News - Research

Tay-Sachs Gene Therapy (TSGT) Consortium

• What is the Tay-Sachs Gene Therapy (TSGT) Consortium
• NIH Awards $3.5 Million Dollar 4-Year Grant to Tay-Sachs Research Team
• Goals Set to Accelerate TSGT Timeline
• TSGT Results and Progress Summary - September 2011

• February 2012
• September 2011
• August 2011
• December 2010
• July 2010
• May 2010
• March 2010
• January 2010
• 2009
• September 2008
• January 2008

What is the Tay-Sachs Gene Therapy (TSGT) Consortium?

The Tay-Sachs Gene Therapy (TSGT) Consortium was formed in 2007 with the goal of initiating a gene therapy clinical trial for Tay-Sachs disease and Sandhoff disease. Read more about the TSGT Consortium results and progress toward a cure. Clinical trials could begin as early as September 2012!
The TSGT Consortium includes scientists and clinicians from the following institutions:

• Auburn University, AL
• Boston College, MA
• Cambridge University - UK
• Massachusetts General Hospital/Harvard Medical School
• University of Massachusetts Medical School
• New York University Medical School

The TSGT Consortium is led by Dr. Miguel Sena Esteves, recipient of the 2011 Outstanding New Investigator Award from the American Society of Gene & Cell Therapy. Often referred to as the "unit of heredity," a gene is composed of a sequence of DNA required to produce a functional protein.
Visit the TSGT website at www.tsgtconsortium.com to read about all the scientists and clinicians in the TSGT Consortium.

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NIH Awards $3.5 Million Dollar 4-Year Grant to Tay-Sachs Research Team

The Tay-Sachs Gene Therapy (TSGT) Consortium has received official notification from the National Institutes of Health (NIH) - more percisely the National Institute of Neurological Disorders and Stroke - that it has been awarded a 4 year, $3,545,985 research grant!!!   The grant period begins September 1, 2009 with a first year distribution of $1,054,908!!!

The Tay-Sachs Gene Therapy Consortium consists of research experts from Massachusetts General Hospital, Auburn University, Boston College, NYU, University of Massachusetts and Cambridge University in England.  They are using gene therapy applications to try to halt the progressive neurological degeneration of Tay-Sachs disease. The first year of research produced unbelievable success in small animal models and vector distribution throughout the brain. The second year of research focused on large animal models. There are naturally occuring models in both cat and sheep populations.  In year 3 of 4 (depending on success with large animal models) the Consortium will be preparing a clinical trial protocol and seek approval from regulatory agencies in the US (FDA) and UK.

This is a huge step in our battle to find a cure for Tay-Sachs disease, made possible by the many generous donors that have supported us!!!   We are blessed to have such a talented group of researchers working on our cause!!!   Prior to obtaining this NIH grant, NTSAD’s Research Initiative awarded $577,000 in grant support in the last two years to the Tay-Sachs Gene Therapy Consortium.  These funds were critical in advancing the project and enabling the scientific team to apply for the NIH funds.  Thousands of people financed this effort through the Cure Tay-Sachs Foundation (CTSF), Cameron & Hayden Lord Foundation, NTSAD New York Chapter, Harry Hoffman Fund, Evan Lee Ungerleider Foundation, Mathew Forbes Romer Foundation and the Sophia Pesotchinsky Fund. This is a huge step in our battle to find treatments and cures for Tay-Sachs disease!

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Goal Set to Accelerate Tay-Sachs Gene Therapy Consortium Timeline by One Year

The Tay-Sachs Gene Therapy (TSGT) Consortium's investigators, funders and other team members met recently. The meeting resulted in an unprecedented collaboration to explore accelerating the time line to human clinical trials by one year, with a goal of starting clinical trials as early as September, 2012. The team is addressing the best approach for vector manufacturing, which poses the main technical challenge to the aggressive schedule.

In order to speed the start of human trials, several conditions need to be met at an additional cost of $1.2 million. Toxicity studies would need to be undertaken at our cost. More animal studies and vector manufacturing need to be undertaken and are not covered by the NIH grant.

National Tay-Sachs & Allied Diseases Association (NTSAD) and its family foundation affiliates have already pledged 75%, or $900,000, of the $1.2 million that needs to be raised. The team is seeking an additional $300,000 by the end of 2011.

In parallel, a new team will seek funding and drug development partners for the clinical trial phases.
Previously, families, friends and supporters generously donated $920,000 that was awarded in grants through NTSAD's Research Initiative. The majority of these funds were raised by family foundations affiliated with NTSAD: Cure Tay-Sachs Foundation, Cameron and Hayden Lord Foundation, Mathew Forbes Romer Foundation, Pesotchinsky Family Fund, and the NTSAD New York Chapter.

The grants led to the TSGT Consortium's impressive scientific results, which demonstrated proof of concept in cat animal models, and were presented last week by TSGT team members Douglas R. Martin, PhD, and Miguel Sena-Esteves, PhD, at the Annual Meeting of the American Society of Gene and Cell Therapy.

We are proud to report that Miguel, who is the program manager of the TSGT Consortium and associate professor of neurology at the University of Massachusetts Medical School, has received the 2011 Outstanding New Investigator Award from the American Society of Gene & Cell Therapy (ASGCT) for his contributions to the field of gene and cell therapy.

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Tay-Sachs Gene Therapy Consortium Progress Update
February, 2012

The following summarizes progress by the TSGT Consortium toward initiating a human clinical trial for Tay-Sachs disease (TSD).

1. Experiments in GM2 cats
GM2 cats were treated prior to or near the time of disease onset with AAV gene therapy delivered directly into the brain ("standard therapy group"). Treated GM2 cats lived an average of 16.7 months, or 3.7 times longer than untreated GM2 cats, which die at ~ 4.5 months of age. Remarkably two AAV-treated GM2 cats are still alive at 24 and 20.7 months of age, a remarkable outcome that surpasses our initial expectations. Of all treated cats, only one died of typical neurological disease progression. Some of the treated cats succumbed to other problems such as heart disease or bladder and bowel dysfunction, which will be closely monitored in the human clinical trial Overall, however, AAV gene therapy has shown profound therapeutic benefit in GM2 cats, and has largely corrected the central nervous system disease components (debilitating whole body tremors and balance difficulties found in untreated cats).

Another remarkable finding from our studies was that injection of a 10-fold lower vector dose increased the average lifespan to > 1 year of age (13.1 months), or 2.9-fold longer than untreated GM2 cats. Most cats in this group reached the study endpoint because of hind limb weakness, even though severe whole body tremors and balance difficulties were corrected. Two cats in this group lived to 15.3 and 16.9 months of age. The therapeutic benefit from the low dose of AAV gene therapy was far better than expected.

Other treatment groups have been initiated in GM2 cats, including tests of different injection routes and ages. Though it is too soon to draw firm conclusions from these experiments, the results are encouraging. For example, cats with mild clinical disease at the time of treatment are currently 13.8 months old (compared to non-treated cats that die by 4.5 months old). Also some of the longest surviving GM2 cats were treated with the same vectors as before but using a different combination of delivery routes thought to be safer and easier to implement in patients.

2. Experiments in Tay-Sachs Sheep
In year 1 of the research project, two Tay-Sachs sheep were treated with gene therapy and two sheep were reserved as untreated controls to carefully characterize disease progression in this new animal model. Though untreated sheep lived to ~ 8 months of age, both sheep treated with AAV gene therapy lived > 14 months and maintained a good quality of life until ~ 1 week prior to euthanasia, at which time they began to have difficulty standing. Both animals in Year 1 were symptomatic at the time of treatment, so these initial therapeutic results are encouraging. Analysis of the brain of these two sheep has produced important findings: 1) Enzyme activity is found throughout the much larger sheep brain after AAV gene therapy, similar to the findings in cats and mice. This suggests that the principles we are exploiting to supply functional enzyme to the entire brain can be harnessed with remarkable success in larger brains and across species; 2) The vector formulation identical to that proposed for the clinical trial generated the highest enzyme activity and distribution throughout the brain. Work is still underway to characterize the changes in GM2-ganglioside levels throughout the brain and spinal cord of these animals.

In Year 2 of the project, 10 Tay-Sachs sheep were treated with gene therapy and 2 remain as untreated controls. The purpose of Year 2 experiments is to learn whether the results from year 1 can be reproduced. Some of the sheep treated with gene therapy were assigned to short-term studies to analyze the production of HexA with different vector combinations. Other sheep were assigned to long-term studies to learn whether treated sheep in Year 2 will live as long as or longer than those from Year 1. Tissues from short-term sheep have been collected and are currently being analyzed.

3. New AAV vector design being tested
Development of a new generation of AAV vector for future clinical trials continues. Ongoing experiments include some of the animal models and delivery routes tested for the current generation of AAV vectors that will be used in the first clinical trial.

4. Pilot study for production of clinical grade AAV vectors for clinical trial
As stated in the last progress report, production of AAV vector for the clinical trial is very expensive, largely because clinical grade AAV vectors must be produced according to FDA-mandated "good manufacturing practice (GMP)" standards. After contacting most U.S.-based facilities that produce GMP-grade AAV vectors, we estimated the manufacturing cost to be $860,000-$1,000,000. However, we identified a new state-of-the-art GMP facility that had produced other clinical grade material but had not yet generated AAV vector, and therefore was willing to make our vectors at a lower cost. After our regulatory consultant visited the new facility and verified that it meets FDA requirements (and potentially the European Regulatory Agency as well), we initiated a 4-month pilot program to validate the new facility's ability to produce functional AAV vectors. The program is on track and several lots of AAV vectors have been generated to test the efficiency of the production process in this facility. Studies are underway to compare the research grade AAV vectors used in animal experiments and those generated at the GMP facility.

5. Feasibility and safety study in monkeys
We have received approval from the animal welfare and biosafety committees for upcoming safety studies in monkeys, and the animals are scheduled to arrive within the week. The monkeys will undergo a quarantine period to ensure that all animals are healthy before testing begins. Monkeys were selected for this study based on rigorous criteria that required screening close to 100 monkeys before making the final selection of animals. These studies are critical to demonstrate the safety of the vectors and injection procedures. As a result the surgical procedures, equipment (infusion cannulas) and brain imaging technologies used to guide the injection needles to the desired targets in the brain will be the same as, or as close as possible to, those that will be employed in the clinical trial patients.

6. Human studies and design of the human clinical trial
Our retrospective study in 97 infantile Tay-Sachs disease patients published in Pediatrics in 2011, the premier journal in this field, allowed us to establish a clinical severity scoring system for this patient population. More recently we also collected retrospective data on 10 juvenile patients to understand disease progression that led to the development of a new clinical severity scoring systems for these patients. Based on these studies we initiated a prospective study with 6 infantile and 5 juvenile patients to assess disease progression and the usefulness of the new disease severity scoring systems in capturing accurately disease status at any given time. A neuropsychologist performed the Vineland Scale for Adaptive Living Skills, and the Gross Motor Functional Classification System was applied to all patients as well.

In addition to developing and assessing new clinical scoring systems in living patients, we have developed a new brain MRI scoring system. Two expert neuroradiologists reviewed a total of 10 brain MRIs of infantile GM2 patient to develop the new scoring system. The findings from this study have been accepted for an oral presentation at the American Society for Neuroradiology (NYC, 2012). Recently we reviewed 5 additional brain MRIs, successfully applying the scoring system to juvenile patients as well, despite some differences in deep brain structures that appear less compromised than in infantile patients.

A February meeting of physicians, regulatory consultants, and scientists was held in Boston to discuss the design of the first human clinical trial of AAV gene therapy for Tay-Sachs disease. After a summary of results in mice, cats and sheep, discussions of potential biomarkers and clinical trial design ensued. Inclusion and exclusion criteria were covered, and measures by which safety and efficacy can be measured were discussed in great detail. The next step is to continue our discussions with the FDA initiated in Feb 2011. According to FDA guidelines, the chief goal of the first clinical trial for any disease is to demonstrate safety of the potential therapy, though efficacy may also be evaluated for rare diseases.

Numerous regulatory steps must be met before the clinical trial can begin, such as filing an "investigational new drug (IND)" application with the FDA and receiving approval from the "institutional review board (IRB)" at the Massachusetts General Hospital where we are planning to conduct the clinical trial. The next step for our program is to conduct a pre-IND meeting with the FDA scheduled for the beginning of March 2012. For this meeting we have submitted a 182-page document describing the results from therapeutic efficacy experiments in animal models of GM2-gangliosidoses, design of upcoming safety studies in mice and monkeys, manufacturing specifications and proposed clinical trial design. Discussions with the Institutional Review Board at the Massachusetts General Hospital will be initiated soon.

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TSGT Results and Progress Summary - September 2011

Exceptional, promising therapeutic results have been obtained in several animal models of Tay-Sachs and Sandhoff disease, as well as GM-1. This is a summary of the latest progress report showing efforts are on track for launching human clinical trials for September 2012. Full progress reports are available below.

CATS
Affected cats treated before symptoms presented with the gene therapy are alive and well an average of 3.2 times longer than an untreated affected cat. This is expected to increase as treated affected cats continue to live well in good condition. Another group of affected cats received a low dose of therapy. Most are alive and well an average of 2.8 times longer than an untreated affected cat. Other treatment groups have been started in affected cats to look at different injection routes and age of treatment. It is too soon to draw firm conclusions but results are encouraging.

SHEEP
Two Tay-Sachs sheep showing symptoms received gene therapy and lived in good health almost twice as long as untreated Tay-Sachs sheep. Brain tissues show above-normal levels of hexosaminidase activity have throughout the brain. These are outstanding results!

Thanks to extraordinary effort from Jacob sheep breeders, 12 Tay-Sachs sheep have been produced for year 2 of the project. Studies will repeat and validate the successful results demonstrated last year and look at short-term (6-9 months) and long-term (12-15 months) benefits of a new gene therapy vector (vehicle to carry corrected gene into the brain). These results will supplement mouse and cat studies and contribute to the design and initiation of human clinical trials.

NATURAL HISTORY STUDIES
The Infantile Tay-Sachs and Sandhoff retrospective study of 97 children is completed and accepted for publication in Pediatrics, the premier clinical journal of the field! The prospective study is ongoing and recruiting infantile and juvenile children. Contact \n Kim@ntsad.org for details.

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Tay-Sachs Gene Therapy Consortium Progress Update
September 2011

The TSGT consortium continues to make excellent progress and is on schedule to meet the accelerated timeline of initiating the clinical trial in the second half of 2012. The following is a summary of the different activities and developments in the last 6 months.

1. Experiments in GM2 cats

Cats with GM2 gangliosidosis have been treated prior to disease onset with AAV gene therapy delivered directly into brain structures ("standard therapy group"). Currently, most AAV-treated cats remain alive and reasonably healthy at ages ranging from 12.0 to 18.8 months. Though most cats have gait abnormalities in the hind limbs, all are able to walk, eat and use the litter pan with little difficulty, and no serious vision problems have been noted. Presently, the life span of the AAV-treated cats in the standard therapy group averages 3.2 times longer than that of untreated GM2 cats (~4.5 months), and this average life span extension will continue to increase since most of the cats remain alive and in good condition.

Additional GM2 cats have been injected in the same brain targets as the standard therapy group described above but with only one-tenth the vector dose ("low dose group"). Two cats in the low dose group remain alive and in good condition at 15.5 and 14.6 months of age. Of the remaining cats, three lived to 13.0, 13.0 and 13.1 months while one was euthanized at 7.5 months of age due to severe knee joint problems. At present, cats in the low dose group are living an average of 2.8 times longer than untreated GM2 cats.

Brain and spinal cord from 3 AAV-treated GM2 cats in the standard therapy group were analyzed at 16 weeks after injection of AAV vectors for enzyme distribution and GM2-ganglioside content. We found hexosaminidase widespread throughout the brain and spinal cord at or above normal levels, albeit not uniformly distributed, and dramatic reductions in GM2-ganglioside levels. A characteristic of the disease in untreated GM2 cats is the loss of white matter (myelin) in the brain, which is reflected in the reduction of specific myelin-associated lipids. In the brain of AAV-treated GM2 cats these myelin-associated lipids were found at levels closer to normal, suggesting that loss of white matter was attenuated considerably by the treatment. We are continuing to analyze the tissues for potential immune responses to the vector and/or enzyme.

Other treatment groups have been initiated in GM2 cats, including tests of different injection routes and ages. Though it is too soon to draw firm conclusions from most of these experiments, the results are encouraging. For example, cats with mild clinical disease at the time of treatment are currently ~8.5 months old (compared to non-treated cats that die by 4.5 months old). Though they have a noticeable body tremor (a typical symptom of neurological disease), they require no advanced levels of care and are able to walk, eat and use the litter pan independently.

2. Experiments in Tay-Sachs Sheep

In year 1 of the research project, two Tay-Sachs sheep were treated with gene therapy and
two sheep were reserved as untreated controls to carefully characterize disease progression in this new animal model. Though untreated sheep lived to ~ 8 months of age, both sheep treated with AAV gene therapy lived > 14 months and maintained a good quality of life. Gene therapy-treated sheep (both males) were able to walk/trot and butt heads in the pasture, maintained vision and had good appetites until ~ 1 week prior to euthanasia, at which time they began to have difficulty standing. Considering that both animals were symptomatic at the time of treatment, and that this is the first ever attempt to treat sheep with gene therapy, the results of project year 1 are outstanding. Currently, biochemical analyses of tissues taken at the time of euthanasia are underway, and above-normal levels of hexosaminidase activity have been documented throughout the brain.

Thanks to extraordinary effort from Jacob sheep breeders, 12 Tay-Sachs sheep have been produced for year 2 of the project. Gene therapy treatments that proved beneficial in year 1 will be repeated in year 2, and treated animals will be followed short-term (for biochemical tests of gene therapy effectiveness in the brain) or long-term (for clinical evidence of long-term therapeutic benefit). In addition to confirming year 1 results with repeated treatments in year 2, a new gene therapy vector will be tested and is hoped to be even more efficient than the year 1 versions. Sheep designated for long-term follow-up were treated in July 2011, and these experiments are anticipated to last until the Fall of 2012. Sheep designated for short-term follow-up are currently being treated, and these experiments will end in early 2012. It is expected that results from gene therapy treatments in sheep will supplement data gained from mouse and cat experiments, thereby contributing to the design and initiation of a human clinical trial for Tay-Sachs Disease.

3. New AAV vector design being tested

A new AAV vector design has been developed and testing is currently underway. We have made versions carrying human, cat, sheep, mouse, or monkey subunits for testing in animals. Tests conducted in cell culture have shown promising results, and as such we are now testing the new AAV vectors in TSD sheep (see above) and GM2 cats. Efficacy tests in mice will begin soon. The new design would use a single AAV vector instead of the two-vector formulation we have been testing. This new system could have significant advantages over the old system: Cut production cost of clinical grade material in half; higher efficiency at lower doses; compatible with alternative routes of delivery not possible with the present two vector system.

4. Pilot study for production of clinical grade AAV vectors for clinical trial

Production of AAV vector for the clinical trial is the largest expense in our pre-clinical development program. We have been in contact with different academic centers with the capabilities to produce clinical grade AAV vectors according to FDA-mandated "good manufacturing practice (GMP)" standards, but the cost was exceptionally high ($430,000-$500,000/AAV vector for a total of $860,000-$1,000,000). This year we became aware of a new state-of-the-art GMP facility, and initiated discussions with the director about their interest/ability to produce clinical-grade AAV vectors. The projected AAV production costs at this facility are considerably lower than any quotes we obtained from other academic centers. We visited the facility at the end of June 2011, and after our regulatory consultant indicated that it meets FDA requirements (and potentially the European Regulatory Agency as well), we developed a validation program for production of AAV vectors. This program is being supported by the Cure Tay-Sachs Foundation and is designed to test AAV vector yields, reproducibility, and scalability of the process. To facilitate the set up of the AAV production process at the GMP facility, one of its members traveled to the University of Massachusetts Medical School to learn the production and purification process. The 4-month pilot program is currently on track.

5. Feasibility and safety study in monkeys

We are currently in advanced stages of planning a study in monkeys to test the feasibility and safety of the injections planned for the clinical trial. For these studies we developed AAV vectors carrying monkey hexosaminidase alpha and beta-subunits to avoid potential confounding immune responses against an enzyme from another species. We are working with a primate research center to conduct these studies.

6. Human studies to support clinical trial design

We completed our retrospective study of 97 patients with infantile Tay Sachs and recently had our paper accepted in Pediatrics, the premier clinical journal in the field. Currently we are analyzing the surveys on late onset Tay-Sachs (LOTS) with a focus upon juvenile patients. We have found considerable overlap in onset and progression rate between juvenile and adult patients and are seeking to validate the information through medical records and compare the clinical data to mutation status. Our goal is to quantify deterioration in gait and speech to establish a solid outcome measure for a future trial.

In our prospective study we have so far enrolled 3 infantile and 2 juvenile patients. They were independently assessed by a neurologist and neuropsychologist at the Massachusetts General Hospital. We are recruiting more patients and we will be performing 6 (infantile) and 12 (juvenile) month follow-up evaluations. We have collected 25 brain MRIs on Tay Sachs patients and plan to establish an MRI scoring system that can be used as a biomarker for future studies and trials.
In parallel to our assessment of childhood GM2, we are evaluating adults with LOTS. Here the selective weakness of individual muscles (triceps, quadriceps) with little or no impairment of other muscle groups is remarkable. It may represent a target for future therapy development. A metabolic imaging study employing MR spectroscopy is nearing completion.

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Tay-Sachs Gene Therapy Consortium Progress Update
August 2011

TSGT Consortium Research Results
Exceptional, promising therapeutic results have been obtained in several animal models of Tay-Sachs and Sandhoff disease, as well as GM1 gangliosidosis.

• Treatment proof of concept was demonstrated and presented at the May 2011 American Society of Gene & Cell Therapy (ASGCT) meeting. Cats with GM2 gangliosidosis, who usually die at 3.5-4 months of age and cannot move, see, or use a litter box, were still alive and moving freely at age 13-14 months.
• Sheep with GM2 gangliosidosis generally die at the age of 7-8 months; the first 2 treated sheep lived almost twice as long.
• Similar results have been achieved with GM-1 cats.
• Based on this demonstrated proof of concept in animal models, the TSGT Consortium believes that it can achieve the other milestones required to begin human clinical trials.

TSGT Consortium Financing

More than $900,000 in grants has come from family foundations and other donors working in concert with the National Tay-Sachs & Allied Diseases Association (NTSAD), including:

• CureTay-Sachs Foundation
• Cameron and Hayden Lord Foundation
• Mathew Forbes Romer Foundation
• Pesotchinsky Family Fund
• NTSAD-NY Area Chapter

Also, a multi-year $3.5 million NIH grant was awarded in August, 2009.

Opportunity to accelerate research

The Tay-Sachs Gene Therapy (TSGT) Consortium investigators, funders, and other team members met recently to explore accelerating the time line to human clinical trials by one year, with a goal of starting clinical trials as early as September 2012, a year earlier than planned. mHowever, several conditions must first be met:

• Raise an additional $1.25 million, of which 75 percent ($900,000) has been pledged by NTSAD and its family foundation affiliates.
• Conduct sheep and primate studies; conduct toxicity studies.
• Undertake vector manufacturing, the main technical challenge to the accelerated schedule.

Other diseases that may benefit from the TSGT research

• Other lysosomal storage diseases (LSDs) and leukodystrophies affecting the central nervous system.

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Tay-Sachs Gene Therapy Consortium Progress Update
December 2010

The TSGT Consortium is on schedule in its NIH-funded program aimed at initiating a human clinical trial in late 2013. Financial support to conduct the clinical trial is presently not secured. The TSGT will continue to pursue its policy of working closely with the patient community/private foundations and will apply for NIH support to conduct the clinical trial.

In the second half of 2010 the TSGT developed a clinical trial plan and recently submitted to the Food and Drug Administration (FDA) an information package summarizing our development program for the Investigational New Drug (IND) application necessary to initiate the clinical trial. A teleconference is scheduled with officials at the FDA in the beginning of February 2011.

As a result of the Natural History study we developed a clinical severity scoring system for Infantile TSD and SD patients. This is critical for the clinical trial. We have designed additional studies to extend and validate this scoring system in Juvenile patients, and also to develop an MRI-based scoring system to describe changes over time in the brain of Infantile and Juvenile TSD patients.

In January 2011 we will submit an application to the NIH Rapid Access to Investigational Drugs (NIH-RAID) program to support manufacture of clinical grade (GMP) AAV vectors for the clinical trial.

Therapeutic Efficacy Experiments in Large Animal Models of GM2-gangliosidoses

Many of the critical therapeutic efficacy experiments in GM2 cats originally scheduled for Year 2 of the NIH-funded project are well ahead of the originally planed timeline. Currently, 9 GM2 cats have been treated with high-dose AAV gene therapy via bilateral injection of the thalamus and cerebellum (brain targets). These cats range in age from 2.3 to 11.4 months, whereas untreated GM2 cats live to only 4.5 months. The four oldest cats treated by high-dose AAV therapy are 11.4, 10.1, 8.3 and 8.3 months. Although the two oldest cats have pronounced hind limb weakness, they are still able to walk, eat and use the litter box independently. Other than gait abnormalities due to hind limb weakness, they behave as relatively normal cats, playing with toys and siblings.The 8.3 month-old cats have very mild hind limb weakness and appear to be less affected neurologically than their older counterparts at the same age. The remaining younger cats treated with high-dose therapy are also doing well, and we have not observed any evidence of vector toxicity in cats to date. In a second arm of the study, 6 GM2 cats were treated with a low dose of AAV gene therapy, in preparation for dose escalation groups in the human clinical trial. Low-dose cats currently range in age from 5.7 to 6.8 months, all having lived longer than untreated GM2 cats (4.5 months). While all low-dose cats still walk easily and independently, they have hind limb weakness coupled with subtle but
definite intention tremors, a typical sign of disease progression in untreated GM2 cats that is not apparent in the high-dose treatment group. Therefore, we are making progress toward defining the actual target dose for the human clinical trial.

In the sheep model of Tay-Sachs Disease, two of four affected sheep were treated with AAV gene therapy. Two of the affected sheep were left as untreated controls because this is a very new animal model whose disease progression is not yet wellcharacterized clinically. One of the untreated control sheep was euthanized before it reached a severe stage of disease so that tissues from a disease midpoint could be analyzed. The second untreated control sheep reached the endpoint of disease
progression at 8.0 months of age and deteriorated very rapidly over the final 2 weeks of life. Both AAV-treated sheep currently are ~9.8 months old and walk independently but with subtle yet definite fore limb abnormalities. The treated sheep eat well and interact with humans and other sheep normally. While these results are encouraging, it is essential to remember that clinical disease progression in the sheep TSD model has not yet been thoroughly characterized and that we have no real idea of its natural variability from animal to animal. In fact, the untreated control animal that reached the endpoint at 8.0 months of age was, from the outset of the study, the most severely affected animal of the group. Therefore, it will require several more months of observation and analysis before any conclusions can be drawn regarding gene therapy success in TSD sheep

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Tay-Sachs Gene Therapy Consortium Progress Update
July 2010

1. We have now injected 12 GM2 kittens with AAV vectors encoding the feline subunits of hexosaminidase. Also, due to recent substantial increases in colony productivity, we have begun treating cats for year 2 of the NIH-funded project ahead of schedule.

The oldest treated GM2 cat is now > 6 months old, and the second oldest is almost 5 months old. The other 10 kittens range between 1 and 3 months of age.

Untreated GM2 kittens survive until ~ 4.5 months of age and by the end have a severe whole body tremor and are unable to support their weight on four limbs. In contrast, the oldest treated GM2 cat is able to move around easily, including running after toys and making quick stops and changes in direction. It is also under these conditions that her major clinical symptom becomes apparent: instability/weakness in the rear legs. A new piece of sophisticated equipment to analyze gait in great detail has shown some mild abnormalities in the treated cat, especially on the left side.

evertheless, there are no apparent balance difficulties when sitting or standing, contrary to what is observed in untreated GM2 kittens at later stages of disease. It is important to note that, although clinical disease is evident in the rear legs, this cat is 1.5 months older than the expected life span for an untreated GM2 cat and is doing very well. In addition, no progression of clinical signs has been noted in this 6 month-old AAV-treated cat for the past 5 weeks.

The 5-month old treated GM2 cat has very slight rear leg weakness, no balance difficulties and no body tremor. Once again, we have noted no clear progression of clinical signs for the past 4-6 weeks.

The younger 1-3 month-old AAV-treated GM2 kittens appear normal thus far, but it is too early to tell whether we will see the same or better results than in the two older cats. Treated and untreated GM2 cats are being subjected to numerous tests including neurological exams, MRI, gait analysis and biochemical analysis of cerebrospinal fluid to determine the extent of disease progression/correction.

2. Four Tay-Sachs sheep along with mothers and siblings arrived at one of the TSGT's member institutions at 5 a.m. on April 17, 2010. At weaning, the mothers and siblings were transported back to their home in Texas. During an extremely busy spring season, we completed the following steps necessary to begin pilot studies of AAV gene therapy in affected sheep, the first clinically relevant model of Tay-Sachs Disease: (1) DNA testing of >50 sheep to identify those affected with Tay-Sachs Disease, carriers and normals; (2) Generation of AAV vectors carrying the sheep alpha and beta genes (the beta gene having to be cloned for the first time) to minimize immune responses in treated sheep; (3) Testing of the AAV vectors in cultured cells to confirm functionality; (4) Initiation of controlled studies of disease progression in untreated sheep, which will allow us to assess whether treatment has had any impact on disease course; (5) Careful definition of the brain injection coordinates necessary to treat Tay-Sachs sheep with AAV gene therapy. Challenges were encountered in this process because of the wide variability in Jacob sheep, which may have from 2 to 6 horns, making standardization of injection coordinates impossible. We learned that each sheep must undergo an MRI prior to surgery for accurate identification of injection coordinates. In addition, a board-certified veterinary anesthesiologist and an external collaborator from the University of Tennessee were brought in to ensure successful injection of these extremely valuable sheep.

To date, we have learned that disease onset in affected sheep begins at 1-2.5 months of age with occasional stumbling due to "knuckling" of the front hooves. There is mild variability of disease onset and progression. For example, the youngest TS sheep also has the most significant clinical disease, with obvious front limb gait defects and a tendency to lie down much more frequently than the other affected or normal sheep. By contrast, a second TS sheep born 1 week prior to the most severely affected sheep has very mild clinical disease (an almost imperceptible front limb gait defect - "walking down in the fetlock"). The remaining 2 TS sheep began to show clinical signs at similar ages (~8 weeks) and have been treated with AAV gene therapy (see further description below). To date, TS sheep have shown no abnormalities in general health measures (weight, temperature, heart rate), routine blood work (complete blood count, serum chemistries), MRI, ophthalmology exams or response to anesthesia. Other than the gait defects, the only obvious difference in TS sheep is their outgoing / curious nature compared to normal siblings.

In early June, two of the four Tay-Sachs sheep were treated by injection of large amounts of gene therapy vectors into the brain. One sheep was treated with vectors expressing both the Hex alpha and beta subunits while the other sheep was treated with a vector expressing the alpha subunit alone. This experiment should provide valuable information regarding the need for co-expression of both subunits in human clinical trials or whether treatment with a single subunit will be sufficient. Both sheep tolerated the injection procedure well and are at similar mild stages of disease progression currently. Initial tests to evaluate vector function and therapeutic effect are underway.

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Tay-Sachs Gene Therapy Consortium Progress Update
May 2010

It took more than a decade to solve the mystery. Dr. Brian Porter, a veterinary pathologist at Texas A&M took over the cases of the unknown lipidosis and consulted with Dr. Joseph Alroy, another veterinary pathologist at Tufts University in Boston. He recommended asking Dr. Ed Kolodny at New York University Medical Center to help. Dr. Kolodny's lab began putting the pieces of the puzzle together, first with the discovery in the sheep of a reduced level of hexosaminidase A (Hex A), the enzyme deficient in Tay-Sachs disease, and then in the sheep's brain of an increase in GM2-ganglioside. The next step was the cloning and sequencing of the sheep's Hex A gene which was difficult due to differences from the human Hex A gene. Finally, a mutation was found in the Hex A DNA isolated from tissue of an affected sheep. That this change actually was the cause of the neuro-degeneration was shown when the altered gene placed into cultured cells failed to produce active Hex A.

Fred and Joan Horak were elated that a cause had finally been found to explain why their lambs had died, and the International Tay-Sachs Gene Therapy Consortium quickly realized the potential of Jacob sheep as a large animal model for gene therapy trials. Without very much persuasion, the Horaks agreed to enlarge their flock in favor of carriers in the hope that perhaps two years hence a few affected lambs could be produced. They were handicapped by the lack of rams carrying the Tay-Sachs gene and therefore the first year of breeding, 2008, resulted in more carriers of the trait but no affected animals. The same outcome was anticipated in the 2009 season but everyone was surprised by the birth of four lambs with Tay-Sachs disease. By 3 months of age, three of these lambs were already showing neurological signs. A fourth lamb was also identified as affected with Tay-Sachs.

What next? The Gene Therapy Consortium moved into high gear. The Horaks donated the affected animals and others from the flock to the veterinary school in Auburn, Georgia, where Dr. Douglas Martin is carefully watching the sheep for additional signs of Tay-Sachs disease and preparing to treat them by gene therapy. Dr. Kolodny's lab has supplied the sequences of the sheep Hex A and Hex B genes to Dr. Miguel Esteves at the University of Massachusetts and he has converted them into a form which should multiply, diffuse throughout the sheep brain, and make the missing protein. Dr. Martin, whose gene therapy work with Sandhoff cats shows promise, will make the intra-cerebral injections in early June.

The researchers realize that more studies are needed before gene therapy for Tay-Sachs disease is ready for human trials. They are hoping that the next breeding season, October 2010 through March 2011, can produce another 6-8 affected animals. Fred and Joan are prepared to do their part. Mother Nature, teamwork, and financial support remain the pillars upon which rest the hopes of so many families with lysosomal storage disorders (LSD*.) Let us hope that the saga of the Jacob sheep will bring healing and renewed life to children with Tay-Sachs and other LSDs.

*Lysosomal storage disorders (LSD) are caused by a genetic mutation that prevents an essential enzyme from breaking down molecules into parts for reuse by the brain cell. In most of the LDSs, the continued buildup of these molecules leads to the eventual death of the cell and overall deterioration of the central nervous system.

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Tay-Sachs Gene Therapy Consortium Progress Update
March 2010

1. The performance of AAV-treated GM2 mice in behavioral tests of motor and neurological functions is comparable to that of age-matched normal mice.

2. In a related project sponsored by Auburn University, University of Massachusetts Medical School and the National Institutes of Health (NIH), a GM1 cat treated by AAV injection is now 6.5 months of age and its behavior continues to be indistinguishable from a normal littermate. Disease onset in this model typically occurs at 3.5-4.0 months of age. Neither the veterinary neurologist that has been following this cat on a regular basis, nor researchers with many years of experience with this cat model are able to identify any signs of disease. A number of AAV-treated GM1 cats are currently in the pipeline.

3. An AAV-treated GM2 kitten is now 2 months of age and continues to do well. By 3.5 months of age we will know better whether the AAV-injections are having an effect since untreated GM2 kittens by this age are capable of standing but not ambulating. By 2-2.5 months of age untreated GM2 kittens usually display whole body tremor. Another GM2 kitten has been treated in late March, 2010.

4. Breeding of the GM2 cat colony continues at an accelerated pace to generate GM2 kittens for therapeutic efficacy experiments.

5. The AAV vectors encoding feline alpha and beta-subunits have been tested in heterozygote (HZ; carriers of one normal and one mutant copy of the gene) GM2 cats and levels of enzyme expression in the brain and cerebellum are up to 60-fold higher than normal at 1 month after injection. Importantly there was no evidence of inflammation.

6. The Tay-Sachs sheep colony has been very productive and we have now identified four affected lambs born in the last two months. Two affected lambs and age-matched controls will be used to characterize in detail disease progression using a battery of tests to assess neurological function.

7. We are currently cloning the sheep alpha and beta genes and will produce AAV vectors carrying these genes for injection into affected lambs.

8. Two affected lambs will be injected with AAV vectors encoding the sheep alpha and beta genes. This experiment is very important to demonstrate the effectiveness of our approach in the only Tay-Sachs disease animal model available that displays a severe phenotype.

9. The retrospective natural history study is in its final phase. The results of this study will be presented at the annual family meeting of the NTSAD. Prospective studies in LOTS patients are currently underway and studies in infants are in preparation. Both aim to develop biomarkers and validate imaging and clinical scoring scales that will be used to assess treatment effects in the clinical trial.

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Progress Report January 2010

1. The NIH funded studies are well under way and showing very good results. Detailed analysis of the behavior of GM2 mice treated by our gene therapy approach is showing exceptionally positive results that meet or exceed our pre-defined success criteria.

2. We have cloned the genes for alpha- and beta-subunits of cat hexosaminidase (not available until now) into AAV vectors and demonstrated their ability to generate large amounts of HexA activity in cultured cells. These are critical for our efficacy studies in GM2 cats.

3. The support that the TSGT Consortium received over the last few years allowed the expansion of the GM2 cat colony, which is highly productive at the moment with 13 new kittens born in the last month alone. Eight new litters are anticipated in the next few weeks. This increased productivity is crucial to our project to generate the necessary GM2 kittens for the efficacy studies, and testing of additional delivery routes.

4. Tay-Sachs Natural History study. We have received 145 questionnaires of which our target group, infantile GM2, comprise 90 (62.1%). We continue to work with the NTSAD to increase the number of responses to our questionnaires. We have a preliminary report on milestones gained/lost and aberrant behavior (seizures, etc.) in 60 infants, but are awaiting complete data entry. In addition we are also surveying the literature to pick up all reported cases. We anticipate submitting a report of our findings for publication in the first half of 2010.

5. We have started an imaging study in late onset GM2 patients to assess structural and metabolic changes in the CNS. First results have been accepted for presentation at the International Society for Magnetic Resonance in Medicine (Stockholm, Presentation May 2010).

6. Also we are analyzing existing MR imaging data in infantile Tay-Sachs patients with the goal of developing an MRI scoring system. For this initial phase we will need at least 12 MRIs, and thus far we have collected 5 through the NTSAD. We are working with several physicians in different centers in the United States and England to collect data from additional patients. In a second stage, we will use a second set of MRIs to validate the scoring system.

Finally we are in the planning stages of a prospective study to assess disease progression in infantile patients using the new imaging and clinical scoring systems, validate MR spectroscopy findings from our studies in LOTS patients and their applicability to infantile patients, and also validate biochemical markers of disease progression. These studies will give us the tools to assess accurately the effect of gene therapy on disease progression in the clinical trial.

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Tay-Sachs Gene Therapy Consortium Progress Update
2009

The Discovery of Tay-Sachs Disease in Rare Sheep
Tay-Sachs disease has been discovered in a rare breed of sheep known as Jacob Sheep.

Their discovery is due to the persistence of Fred and Joan Horak, Texas farmers who noticed, in 1999, among their flock of Jacob sheep, two lambs that had developed signs of a nervous system disorder while still very young.  Veterinary studies convinced them that the sheep had a lipid storage disease, but no enzyme deficiency could be found.

Fred and Joan realized that they had encountered an inherited disease and because they kept good records on their flock, they knew the parental origin of the affected sheep. They could have removed these animals from their breeding stock, but instead kept the gene around so that perhaps someday it would be better understood.

It took more than a decade to solve the mystery.  Dr. Brian Porter, a veterinary pathologist at Texas A&M, referred the Horaks to Dr. Edwin Kolodny at New York University Medical Center, a member of the Tay-Sachs Gene Therapy (TSGT) Consortium. Dr. Kolodny's lab began putting the pieces of the puzzle together and discovered that the Jacob sheep had Tay-Sachs disease.

Fred and Joan Horak were elated that a cause had finally been found to explain why their lambs had died, and the Tay- TSGT Consortium quickly realized the potential of Jacob sheep as a large animal model for gene therapy trials. The Horaks agreed to enlarge their flock in favor of carriers in the hope that a few affected lambs could be produced.  In the breeding 2009 season, four lambs were born with Tay-Sachs disease.  By three months of age, three of these lambs were already showing neurological signs.

The Horaks then donated the affected animals and others from the flock to NTSAD and the veterinary school at Auburn University where Dr. Douglas Martin and Dr. Miguel Sena-Esteves quickly mobilized to start gene therapy experiments in these sheep.  The initial results have been promising, although more animals will be needed to perform further gene therapy studies.  Fred and Joan are prepared to do their part. 

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Tay-Sachs Gene Therapy Consortium Progress Update
September 2008

Since its formation in the summer of 2007, the Tay-Sachs Gene Therapy (TSGT) Consortium has been diligently working toward the goal of starting a clinical trial for Tay-Sachs and Sandhoff in the next several years. Currently the consortium is working to answer these (very simplified) questions:

• In mice, what is the most effective combination of Tay-Sachs and/or Sandhoff genes in their natural configuration or modified to increase the enzyme production?
• In mice and cats, which type of AAV sub-type is most effective? Based on results in the mice and cat models, the consortium expects to conclude which type is the most efficient AAV particle by mid-summer 2008. If it works well in mice and cats, it is reasonable to conclude that it may also be effective in humans.

The Consortium is pleased to share these (very simplified) results:

• It is essential to inject mice early for maximum benefit. Affected mice treated at 1 month are alive and in good health at 18 months; the humane endpoint of untreated affected mice is 4 months. The Consortium anticipates these mice will reach 24 months of age, which is the normal life span of mice. Mice treated at later stages showed some benefit but not as remarkable as the mice treated early.
• Two treated affected cats lived to 7 and 8 months of age; normal lifespan for untreated affected cats is 4.5 months.
• The detailed comparison of the lipids (fats) in the brains of affected mice, cats and humans is nearly complete. The Consortium is also measuring the GM2-ganglioside levels in treated affected cats, and characterizing the changes in myelin (white matter) in mice and cats.

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Tay-Sachs Gene Therapy Consortium Progress Update
January 2008

Since forming last summer, the Tay-Sachs Gene Therapy (TSGT) Consortium has been busy working toward the goal of starting a clinical trial for Tay-Sachs and Sandhoff in the next three years. In fact, participating laboratories hired new personnel to accelerate the rate of progress. Currently the consortium is working to answer these (very simplified) questions:

• In mice, what is the relationship between the amount of vector (a vector transports the correct genetic code into the cells) and the therapeutic effect?

• In mice, which is more effective: a vector that carries the corrected Tay-Sachs and Sandhoff gene OR a different vectors for each the Tay-Sachs gene and the Sandhoff gene?

• Demonstrating effectiveness in larger animal models is an important step towards a clinical trial; do we get the same results in cats as in mice?

In the coming months, in collaboration with the Consortium, NTSAD will be sending a Natural History Survey to families and individuals affected by Tay-Sachs and Sandhoff. The survey will provide a detailed description of the diseases from the earliest symptoms to terminal stages. This is a vital component of the clinical trial because it provides a quantified baseline to compare the effectiveness of the gene therapy in patients.

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