still felt a little sad, these words from his parents made him feel much better. “Keep a positive attitude and keep loving each other,” Bryan repeated to himself.
The next week at school, Bryan’s dad came to pick him up and Bryan’s friend Joe said, “Your dad walks funny!” This time Bryan knew the reason why his dad walked differently. He was no longer angry at his dad. He wasn’t even angry at Joe. He realized that it was not his dad’s fault for changing; it was the Huntington’s disease that was making his dad act differently from before. Bryan knew that it would be hard for his friends to understand why his dad was acting differently, because they did not know the things that he had learned about Huntington’s disease. Maybe someday he could teach them about the huntingtin protein and nerve cells, so that they would understand too. But for now, his main focus was to keep a positive attitude and not allow these comments to hurt him. He knew that he would be much happier this way.
“Yeah,” said Bryan, “My dad has a medical problem. But he’s a real sport about it, and I’m learning to be a sport about it too. We walk and run less these days, but we still have fun together doing new things.”
On the ride home, Bryan’s dad said, “Hey Bryan, I know I don’t throw the baseball as well as I used to, but I can sure catch fish. I was thinking that this weekend we could go down to lake and go fishing.”
“That sounds great, Dad!” Bryan said. He loved to go fishing, but he was even more excited by the chance to hang out with his dad.
Bryan and his dad were wonderful fishermen! They both reeled in six fish! As they were getting ready to go, Bryan’s dad said, “Wow Bryan, this has been a blast! Thank you so much for coming out here with me.”
Bryan quickly replied, “No Dad, thank you!”
Bryan realized that although his dad was changing and could no longer do some of the things he had done before, they could still do plenty of other fun things together. He knew that his dad’s Huntington’s disease would present tough problems in the future, but he loved his dad and he knew that his dad loved him. No matter what the future had in store, nothing could ever change that.
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HOPES is a team of faculty and undergraduate students at Stanford University dedicated to making scientific information about Huntington's disease (HD) more readily accessible to the public. Our goal is to survey the rapidly growing scientific literature on HD and to present this information in a web source. We seek to provide information about causes, symptoms and treatment of HD that reflects current scientific understanding of HD. To date, HOPES resources have reached out to families in over 47 countries.
We emphasize that we are neither medical professionals, nor affiliations of the researchers and laboratories mentioned on our pages. The information we present is intended for educational purposes only and should not be construed as offering diagnoses or recommendations. We operate as a not-for-profit public service organization, and our funding is entirely from private sources.
Meet the HOPES Team
Stephanie is a sophomore with a penchant for pretty things, paddling dragonboats, pushing pixels, the printed word, procrastination, and photography. She joined HOPES because she loves science, loves websites, loves people, and loves writing. Relatedly, her main life goals are to become the next Dr. Sanjay Gupta, and to spread the gospel of bubble tea around the world. Stephanie also enjoys alliteration, collecting Stuff that Looks Like Other Stuff, snacking when she ought to be sleeping, making lists, tweeting excessively, watching unintellectual television, changing her mind about what to major in, and speaking in third person.
Pooja Bakhai is a senior majoring in human biology with a focus in global health and infectious disease, and minoring in music. She is the President of Global Health Volunteers at Stanford, plays clarinet with the Stanford Philharmonic Orchestra, and is looking forward to researching the viral etiology of Chronic Fatigue Syndrome. Huntington's disease is present in her father's side of the family, and this has inspired her to work for HOPES. As a future doctor, Pooja aims to eliminate health disparities and work with underserved populations in the United States and around the world.
Amy is a student researcher who is thrilled to be joining HOPES. A member of the Stanford class of 2013, she plans on majoring in Biology, with an emphasis on molecular and cellular biology. She joined HOPES because she is very interested in the biological workings of Huntington's Disease and its treatments. She also thinks that compiling all the research on the forefront of HD into one website written to be accessible to everyone is a brilliant idea. Her favorite part of HOPES is getting to meet so many remarkable people who share her goals. In her free time, Amy enjoys dancing, writing for the paper, reading, and watching bad British television.
Aimee is having the time of her life during her third year at Stanford. She is a proud member of the class of 2012 and majoring in Human Biology. Aimee is a student researcher for HOPES. She decided to join HOPES because she likes the way HOPES serves the HD community and the potential number of people it reaches around the world. In her spare time, Aimee likes to read, run, write, play video games, and watch the sunset. Her long-time wish is to fly.
Yi is a student researcher and a proud member of the Stanford Class of 2011. He joined HOPES because of his desire to combine his interests in scientific outreach and education with his skills as a writer. His favorite part about HOPES so far has been the opportunity to meet so many engaging and inspiring people from the HD community. At Stanford, Yi is majoring in Anthropology with a minor in Modern Languages, although he really wishes that he could just stay in school forever and get a Bachelor's degree in everything. In his free time, he enjoys reading The New York Times, eating plain yogurt and thinking about how great it would be to run a marathon. He also hates beets. Boy does he hate beets.
Tiffany is a student researcher with HOPES. She is a Biology major hoping to go to medical school and member of the Stanford Class of 2010. Tiffany became interested in HOPES after working at an Alzheimer's Research lab, which prompted her to learn more about other neurodegenerative diseases. Since becoming a member of HOPES in 2008, Tiffany has found inspiration in working with the HD community and treasures every opportunity she has had to hear the stories of people who are affected by HD. As a frequent participant in science outreach activities, she sees HOPES as a way to pursue her academic interests while serving the HD community. When she is not working on pre-medical classes or HOPES, Tiffany enjoys dancing with her school jazz group Urban Styles, eating cereal and watching David Attenborough's Planet Earth documentaries.
Faren graduated in June 2009 with a BA in Human Biology but couldn't bear to leave the HOPES team behind. She stepped down from the position of Project Leader and is enjoying getting back to researching and writing articles. Her favorite job within HOPES is answering e-mail, so please send feedback and questions! In her free time, which she finds quite abundant post-college, Faren likes to cook, eat, and teach dance. Oh and study for the MCAT…
Morgan Thompson, Ph.D. – Editing Consultant
Danielle Simmons, Ph.D. – Editing Consultant
William Durham, Ph.D. – Faculty advisor, Bing Professor in Human Biology, Department of Anthropological Sciences
Past Team Members
Danny Neumann, Web Developer, B.S. in Geophysics
Andrew Pipathsouk – Researcher, B.S. in Mechanical Engineering
Chelsea Garnett – Researcher, B.A. in Human Biology
Jean Ansolabehere – Researcher, Senior in Human Biology and English
Natalie Justicz – Graphic Artist, Senior in Human Biology
Adam Hepworth – Project Leader & Researcher, Senior in Human Biology and Philosophy
Mesa Schumacher – Graphic Artist, B.A. in Anthropological Science
Amy Frohnmayer – Researcher, Psychology
Eric Whitney – Researcher, Human Biology
Jonathan Dyal – Project Leader & Researcher, B.A. in Human Biology
Justine Seidenfeld – Researcher, B.A. in Human Biology
Matt Woloszyn – Researcher, English
Alicia Follmar – Researcher, Human Biology
Aliyya Haque – Researcher, B.A. in Human Biology
Christina Chen – Project Leader & Researcher, B.A. in Human Biology, '07
Devon McGee – Project Leader & Researcher, B.A. in Human Biology, '06
Stanislav Jourin – Art Director, M.S. in Digital Imaging & Design at NYU, '04
Kim Taub – Project leader & Researcher, B.A. in Human Biology, '05
Taylor Altman – Researcher, B.A. in Human Biology
Jia Hou – Researcher, B.A. in Human Biology
Agnieszka Milczarek – Researcher, B.S. in Biology and B.A. in English
Leon Hsu – Researcher, B.A. in Human Biology, '05
Clare Tobin – Researcher, B.A. in Human Biology
Michael Morici – Researcher, B.S. in Biology
Tonkid Chantrasmi – Web developer, Graduate Student in Mechanical Engineering
Shawn Fu – Web developer & graphic artist, B.S. in Biology, '03
Amy Hsu – Project leader, B.S. in Mathematical and Computational Science, '02
Matt Stenerson – Project leader & researcher, B.A. in Human Biology, '03
Cathy Barnard – Researcher, B.A. in Human Biology, '04
Kaizad Cama – Researcher, B.A. in Human Biology, '03
Peter Chang – Researcher & web developer, B.S. in Biology '03
Darwin Chen – Researcher, B.A. in Human Biology, '02
Tucker Cunningham – Web developer & graphic artist, Senior in Computer Science
Jaclyn Czaja – Researcher, B.A. in Human Biology, '02
Karen Hammond – Researcher, B.A. in Psychology at Carleton College, '03
Kelvin Ho – Web developer & graphic artist, B.S. in Computer Science, '04
Tracy Ho – Researcher, Senior in Human Biology
Vinita Kailasanath – Researcher, B.A. in Human Biology and American Studies, '03
Shashikant Khandelwal – Web Developer, Graduate Student in Computer Science
Ruth Lo – Researcher, B.A. in Human Biology, '04
Jean-Gabriel Morard – Web developer, Graduate Student in Computer Science
Melissa Schapiro – Researcher, B.S. in Biology & Psychology, '04
Gina Schiel – Researcher, Senior in Human Biology
Elyn Tan – Researcher, B.A. in Human Biology, '03
Joakim Vinberg – Web developer and graphic artist, B.S. in Biotechnology, '04
Belinda Fu, M.D. – Faculty advisor, Lecturer, Department of Human Biology
Joanna Mountain, Ph.D. – Faculty advisor, Assistant Professor, Department of Anthropological Sciences and Department of Genetics
Ronald Barrett, R.N., Ph.D. – Faculty advisor, Assistant Professor in Medical Anthropology
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.
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.
The nucleus of a mammalian cell is enclosed by a nuclear envelope, a membrane that features many small openings or “pores.” (The nuclear membrane and its pores can be seen in Segment 4 of the “Basics of HD” video: click here to view that segment.) These pores allow different molecules to move back and forth between the nucleus and the cytoplasm. But these pores are very small and allow only smaller molecules to cross the nuclear envelope. Larger molecules require other, more complex mechanisms to be transported into the nucleus, and these mechanisms often take longer as well.
The alteredhuntingtin protein associated with HD normally resides in the cytoplasm of a nerve cell because it is too big to be able to easily cross the envelope into the nucleus. But when that intact protein is cut up into small fragments, those fragments can easily move into the nucleus and cause dangerous problems for the cell. Proteases are a family of proteins that break up other proteins into smaller pieces. Studies have shown that a specific group of proteases called caspases play a big role in cutting up altered huntingtin into small fragments that can move from the cytoplasm into the nucleus.
A recent study from the lab of Michael Hayden at the University of British Columbia has shown that a particular caspaseprotein, named caspase-6, may be responsible for the type of huntingtin fragments that lead to nerve cell death and symptoms in HD. In the study, scientists used a mouse model of HD and changed the alteredhuntingtin protein so that caspase-6 could no longer cut it into fragments. They found that these mice showed no evidence of nerve cell death and they never developed any symptoms of HD. This finding suggests that a drug that inhibits the activity of caspase-6 may be a treatment for HD.
Background on caspases and HD
Caspase-6 is not the only protein to cut up altered huntingtin into fragments. Previous studies have shown that caspases can be divided into three rough categories (for more on caspases, click here). There are “ICE-like” caspases (named for their similarity to another kind of protease called the interleukin-1b converting enzyme), “initiator” caspases, and “effector” caspases. ICE-like caspases include caspase-1, 4, and 5. These three seem to play a role in fragmenting proteins involved in processes like inflammation, rather than fragmenting the huntingtin protein. Initiator caspases include caspase-3, 7, and 2, and convert the inactive form of an effector caspase to an active form by cutting off one or two small fragments from the inactive effector caspase. The three effector caspases include caspase-6, 8, and 9, which are the caspases that (when activated by the initiator caspases) actually break down most other proteins.
While it was known for some time that both caspase-3 and caspase-6 break down huntingtin protein into fragments, it was not known if all of the resulting fragments enter the nucleus and cause nerve cell death. It seemed possible that the fragments that were particularly toxic to the nerve cell were specifically generated by one of the two caspases. So in their study, Hayden and co-workers used a mouse model of HD, and mutated the alteredhuntingtin protein so that either the caspase-3 or caspase-6 protein would not find its usual target. This involved changing the specific amino acid sequence that caspase-3 (or -6) usually targets, and only changing that part so that the rest of the huntingtin protein acts the same. Most of the target sites for caspases are only 4 amino acids long, so it is not difficult to selectively change that part.
Hayden’s group generated one mouse that had all four of the caspase-3 target sites changed and inactivated, one mouse that had the single caspase-6 target sites changed and inactivated, and one mouse that had all of the caspase-3 sites and the caspase-6 site changed and inactivated. They tested all of the types of mice to make sure that they were expressing similar amounts of the huntingtin protein, and that the expanded polyglutamine chains were roughly the same length. In so doing, the researchers ensured that the main difference between these mice was the ability for caspases-3 and 6 to fragment the huntingtin protein.
One way to test for nerve cell death is simply to measure the weight of the brain at a certain age in HD mice and compare it with the weight of the brain in other strains of mice. The less the brain weighs, the more you can assume there is nerve cell death. Previous studies have shown that mice with alteredhuntingtin protein (that can be targeted by both caspase-3 and 6) lose about 10% of their brain mass as compared to healthy, wild-type mice without the alteredhuntingtin protein. This loss of brain mass can be attributed to nerve cell death due to the HD associated protein. The first thing that the Hayden group observed was that the mice with alteredhuntingtin protein resistant to both caspase-3 and caspase-6 did not have that 10% loss of brain mass. Instead, the mice were much more similar to the healthy, wild-type mice.
Then, to determine which of the caspases—3, 6, or both—were necessary for brain mass loss, Hayden and coworkers tested each of the other two mouse lines they had generated. They found that the mice with huntingtin protein resistant to caspase-3 cleavage had similar brain mass loss as mice with the HD associated huntingtin protein. In other words, fragments generated by caspase-3 are not the fragments that cause nerve cell death. But these mice still generated fragments due to caspase-6.
Next, they tested the mice with huntingtin protein resistant to caspase-6 cleavage, and they found that these mice had no significant brain mass loss. They were similar to healthy, wild-type mice and to the mice that had huntingtin protein resistant to both caspase-3 and 6 cleavage. Notably, these mice were still generating fragments due to caspase-3. But since these mice had no evidence of brain mass loss, it is evident that fragments selectively generated by the action of caspase-6, but not caspase-3, are toxic and cause nerve cell death.
Additionally, Hayden and his group tested the motor coordination of each type of mouse. What they found was that both the mice resistant to caspase-6 action and the mice resistant to caspase-3 and -6 action, were able to perform normally, just like healthy, wild-type mice. The mice resistant to only caspase-3 action performed poorly, just like mice with the regular HD-associated huntingtin protein. This result shows that selective inhibition of caspase-6 not only prevents brain mass loss, it also prevents motor symptoms of HD.
Finally, they looked specifically at the location of fragments of huntingtin protein within the nerve cell. Mice with the HD-associated huntingtin protein and mice that have caspase-3 resistance (but generate fragments cut by caspase-6) both have fragments that enter the nucleus early in the mouse’s lifetime. Both healthy, wild-type mice and mice that are resistant to caspace-6 (but generate caspase-3 fragments), show little to no signs of huntingtin fragments entering the nucleus. In the caspase-6 resistant mice, researchers saw some fragments enter the cell very late in life, but they still did not cause nerve cell death or symptoms. This points to the idea that it is the action of fragments (created selectively by caspase-6) inside the nucleus that causes toxicity and nerve cell death. If fragments created selectively by caspase-6 are the ones to enter the nucleus, then caspase-6 inhibition might prevent that toxicity and might prevent HD symptoms.
Directions for the future
A few uncertainties remain to be considered in the Hayden Lab study. Most significantly, it is unknown whether altering the specific amino acid sites that caspase-3 and -6 target has any effect on the rest of the huntingtin protein itself. Perhaps in addition to being targets for caspases, those sites determine huntingtin structure, stability, or clearance. If so, we cannot know whether the lab’s findings on the role of caspase-6 would hold true in human patients. Furthermore, caspase-6 might have other important functions in the cell that an inhibitory drug would impede. Finally, we do not know if there are other caspases or caspase sites that play a significant role in creating the specific huntingtin fragments that lead to nerve cell death in humans. More work will have to be done to answer all of these questions and ensure that any caspase-6 inhibitors developed as drugs are safe and effective. This area of research will be important to watch for the next few years.
For further reading
Graham RK, et al. (2006). Cleavage at the caspase-6 site is required for neuronal dysfunction and degeneration due to mutant huntingtin.Cell. Jun 16;125(6):1179-91
This is the main paper discussed in this article: a fairly technical research paper.
Slow EJ, et al. (2003). Selective striatal neuronal loss in a YAC128 mouse model of Huntington disease. Hum Mol Genet. Jul 1;12(13):1555-67.
This paper discusses the creation of the mouse model used in the study described by Graham et al (2006).
Thornberry NA, et al. (1997). A combinatorial approach defines specificities of members of the caspase family and granzyme: Functional relationships established for key mediators of apoptosis. J Biol Chem. Jul 18;272(29):17907-11.
A more general review of caspases and their three different functions. Still technical, but more comprehensible.
Gutekunst CA, et al. (1999). Nuclear and neuropil aggregates in Huntington’s disease: relationship to neuropathology. J Neurosci. Apr 1;19(7):2522-34.
This paper is a technical but readable research article about where it is in the nerve cell that huntingtin protein and huntingtin fragments tend to localize.
Como et al. (1997) performed a randomized, double-blind study of 30 nondepressed patients with HD. 17 subjects received fluoxetine for 4 months, while 13 received a placebo. The study did not find that fluoxetine was helpful; patients receiving fluoxetine did not demonstrate improvements in motor or cognitivesymptoms, and did not improve in measures of functional capacity (the ability to perform day-to-day tasks. While this study suggests that fluoxetine is not helpful for nondepressed people with HD, the researchers had previously reported that treating 8 depressed HD patients with fluoxetine helped those patients deal with symptoms much better – so fluoxetine may be useful to treat depression in HD.
The first case study was on a 60-year-old woman who had symptoms of HD beginning in her mid-forties and symptoms of OCD beginning at age 25. She had not been successfully treated for her chorea, declining cognitive functioning, or aggressive behavior. Before treatment, her symptoms were so bad that her speech could not be understood and her cognitive functioning so impaired that she could not even take the MMSE. She was given the HDMRS, and her motor functioning scored 20 on a scale of 25 (with 0 as the least impaired, and 25 as the most impaired). She began treatment with fluoxetine, and after a month she was clearly less agitated and had a better mood. Her motor performance progressively improved during treatment, scoring 12 on the HDMRS after 4-6 months. The improvement in her motor functioning allowed her to begin walking again and speak coherently. Perhaps most surprising was her improvement in cognitive functioning. Cognitive improvement began after about 4-6 months of treatment, and after about a year she could take the MMSE and scored 12 out of 25. She continues to improve 6 years after beginning treatment with fluoxetine. Additionally, her movement became worse during the two periods in which she stopped taking the medication.
The second case study was on a 55-year-old woman who had symptoms of HD for the past 8 years. Her main symptom was the involuntary movements characteristic of HD, and she mostly retained her cognitive functioning. When tested before treatment began she received a score of 13 on the HDMRS and 19 on MMSE. She began treatment with fluoxetine and another drug to treat her insomnia (since fluoxetine was making the insomnia worse). She began to improve in her motor performance after about 2 months of treatment and reached the height of her improvement after 6 months, with a score of 8 on the HDMRS. She maintained this level of motor functioning for the next year and was able to return to her job. The patient did not change significantly in cognitive functioning, maintaining a score of 20 on the MMSE for as long as she was observed. She went off of the medication for a period of 3 months after a year of treatment and her motor performance deteriorated during this time. When she started taking fluoxetine again, she regained her previous level of motor functioning.
These two case studies show that fluoxetine may be beneficial to people with HD who have not responded well to other treatments for both behavioral and movement symptoms. The motor functioning probably improved as a result of increased serotonin signaling in the brain. It is unclear how the patient in Case One had such impressive cognitive improvement; this has never been seen before and may only be partially due to the beneficial effects of serotonin. It is possible that the reason why fluoxetine was so helpful in these two cases has to do with them both having a history of OCD in their families. In other words, a possible reason for success in these cases had to do with improvements in their OCD rather than, or in addition to, HD. It is important to note that this study only represents two cases of people with HD. Based on these cases, it seems that fluoxetine could potentially be beneficial to people with HD who also have a family history of OCD, however, more research needs to be done before making assessments about fluoxetine’s effect on people with HD.
Grote et al. (2005) studied fluoxetine in a mouse model of HD and found that fluoxetine might help fight some of the effects of the disease. R6/1 mice were either treated with fluoxetine or a placebo. Scientists found that there was no improvement in motor symptoms, but HD mice treated with fluoxetine had improvements in cognitivesymptoms; untreated HD mice tend to repeatedly explore the same paths in a maze, and the treated HD mice behaved more like normal mice by exploring the maze more thoroughly. Treated HD mice also showed fewer symptoms of depression than untreated HD mice.
The results were more than just behavioral; when the researchers looked at the brains of these mice, they found that fluoxetine reversed many of the problems HD causes in the brain. Treated HD mice had much larger dentate gyruses than untreated HD mice, and had an increase in neurogenesis.
Altogether, research results on fluoxetine are mixed; an animal study reports some improvement, while a small clinical trial does not.
For further reading
Como PG, Rubin AJ, O’Brien CF, Lawler K, Hickey C, Rubin AE, Henderson R, McDermott MP, McDermott M, Steinberg K, Shoulson I. A controlled trial of fluoxetine in nondepressed patients with Huntington’s disease. Mov Disord. 1997 May;12(3):397-401. This medium-difficulty study describes the clinical trial of fluoxetine
DeMarchi, et al. Fluoxetine in the treatment of Huntington’s disease. 2001. Psychopharmacology 153: 264-266. This is a scientific article of medium difficulty that describes two case studies of people with HD that were treated successfully with fluoxetine.
Duan, et al. (2004) noted that SSRIs are very helpful in treating psychiatricsymptoms in people with HD, yet no one had tested how they might affect neurodegeneration and the progression of the disease. These researchers created four different experimental groups of mice: transgenic (mouse models of HD) and nontransgenic mice that were injected with paroxetine, and transgenic and nontransgenic mice that were injected with a placebo. The progression of the disease was observed in the mice each day, and they were weighed each week. Their motor performance was tested by placing them on a rotating rod and recording the amount of time that they were able to stay on. Levels of different brain chemicals were measured, including serotonin and the related molecule 5-HIAA.
The mice received treatment (or placebo) starting at eight weeks of age. At 14 weeks it was found that HD mice had lower levels of serotonin and 5-HIAA in the striatum compared to the nontransgenic mice. Serotonin was increased in both transgenic and nontransgenic mice that received paroxetine. Furthermore, administration of serotonin did not affect the levels of any other important brain chemicals tested, such as dopamine.
Paroxetine also delayed the beginning of behavioral symptoms by an average of 2 weeks in the HD mice, and even helped them survive for an average of 15 days longer than the previous maximum life span (this is a significant amount of time in the life of a mouse). While weight loss is a problem both for HD mice and people with HD, paroxetine slowed the loss of weight in HD mice compared to untreated HD mice. At 16 weeks the HD mice treated with paroxetine performed significantly better than the untreated HD mice on the motor tests.
In order to assess the drug’s effect on the neurodegenerative process, the researchers examined the brains of HD mice and compared them to nontransgenic mice. While untreated HD mice showed deterioration of the brain with larger lateral ventricles and a thinner cerebral cortex, HD mice treated with paroxetine had less enlarged ventricles. Having less enlarged ventricles means that the HD mice treated with paroxetine lost fewer nerve cells in their brains. (For more information on HD and the brain, click here.)
This study suggests that paroxetine not only improves serotonin signaling in HD mice, but it also slows neurodegeneration and improves overall survival. This slowing of the neurodegenerative process may also help to slow the typical weight loss caused by HD. By using an SSRI such as paroxetine to increase serotonin in the brain, serotonin-induced signaling is increased, as is the expression of the important brain-derived neurotrophic factor (BDNF) . (For more information on BDNF, click here.) Additionally, this research found that paroxetine was helpful when given both before and after the onset of motor symptoms. This finding is important because it may indicate that paroxetine might still have beneficial effects even if it is given later in the course of the disease. SSRIs appear to be safe for long-term use in humans, so starting paroxetine early should not rule out its later use as well.
While these results are hopeful, it is important to remember that the study was conducted on mice made to look like they have HD, not on humans or people with HD themselves. Even if paroxetine is relatively safe for use in human use, it nevertheless may not help neurodegeneration in humans the same way that it is indicated in the mouse study. Overall, the use of paroxetine to treat both the symptoms and progression of HD is a promising idea that needs more investigation.
For further reading
Duan, et al. Paroxetine retards disease onset and progression in Huntington mutant mice. 2004. Annals of Neurology 55(4): 590-594. This is a scientific article that describes a study done on mice treated with paroxetine.
In order to understand how SSRIs increase the amount of serotonin signaling in the brain, we must first understand how neurotransmitters like serotonin work. Neurotransmitters are important molecules in the brain that help nerve cells communicate with each other. A message is passed within a nerve cell electrically, but when it comes to the end of the nerve cell and the message must be passed to another nerve cell, the message must be converted to a chemical signal. This is where neurotransmitters come in: they are the chemicals that carry the message between nerve cells. The space between two nerve cells is called the synapse. The nerve cell that is sending the message is called the presynaptic cell and the nerve cell that is receiving the message is called the postsynaptic cell. When the presynaptic cell gets the signal to pass on the message, it releases the stored neurotransmitters into the synapse. Once in the synapse, the neurotransmitters can be taken up by receptors on the postsynaptic cell, and the message begins to be passed through the new cell. In order to prevent too much signaling, the neurotransmitter cannot stay in the synapse for too long. The presynaptic cell begins to take back the neurotransmitter, storing it for the next time that a message needs to be passed across the synapse. This recycling of neurotransmitter is called “reuptake.” (For more information on the neurobiology of HD, click here.)
Histone deacetylases (HDACs) are enzymes involved in expression of DNA. Blocking these enzymes with HDAC inhibitors such as sodium butyrate and suberoylanilide hydroxamic acid (SAHA) has beneficial effects on fruit-fly and mouse models of HD. Why are HDACs significant in the study of HD and how could they lead to HD therapies? To answer these questions, we will explore histones and their role in gene expression, learn about certain histone-modifying enzymes, and then address how regulation of these enzymes can have potentially beneficial effects for an individual with HD.
Histones and DNA
If you were to stretch out all of the DNA in the human genome, it would span approximately 102 centimeters. This length is tremendous given that the average diameter of a mammalian cell’s nucleus, which is where DNA is stored, is only 5 micrometers. Given that the cell’s total length of DNA is over 20,000 times longer its storage space, fitting DNA into the nucleus requires packing it within a tightly-condensed state known as chromatin. In its chromatin form, DNA is organized in structures called nucleosomes, which consist of both DNA and proteins known as histones. Each nucleosome has 147 base pairs of DNA wrapped around a scaffolding of eight histones like twine around a spool. Thus, histones provide the structural support for packaging DNA.
Not only do histones provide a backbone for packaging DNA, they also regulate the function of DNA, which is to replicate the body’s cells and to transcribe all of the body’s protein building blocks. When the cell needs to divide or transcribe its genes, signaling proteins are sent to interact with the “tail” of amino acids on the histones, causing them to unwind their DNA. When DNA is unwound, its distinct strands can be accessed by the cell’s replication or transcription machinery.
DNA accessibility can be regulated by histones by adding and then subsequently removing molecules such as acetyl groups, methyl groups, and phosphate groups to or from specific sites on the histone tail. These histone modifications are performed by enzymes that are specialized to attach or remove a specific group. For example, enzymes known as histone methyltransferases are responsible for attaching methyl groups to histones. These attached groups are small compared to the size of the structure they are attached to, but their presence or absence on the histone have a big effect on the accessibility of the DNA in its tightly-condensed chromatin form. For instance, adding a methyl group to the ninth lysine on histone 3 represses transcription, while adding it to the seventeenth arginine activates transcription. The significant effect of this methyl group and other histone modifiers has been attributed to two different mechanisms:
The binding of modifiers disrupts contacts between nucleosomes, resulting in the unraveling of the DNA.
The bound modifiers attract or repel other proteins that initiate a variety of enzymatic activities (e.g. prepare the DNA for transcription).
Clearly, it is very important for cells to precisely control which modifications go on which part of the histone. The huge variety of different combinations of attached groups has led scientists to suggest that these modifications serve as a “code” that tells the cell what to do with the DNA wound around the histone.
Histone acetylation and HD
Because the chromatin structure is so important in regulating in the accessibility of DNA, histones and the enzymes that modify them play significant roles in controlling which genes are expressed and which are turned off. This method of regulation is very relevant in the nervous system, which depends on the coordinated expression of many different genes in order to develop and maintain itself. For example, recall that all neurons are descendants of embryonic stem cells, cellular progenitors that have the potential to turn into many other types of cells. What these stem cells ultimately become depends on the signals that they receive from their environment. (For more information on stem cells, click here.) One way that these signals may “tell” stem cells what to become is through histone modifications. This makes sense when you consider the importance of chromatin structure in determining the access of genes—turning different genes “on” and “off” results in stem cells taking on different fates. As we will soon see, in an organ as critical and complex as the brain, even small changes in gene expression can have big effects on function.
The addition of a chemical structure known as an acetyl group to histones is associated with activating transcription. This addition, which is known as acetylation, is performed by enzymes known as histone acetyltransferases (HATs) and produces a more transcriptionally-active form of chromatin. Conversely, the removal of attached acetyl groups by enzymes called histone deacetylases (HDACS) represses transcription. Varying the amounts of active HATs and HDACs allows the cell to control the accessibility of its chromatin, and thus which genes are turned on and when. One simple way to think about this relationship is a seesaw, with the left side signifying a certain gene is “on” and the right side signifying that a gene is “off.” If there are more HATs on the left side than there are HDACs on the other side, more histones will be acetylated than not and the seesaw will tilt towards the gene being transcribed. On the other hand, if HDACs outnumber HATs, acetyl groups will be removed faster than they are added and the transcription of the gene is repressed. Of course, the control of gene transcription is more complex than a seesaw of HDACs and HATs. However, this model provides a simple way of understanding what can go wrong if the balance between HDACs and HATs goes changes, as what appears to happen in HD.
Mutant huntingtin and acetylation:
An imbalance in histone modifications occurs in HD and researchers are trying to correct this problem as a therapeutic strategy. The mutated version of the huntingtin protein has been shown to disrupt transcription. (For more information about mutant huntingtin, click here.) Many genes important for the proper functioning of neurons, such as those responsible for neurotransmitter signaling, are down-regulated, or show reduced expression, in the brains of HD patients and animal models. The reduced output of these genes plays a major role in the massive neurodegeneration seen in HD and is likely at least partially due to mutant huntingtin’s effect on histones. Experiments examining the chromatin of genes known to be downregulated in both HD cell lines and HD mice and found that their histones contained fewer acetyl groups than usual. Recalling the well-established role that acetylation plays in transcriptional activation, these results provide a way of understanding how transcription is affected in HD.
Further investigations have provided more insight into what actually causes the reduced acetylation seen in HD. Mutant huntingtin has been observed to interact with both HATs, the enzymes that add acetyl groups, and HDACs, the enzymes that remove them. While there are some indications that mutant huntingtin may directly act on HATs to de-active them, there has been more evidence suggesting that the mutated protein affects the histone-modifying enyzmes by making them unavailable to do their jobs. This latter mechanism is a recurring one in HD—mutant huntingtin sequesters transcription factors and co-activators (molecules that help transcription factors bind to DNA), resulting in abnormal gene expression. But what happened to these coactivators and transcription factors? In order to answer this question, we need to review some of the disease mechanisms that take place in the cells with mutant huntingtin.
A Brief Review of HD Mechanisms
The expanded CAG repeat found in the HD gene produces a mutant form of the huntingtin protein. At some point in the life of mutant huntingtin, enzymes known as caspases cleave the protein, producing huntingtin fragments. These fragments are transported into the nucleus of the HD brain cell, or neuron, where they aggregate to form neuronal inclusions (NIs) in the nucleus. Figure 3 shows a diagram depicting the formation of NIs from the altered huntingtin protein. (For more information about the mutant huntingtin protein and protein aggregation, click here.)
Recent studies showed that the NIs “trap” various co-activators and transcription factors and prevent them from doing their jobs. This discovery led scientists to speculate that one of the ways by which HD progresses is through the loss of transcription of several key genes essential for cell survival. It was then important for scientists to try to identify which molecules are being trapped by the NIs so that this molecular “trapping” can be counteracted.
One such molecule called CREB-binding protein (CBP), which is an HDAC, appears to associate with NIs. CBP is a co-activator because it induces histones to adopt a more open chromatin configuration, allowing transcription factors to bind. If CBPs are trapped by the NIs, transcription factors can’t access DNA and certain genes are not transcribed. Sequestering CBP in HD especially affects p53, a transcription factor that regulates neuronal death, but is better known for its role as a tumor-suppressor protein. When few CBP molecules are available to co-activate p53, it is unable to access and bind to DNA. Without p53, abnormal gene transcription and expression occurs, and scientists speculate that this error in transcription may lead to cell death.
In summary, the altered huntingtin protein forms NIs that trap coactivators such as CBP. Loss of CBP results in the loss of histone acetylation, which in turn results in the inability of the transcription factor p53 to bind to DNA. Drugs such as HDAC inhibitors that could compensate for the loss of the coactivator CBP could, in theory, be possible treatments for HD. The following section summarizes some of the recent studies that test this theory using animal models.
The disruption of localized processes, such as p53 binding, by mutant huntingtin results in global changes to the cell. Gene profiling studies, which examine the expression of a tremendous number of genes, have found significant differences in the messenger RNAs (mRNAs) that are produced in individuals with HD compared to those unaffected by the disease. Recall that mRNAs are transcribed from DNA and form the genetic code that is “read” to produce proteins. The changes seen in the expression profiles of individuals affected by HD suggest that the disease impacts transcription at a global level.
The disruption of localized processes, such as p53 binding, by mutant huntingtin results in global changes to the cell. Gene profiling studies, which examine the expression of a tremendous number of genes, have found significant differences in the messenger RNAs (mRNAs) that are produced in individuals with HD compared to those unaffected by the disease. Recall that mRNAs are transcribed from DNA and form the genetic code that is “read” to produce proteins. The changes seen in the expression profiles of individuals affected by HD suggest that the disease impacts transcription at a global level.
From bench to bedside:
Because of the strong evidence linking histone dysregulation to the disrupted transcription seen in HD, scientists have hypothesized that addressing the imbalances in histone modifications may improve the symptoms of HD. Research been particularly focused on a diverse group of molecules known as HDAC inhibitors, which prevent the deacetylases from removing acetyl groups.
As mentioned above, CBP gets “trapped” in NIs which disrupts DNA transcription. Steffan, et al. (2001) looked into the possibility of reversing or preventing CBP aggregation in NIs. CBP functions as a HAT but this function is lost when it is “trapped” in the NIs. This loss of HAT function (or acetyltransferase activity) may be counteracted by inhibiting HDACs (or reducing deacetylase activity). In other words, blocking the inhibitor of transcription would make up for the low levels of transcription activators. To test this idea, Steffan, et al. used a Drosophila (fruit fly) model of HD, which was genetically modified to express mutant huntingtin. These HD mutant flies show degeneration of nerve cells and decreased survival rates similar to what is observed in people with HD.
To assess the efficacy of reducing deacetylase activity in treating HD, the flies received food containing various HDAC inhibitors, including sodium butyrate and suberoylanilide hydroxamic acid (SAHA). After treatment, the researchers discovered that the flies fed SAHA showed increased histone acetylation compared to untreated flies as well as a slowing in the progression of neurodegeneration. SAHA also increased survival rates: 70% of untreated flies showed early death compared to 45% of SAHA-treated flies. The results of this study suggest that mutant huntingtin reduces levels of acetylation and transcription by sequestering co-activators (such as CBP and others) and trapping them into aggregates.
Similar studies have been carried out in mouse models of HD, which are better than invertebrate models in assessing how the disease progresses in humans. In 2003, Hockly et al. found that HD mice that were administered SAHA through their drinking water performed better on the Rotarod, a revolving rod that is used to assess motor coordination, than HD mice that were given a placebo.
These effects of SAHA have also been investigated at the molecular level, an important validation step that confirms that the drug is doing what it is supposed to do in the cells of HD mice. Experiments performed by Mielcarek et al. in 2011 showed that SAHA reduced the levels of HDAC4, a class of histone deacetylases, in the cortex and brain stem of HD mice. Recalling the see-saw model of histone acetylation (see above), lower concentrations of HDAC would be expected to result in higher levels of histone acetlyation and thus transcriptional activation—genes being turned “on.” The potentially positive effects SAHA could be seen in the significant reduction of mutant huntingtin aggregation and the partial restoration of the HD mice’s levels of brain-derived neutrophic factor (BDNF), a protein that is important for the survival and growth of certain neurons and is inhibited in HD. (For more information on BDNF, click here.) Despite these promising results, the investigators also found that both wild-type and HD mice treated with SAHA showed significant weight loss, a side-effect that needs to be taken into account if the chemical enters clinical trials.
Thomas et al. (2008): Researchers at The Scripps Research Institute in California have developed an HDAC inhibitor that staves off disease progression in a mouse model of HD. HD mice treated with the drug, called HDACi (HDAC inhibitor) 4b, had significant improvements in movement and coordination, and lost less weight. When scientists looked at the brains of the HD mice, they found that HDACi 4b countered some of the negative effects that HD causes. HD mice usually have a smaller striatum and larger ventricles, and have smaller brains overall. (For more information on HD’s effect on the brain, click here.) However, HD mice treated with HDACi 4b had brains that were just as large as the brains of normal mice, and had a normal-sized striatum and ventricles.
Notably, HDACi 4b has very low toxicity – which is important because many HDAC inhibitors had toxic side effects in mice, and therefore can’t be used in humans. The researchers suggest that HDACi 4b should be studied in humans, as it shows potential to help people with Huntington’s disease.
The results of these studies have been replicated and adapted in subsequent studies looking at other HDAC inhibitors. Taken together, the body of evidence presents a compelling case that this class of molecules could be effective in treating HD. However, there are significant challenges in translating basic science research into clinical therapies. (For more information on clinical trials, click here.) In 2006, researchers concluded Phase II of a clinical trial examining the safety and tolerability of phenylbutyrate, a HDAC inhibitor that has been found to increase acetylation and improve motor function in mice models of HD. As of the end of 2011, it is unclear whether phenylbutyrate is still actively being considered as a treatment for HD, although scientists have been publishing based on the results of the Phase II study as recently as 2010.
The development of HDAC inhibitors into viable drugs to treat HD can draw from ongoing research into their use as therapies for cancer. Two HDAC inhibitors (including SAHA, the molecule tested in fly and mouse models of HD) are FDA-approved for the treatment of cutaneous T-cell lymphoma, a cancer of a class of immune cells. Many more are being investigated in clinical trials for a range of cancers, an encouraging indication that these drugs are regarded as generally safe for use. But despite their use in current treatments, scientists are still unclear about how increasing histone acetylation improves clinical outcomes. For example, there is now evidence that the majority of molecules targeted by HDACs for acetyl group removal are not histones, but other protein complexes important for cellular function. Clearly, scientists still have an incomplete understanding of how HDAC inhibitors work at the molecular level.
While further research is needed to determine whether HDAC inhibitors can be developed into a safe and effective drug for HD, early results suggest that these molecules hold promise in improving the lives of individuals affected by this terrible disease.
For further reading
Steffan, et al. “Histone deacetylase inhibitors arrest polyglutamine-dependent neurodegeneration in Drosophila”. 2001. Nature 413: 739-743. Treatment with HDAC inhibitors increased survival rates in a Drosophila model of HD.
Thomas EA, Coppola G, Desplats PA, Tang B, Soragni E, Burnett R, Gao F, Fitzgerald KM, Borok JF, Herman D, Geschwind DH, Gottesfeld JM. The HDAC inhibitor 4b ameliorates the disease phenotype and transcriptional abnormalities in Huntington’s disease transgenic mice. Proc Natl Acad Sci U S A. 2008 Oct 7;105(40):15564-9. Epub 2008 Sep 30. A technical article describing the effect of HDACi 4b on HD mice
Gray SG. Targeting Huntington’s disease through histone deacetylases. 2011. Clinical Epigenetics 2: 257-277. This technical article provides a broad review of histone deacetylases and their role in Huntington’s disease.
Butler R, Bates G. Histone deacetylase inhibitors as therapeutics for polyglutamine disorders. 2006. Nature Reviews Neuroscience 7:785-796. This is another technical review of how histone deacetylase inhibitors may prove useful in treating diseases such as HD.
Kouzarides T. Chromatin Modifications and Their Function. 2007. Cell 128:693-705 This review article provides a good overview of how the addition and removal of different groups affect chromatin structure and function.
Hockly E, Richon VM, Woodman B et al. 2003. Proceedings of the National Academy of Sciences. 100(4): 2041-2046. This is an article from the primary literature about testing SAHA on mice models of HD. that is targeted towards individuals with a science background. Although it may be difficult to understand, the introduction and conclusion are readable.
Mielcarek M, Benn CL, Franklin SA et al. 2011. PLoS ONE 6(11): 1-10. This is another article from the primary literature about SAHA in mice models of HD.
-E. Tan, 11-21-01, updated by M. Hedlin, 10-6-11, Y. Lu, 2-27-12
-Histone update by Y. Lu, 2-27-12
Drug Summary: Memantine is an anti-glutamate and energy-buffering drug. As an NMDA antagonist, memantine prevents the neurotransmitter glutamate from leading to nerve cell degeneration by inhibiting glutamate´s binding to the receptor. Memantine has been clinically used to treat dementia and Alzheimer´s disease. Current research on its effects in other diseases of the central nervous system (CNS), including HD, looks promising because memantine appears to be well-tolerated, and may help learning. It is possible that memantine may even be able to disrupt the progression of HD.
Mechanism of Action
According to a theory known as the excitotoxicity theory, lower energy levels in the nerve cells of people with HD cause them to be overly sensitive to glutamate. As a result, even normal levels of glutamate can overactivate the glutamate receptors on the nerve cells. When these receptors (also known as NMDA receptors) are activated, calcium ions enter the nerve cells. Excessive activation causes a buildup of these calcium ions, which then leads to the death of the nerve cell. (For more on the excitotoxicity theory, click here.)
HD researchers believe that memantine may have strong potential to slow the progression of HD by decreasing the NMDA receptor´s sensitivity to glutamate. Memantine is an NMDA antagonist. As an antagonist, memantine prevents the excessive binding of glutamate to NMDA receptors, inhibiting the pathway to excessive NMDA activation and nerve cell death. Memantine is also a non-competitive antagonist. “Non-competitive” means that memantine binds to a site on the NMDA receptor that is different from glutamate´s binding site. By binding to one portion of the NMDA receptor, memantine changes the overall shape of the receptor, making it more difficult for glutamate to bind to the other portion of the receptor.
Memantine differs from other NMDA non-competitive antagonists in that it allows the NMDA receptor to undergo physiological activity required for normal nerve cell functioning, while at the same time preventing the receptor from the over-activation that leads to nerve cell death. This is important because NMDA receptors still need to be activated to allow the entry of calcium ions, which facilitate learning and memory. But once again, too much activation of the receptor can lead to nerve cell death. Two properties of memantine allow the NMDA receptors to be activated to the optimal level, which allows learning but prevents nerve cell death.
The first property of memantine prevents nerve cell death by decreasing the NMDA receptor´s sensitivity to glutamate. When glutamate binds to the receptor, it increases the cell´s electrical charge. The electrical charge inside the cell first needs to rise to a specific value before the magnesium ion leaves the receptor so that calcium can now enter. In people with HD, the over-excitation by glutamate causes the magnesium ion to leave too easily, allowing the influx of calcium ions responsible for nerve cell death. On the other hand, memantine is not as sensitive as the magnesium ion towards an electrical charge. That is, more glutamate needs to bind to the receptor before memantine will leave the receptor, thereby allowing calcium ions to enter. This is an advantage for those with HD, because memantine can block the pathological pathway by not responding as easily to an excessive amount of glutamate.
Besides inhibiting over-activation by glutamate, the second property of memantine still enables the physiological pathway to learning and memory. Memantine has “fast blocking/unblocking kinetics.” This means that, after glutamate strongly activates the receptor, memantine is still capable of quickly unbinding the receptor, thereby allowing calcium to enter the nerve cell. The fast kinetics of memantine is what allows an appropriate amount of calcium to enter the nerve cell, a process necessary for learning and memory.
Memantine has been clinically used in the treatment of dementia and Alzheimer´s disease. In studies general to all chronic neurodegenerative diseases, therapeutic doses of memantine inhibit disruption of spatial learning and aid learning in general through prevention of the pathological pathway discussed earlier. Researchers are currently testing its efficacy in treating other CNS disorders, including HD. Discussed later in this article, a clinical study on treatment of HD with memantine has also discovered benefits in its ability to slow the progression of HD.
Clinicians have used memantine to treat over 200,000 patients for mostly dementia over the last fifteen years. Although memantine has been well-tolerated in humans, in animals it has produced side effects characteristic of other NMDA receptor antagonists. For instance, memantine can impair the ability to control muscular movements (ataxia), muscle relaxation (myorelaxation), and is sometimes known to cause amnesia. However, these side effects were only seen at high dosages (greater than or equal to 20mg/kg per day)-dosages far higher than the usual 5mg/kg per day used in humans for therapy. In humans, high doses of memantine have been known to result in psychosis in some rare instances. At therapeutic (low-level) doses, memantine does not display the negative side effects found in other NMDA receptor antagonists.
Other tested side effects are drug dependency and abuse. There is some evidence to show that memantine can lead to dependence in animals. A dependency on memantine appeared in rats and monkeys but only at high doses. However, researchers have widely agreed that memantine has little abuse potential based on the many years it has been clinically used, recent clinical studies, and zero reports of abuse in humans.
Research on Memantine
Beister, et al. (2004) conducted a two-year-long study with twenty-seven HD patients recruited from two different clinics. Each patient took up to a maximum of 30mg of memantine per day, depending on his/her individual tolerance for the drug. (Note: mg/day should not be confused with the units mg/kg per day that was used to specify therapeutic-level doses.)
Rating scales established in the HD medical literature measured the progression of HD. The Scale of Abnormal Involuntary Movements, the HD Rating Scale, and standardized video recordings evaluated chorea. For instance, in the videotapes, chorea was measured for the arms, legs, head, and trunk each on a three point scale, with 1 = slight, 2 = moderate, 3 = severe, and half-points possibly assigned. The scores for the different body parts were then averaged together.
The Clinical Global Impression (CGI) scale, the HD Activities of Daily Living (HD-ADL) scale, and the Total Functioning Capacity (TFC) of the HD Functional Capacity Scale were used to measure deteriorations. For example, HD-ADL consists of seventeen items that track the progress of HD through assessing a person with HD´s capabilities in taking care of him/herself in various areas, such as eating, dressing, taking medicine, and maintaining relationships. The person´s capability in each area is evaluated on a 3 point scale, with 0 indicating normal ability and 3 indicating necessary help from others required. The points for each area are summed to get the total HD-ADL score.
The Total Motor Score of the Unified HD Rating Scale (UHDRS) measured motor functioning. The Total Motor Score is reached by summing up points for certain movements, such as being able to carry out a sequence of hand movements or the velocity in moving a certain way. Scores for each movement are graded on a 4 point scale, with 0 being normal and 4 being unable to execute.
Psychometric tests, such as the Short Syndrome Test (SKT), the Brief Test of General Intelligence (KAI), and the Trail-making test were used to measure cognitive abilities.
The results following a two year treatment with memantine suggest that memantine has the ability to slow the progression of HD. Untreated people with HD in the Huntington Study Group (1996) experienced a 21.2% decrease in motor function over two years according to the Total Motor Score of the UHDRS. In comparison, treated patients experienced a decline of only 4.3%.
The scores on competence in daily living tasks also show memantine´s benefits. Researchers compared their results measured by HD-ADL with results measured by TFC of the HD Functional Capacity Scale because the two are similar enough to produce comparable outcomes. Untreated people with HD had a decrease in ability of daily living tasks, demonstrated by their average decline of 0.5 points over six months on the TFC scale. On the other hand, people with HD who received memantine treatment actually gained ability in daily living tasks, with an average increase of 0.28 points. These results translate to a 15.4% decrease in competency of daily living tasks over two years in untreated people with HD but a 9.3% reduction in progression of incompetence in daily living tasks in treated people with HD.
With no statistically significant changes in SKT and KAI, psychometric testing showed no deterioration of cognition in the treated participants.
Furthermore, in the second year of treatment (between 12 and 24 months), there were no significant changes in CGI and HD-ADL scores. This score stability indicates a reduction in progression of deterioration. It also interestingly suggests that memantine´s ability to prevent HD progression is expressed only after long treatment with memantine.
Overall, the researchers concluded that memantine has good potential to slow the progression of HD. However, more studies need to be conducted with control groups to serve as a comparison (control groups do not get treatment, they take a placebo) in order to verify the study´s findings.
Forest Pharmaceuticals, Inc. (2010) ran a phase II clinical trial in which 50 people with mild to moderate Huntington’s disease received either 10 mg of memantine or a placebo twice daily for 12 weeks. Then, for the next 12 weeks, all participants took memantine. When compared to patients taking placebo, patients taking memantine showed improvements on tests of memory and attention, but performed worse on tests measuring motor symptoms. Larger studies will be necessary to confirm these findings.
For further reading
Beister, et al. “The N-methyl-D-asparate antagonist memantine retards progression of Huntington´s disease.” Journal of Neural Transmission Supplement. 2004 Supplement; (68): 117-22. This fairly technical article presents the complete details of the study conducted by Beister, et al. The article concludes that memantine has good potential to slow the progression of HD, but more studies still need to be conducted to confirm results.
Parsons, et al. “Memantine is a clinically well tolerated N-methyl-D-asparate (NMDA) receptor antagonist-a review of preclinical data.” Neuropharmacology. 1999, Jun; 38(6): 735-67. Review. This is a highly technical article that summarizes the findings on memantine in its usage for a variety of diseases, including HD. The article also explains in detail memantine´s mechanism and tolerability.
www.memantine.com This website is easy to understand but centers on memantine´s use for Alzheimer´s disease. However, the website clearly explains memantine´s mechanism as well as provides many research studies. The research posted under “Studies & Literature” is helpful in understanding more about memantine´s effects and good tolerability.
Palmer GC. “Neuroprotection by NMDA receptor antagonists in a variety of neuropathologies.” Current Drug Targets. 2001 Sep; 2(3): 241-71. Review. This is a highly technical article that reviews memantine´s mechanism against glutamate toxicity. It is not very useful in understanding memantine´s effects on HD in particular.
Proc. of Fourth Annual Huntington Disease Clinical Research Symposium, San Pavilion Ballroom at the Hyatt Regency La Jolla at Aventine, San Diego. This technical report describes the results of the phase II clinical trial on memantine
– C. A. Chen, 05.02.05, Updated by M. Hedlin on 9.13.11