The following discussion and analysis of our financial condition and results ofoperations should be read in conjunction with our unaudited condensedconsolidated financial statements and related notes included in this QuarterlyReport on Form 10-Q and the audited financial statements and notes thereto as ofand for the year ended December 31, 2021 and the related Management's Discussionand Analysis of Financial Condition and Results of Operations, included in ourAnnual Report on Form 10-K for the year ended December 31, 2021, or AnnualReport, filed with the Securities and Exchange Commission, or the SEC, on March31, 2022. Unless the context requires otherwise, references in this QuarterlyReport on Form 10-Q to "we," "us," and "our" refer to Taysha Gene Therapies,Inc. together with its consolidated subsidiaries.
Forward-Looking Statements
The information in this discussion contains forward-looking statements andinformation within the meaning of Section 27A of the Securities Act of 1933, asamended, or the Securities Act, and Section 21E of the Securities Exchange Actof 1934, as amended, or the Exchange Act, which are subject to the "safe harbor"created by those sections. These forward-looking statements include, but are notlimited to, statements concerning our strategy, future operations, futurefinancial position, future revenues, projected costs, prospects and plans andobjectives of management. The words "anticipates," "believes," "estimates,""expects," "intends," "may," "plans," "projects," "will," "would" and similarexpressions are intended to identify forward-looking statements, although notall forward-looking statements contain these identifying words. We may notactually achieve the plans, intentions, or expectations disclosed in ourforward-looking statements and you should not place undue reliance on ourforward-looking statements. Actual results or events could differ materiallyfrom the plans, intentions and expectations disclosed in the forward-lookingstatements that we make. These forward-looking statements involve risks anduncertainties that could cause our actual results to differ materially fromthose in the forward-looking statements, including, without limitation, therisks set forth in Part II, Item 1A, "Risk Factors" in this Quarterly Report onForm 10-Q and Part II, Item 1A, "Risk Factors" in our Annual Report. Theforward-looking statements are applicable only as of the date on which they aremade, and we do not assume any obligation to update any forward-lookingstatements.
Note Regarding Trademarks
All brand names or trademarks appearing in this report are the property of theirrespective holders. Unless the context requires otherwise, references in thisreport to the "Company," "we," "us," and "our" refer to Taysha Gene Therapies,Inc.
Overview
We are a patient-centric gene therapy company focused on developing andcommercializing AAV-based gene therapies for the treatment of monogenic diseasesof the central nervous system, or CNS, in both rare and large patientpopulations. We were founded in partnership with The University of TexasSouthwestern Medical Center, or UT Southwestern, to develop and commercializetransformative gene therapy treatments. Together with UT Southwestern, we areadvancing a deep and sustainable product portfolio of gene therapy productcandidates, with exclusive options to acquire several additional developmentprograms at no cost. By combining our management team's proven experience ingene therapy drug development and commercialization with UT Southwestern'sworld-class gene therapy research capabilities, we believe we have created apowerful engine to develop transformative therapies to dramatically improvepatients' lives. In March 2022, we announced strategic pipeline prioritizationinitiatives focused on GAN and Rett syndrome. We will conduct smallproof-of-concept studies in CLN1 disease and SLC13A5 deficiency. Development ofthe CLN7 program will continue in collaboration with existing partners withfuture clinical development to focus on the first-generation construct.Substantially all other research and development activities have been paused toincrease operational efficiency.
In April 2021, we acquired exclusive worldwide rights to TSHA-120, aclinical-stage, intrathecally dosed AAV9 gene therapy program for the treatmentof giant axonal neuropathy, or GAN. A Phase 1/2 clinical trial of TSHA-120 isbeing conducted by the National Institutes of Health, or NIH, under an acceptedinvestigational new drug application, or IND. We reported clinical safety andfunctional MFM32 data from this trial for the highest dose cohort of 3.5E14total vg in January 2022, where we saw continued clinically meaningful slowingof disease progression similar to that achieved with the lower dose cohorts,which we considered confirmatory of disease modification. We recently completeda commercially representative GMP batch of TSHA-120 which demonstrated that thepivotal lots from the commercial grade material were generally analyticallycomparable to the original clinical trial material. Release testing for thisbatch is currently underway and expected to be completed in September 2022.Additional discussions with Health Authorities are planned to discuss thesecomparability data and a potential registration pathway with feedbackanticipated by the end of 2022. For Rett syndrome, we submitted a Clinical TrialApplication, or CTA, filing to Health Canada in November 2021 and announcedinitiation of clinical development of TSHA-102 under the approved CTA in March2022. We expect to report preliminary clinical data for TSHA-102 in Rettsyndrome by year-end 2022. We recently executed an exclusive option from UTSouthwestern to license worldwide rights to a clinical-stage CLN7 program. TheCLN7 program is currently in a Phase 1 clinical
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proof-of-concept trial run by UT Southwestern, and we reported preliminaryclinical safety data for the first patient in history to be intrathecally dosedat 1.0x1015 total vg with the first-generation construct in December 2021.Development of the CLN7 program will continue in collaboration with existingpartners with future clinical development to focus on the first-generationconstruct. We will conduct small proof-of-concept studies in CLN1 disease andSLC13A5 deficiency that we believe can further validate our platform.
We have a limited operating history. Since our inception, our operations havefocused on organizing and staffing our company, business planning, raisingcapital and entering into collaboration agreements for conducting preclinicalresearch and development activities for our product candidates. All of our leadproduct candidates are still in the clinical or preclinical developmentstage. We do not have any product candidates approved for sale and have notgenerated any revenue from product sales. We have funded our operationsprimarily through the sale of equity, raising an aggregate of $319.0 million ofgross proceeds from our initial public offering and private placements of ourconvertible preferred stock as well as sales of common stock pursuant to ourSales Agreement (as defined below). In addition, we drew down $30.0 million and$10.0 million in term loans on August 12, 2021 and December 29, 2021,respectively.
On August 12, 2021, or the Closing Date, we entered into a Loan and SecurityAgreement, or the Term Loan Agreement, with the lenders party thereto from timeto time, or the Lenders and Silicon Valley Bank, as administrative agent andcollateral agent for the Lenders, or the Agent. The Term Loan Agreement providesfor (i) on the Closing Date, $40.0 million aggregate principal amount of termloans available through December 31, 2021, (ii) from January 1, 2022 untilSeptember 30, 2022, an additional $20.0 million term loan facility available atthe Company's option upon having three distinct and active clinical stageprograms, determined at the discretion of the Agent, at the time of draw, (iii)from October 1, 2022 until March 31, 2023, an additional $20.0 million term loanfacility available at our option upon having three distinct and active clinicalstage programs, determined at the discretion of the Agent, at the time of drawand (iv) from April 1, 2023 until December 31, 2023, an additional $20.0 millionterm loan facility available upon approval by the Agent and the Lenders, or,collectively, the Term Loans. We drew $30.0 million in term loans on the ClosingDate and drew an additional $10.0 million term loan on December 29, 2021. Theloan repayment schedule provides for interest only payments until August 31,2024, followed by consecutive monthly payments of principal and interest. Allunpaid principal and accrued and unpaid interest with respect to each term loanis due and payable in full on August 1, 2026.
Since our inception, we have incurred significant operating losses. Our netlosses were $84.0 million for the six months ended June 30, 2022 and $73.0million for the six months ended June 30, 2021. As of June 30, 2022, we had anaccumulated deficit of $319.6 million. We expect to continue to incursignificant expenses and operating losses for the foreseeable future. Weanticipate that our expenses will increase significantly in connection with ourongoing activities, as we:
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Our Pipeline
We are advancing a deep and sustainable product portfolio of gene therapyproduct candidates for monogenic diseases of the CNS in both rare and largepatient populations, with exclusive options to acquire several additionaldevelopment programs at no cost. Our portfolio of gene therapy candidatestargets broad neurological indications across three distinct therapeuticcategories: neurodegenerative diseases, neurodevelopmental disorders and geneticepilepsies. Our current pipeline, including the stage of development of each ofour product candidates, is represented in the table below:
TSHA-120 for Giant Axonal Neuropathy (GAN)
In March 2021, we acquired the exclusive worldwide rights to a clinical-stage,intrathecally dosed AAV9 gene therapy program, now known as TSHA-120, for thetreatment of giant axonal neuropathy, or GAN, pursuant to a license agreementwith Hannah's Hope Fund for Giant Axonal Neuropathy, Inc., or HHF. Under theterms of the agreement, HHF received an upfront payment of $5.5 million and willbe eligible to receive clinical, regulatory and commercial milestones totalingup to $19.3 million, as well as a low, single-digit royalty on net sales uponcommercialization of TSHA-120.
GAN is a rare autosomal recessive disease of the central and peripheral nervoussystems caused by loss-of-function gigaxonin gene mutations. There are anestimated 5,000 affected GAN patients in addressable markets.
Symptoms and features of children with GAN usually develop around the age offive years and include an abnormal, wide based, unsteady gait, weakness and somesensory loss. There is often associated dull, tightly curled, coarse hair, giantaxons seen on a nerve biopsy, and spinal cord atrophy and white matterabnormality seen on MRI. Symptoms progress and as the children grow older theydevelop progressive scoliosis and contractures, their weakness progresses to thepoint where they will need a wheelchair for mobility, respiratory musclestrength diminishes to the point where the child will need a ventilator (usuallyin the early to mid-teens) and the children often die during their late teens orearly twenties, typically due to respiratory failure. There is an early- andlate-onset phenotype associated with the disease, with shared physiology. Thelate-onset phenotype is often categorized as Charcot-Marie-Tooth Type 2, orCMT2, with a lack of tightly curled hair and CNS symptoms with relatively slowprogression of disease. This phenotype represents up to 6% of all CMT2diagnosis. In the late-onset population, patients have poor quality of life butthe disease is not life-limiting. In early-onset disease, symptomatic treatmentsattempt to maximize physical development and minimize the rate of deterioration.Currently, there are no approved disease-modifying treatments available.
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TSHA-120 is an AAV9 self-complementary viral vector encoding the full lengthhuman gigaxonin protein. The construct was invented by Dr. Steven Gray and isthe first AAV9 gene therapy candidate to deliver a functional copy of the GANgene under the control of a JeT promoter that drives ubiquitous expression.
We have received orphan drug designation and rare pediatric disease designationfrom the U.S. Food and Drug Administration, or the FDA, for TSHA-120 for thetreatment of GAN. In April 2022, we received orphan drug designation from theEuropean Commission for TSHA-120 for the treatment of GAN.
There is an ongoing longitudinal prospective natural history study being led bythe NIH, that has already identified and followed a number of patients with GANfor over five years with disease progression characterized by a number ofclinical assessments. The GAN natural history study was initiated in 2013 andincluded 45 patients with GAN, aged 3 to 21 years. Imaging data from this studyhave demonstrated that there are distinctive increased T2 signal abnormalitieswithin the cerebellar white matter surrounding the dentate nucleus of thecerebellum, which represent one of the earliest brain imaging findings inindividuals with GAN. These findings precede the more widespread periventricularand deep white matter signal abnormalities associated with advanced disease. Inaddition, cortical and spinal cord atrophy appeared to correspond to moreadvanced disease severity and older age. Impaired pulmonary function in patientswith GAN also was observed, with forced vital capacity correlating well withseveral functional outcomes such as the MFM32, a validated 32-item scale formotor function measurement developed for neuromuscular diseases. Nocturnalhypoventilation and sleep apnea progressed over time, with sleep apnea worseningas ambulatory function
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deteriorated. Total MFM32 score also correlated with ambulatory status, whereindependently ambulant individuals performed better and had higher MFM32 scoresthan the non-ambulant group, as shown in the graph below.
Patients also reported significant autonomic dysfunction based on the COMPASS 31self-assessment questionnaire. In addition, nerve conduction functiondemonstrated progressive sensorimotor polyneuropathy with age. As would beexpected for a neurodegenerative disease, younger patients have higher baselineMFM32 scores. However, the rate of decline in the MFM32 scores demonstratedconsistency across patients of all ages, with most demonstrating an average8-point decline per year regardless of age and/or baseline MFM32 score, as shownin the natural history plot below.
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A 4-point score change in the MFM32 is considered clinically meaningful,suggesting that patients with GAN lose significant function annually. To date,we have up to eight years of robust data from this study.
Preclinical Data
TSHA-120 performed well across in vitro and in vivo studies, and demonstratedimproved motor function and nerve pathology, and long-term safety across severalanimal models. Of note, improved dorsal root ganglia, or DRG, pathology wasdemonstrated in TSHA-120-treated GAN knockout mice. These preclinical resultshave been published in a number of peer-reviewed journals.
Additional preclinical data from a GAN knockout rodent model that had receivedAAV9-mediated GAN gene therapy demonstrated that GAN rodents treated at 16months performed significantly better than 18-month old untreated GAN rodentsand equivalently to controls. These rodents were evaluated using a rotarodperformance test which is designed to evaluate endurance, balance, grip strengthand motor coordination in rodents. The time to fall off the rotarod, known aslatency, was also evaluated and the data below demonstrated the clear differencein latency in treated versus untreated GAN rodents.
A result is considered statistically significant when the probability of theresult occurring by random chance, rather than from the efficacy of thetreatment, is sufficiently low. The conventional method for determining thestatistical significance of a result is known as the "p-value," which representsthe probability that random chance caused the result (e.g., a p-value = 0.01means that there is a 1% probability that the difference between the controlgroup and the treatment group is purely due to random chance). Generally, ap-value less than 0.05 is considered statistically significant.
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With respect to DRG inflammation, a topic of considerable interest within thegene therapy arena, the DRG have a significantly abnormal histologicalappearance and function as a consequence of underlying disease pathophysiology.Treatment with TSHA-120 resulted in considerable improvements in thepathological appearance of the DRG in the GAN knockout mice. Shown below istissue from a GAN knockout mouse model with numerous abnormal neuronalinclusions containing aggregates of damaged neurofilament in the DRG asindicated by the yellow arrows. On image C, tissue from the GAN knockout micetreated with an intrathecal (IT) injection of TSHA-120 had a notable improvementin the reduction of these neuronal inclusions in the DRG.
When a quantitative approach to reduce inclusions in the DRG was applied, it wasobserved that TSHA-120 treated mice experienced a statistically significantreduction in the average number of neuronal inclusions versus the GAN knockoutmice that received vehicle as illustrated below.
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Additionally, TSHA-120 demonstrated improved pathology of the sciatic nerve inthe GAN knockout mice as shown below.
Results of Ongoing Phase 1/2 Clinical Trial
A Phase 1/2 clinical trial of TSHA-120 is being conducted by the NIH under anaccepted IND. The ongoing trial is a single-site, open-label, non-randomizeddose-escalation trial, in which patients are intrathecally dosed with one of 4dose levels of TSHA-120 - 3.5E13 total vg, 1.2E14 total vg, 1.8E14 total vg or3.5E14 total vg. The primary endpoint is to assess safety, with secondaryendpoints measuring efficacy using pathologic, physiologic, functional, andclinical markers. To date, 14 patients have been intrathecally dosed and twelvepatients have at least three years' worth of long-term follow up data.
At 1-year post-gene transfer, a clinically meaningful and statisticallysignificant slowing or halting of disease progression was seen with TSHA-120 atthe highest dose of 3.5E14 total vg (n=3). The change in the rate of decline inthe MFM32 score improved by 5 points in the 3.5E14 total vg cohort compared toan 8-point decline in natural history.
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Although the change in the MFM32 score was clinically meaningful, we might haveexpected a greater change in the MFM32 score compared to natural history in thefirst year but one patient in the high dose cohort was a delayed responder. Atthe 12-month follow-up visit, the patient had a 7-point decline in the MFM32total score that was similar to the slope of the natural history curve as shownbelow. Notably, from Year 1 post gene transfer to Year 2, this patient's changein the MFM32 score remained unchanged suggesting stabilization of disease at 2years post-treatment. At that 2-year post treatment timepoint, there was a9-point improvement in the patient's MFM32 score compared to the estimatednatural history decline of 16 points. The annualized estimate of natural historyover time assumes the same rate of decline as in Year 1.
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An additional analysis was performed to examine the change in the rate ofdecline in the MFM32 score of all therapeutic doses combined (n=12). As shownbelow, the change in the rate of decline in the MFM32 score improved by 7 pointsby Year 1 compared to the natural history decline in the MFM32 score of 8points. This result was clinically meaningful and statistically significant.
A Bayesian analysis was conducted on the 1.2E14 total vg, 1.8E14 total vg and3.5E14 total vg dose cohorts at Year 1 to assess the probability of clinicallymeaningful slowing of disease progression as compared to natural history. Thistype of statistical analysis enables direct probability statements to be madeand is both useful and accepted by regulatory agencies in interventional studiesof rare diseases and small patient populations. As shown in the table below, forall therapeutic dose cohorts, there was nearly 100% probability of any slowingof disease and a 96.7% probability of clinically meaningful slowing of 50% ormore following treatment with TSHA-120 compared to natural history data.
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There remained consistent improvement in TSHA-120's effect over time on the meanchange from baseline in the MFM32 score for all patients in the therapeutic dosecohorts compared to the estimated natural history decline over the years. ByYear 3, as depicted below, there was a 10-point improvement in the mean changefrom baseline in MFM32 score for all patients in the therapeutic dose cohorts.
In addition to the compelling three-year data, there was one patient at Year 5whose MFM32 change from baseline improved by nearly 26-points in the 1.2E14total vg dose cohort compared to the estimated natural history decline of 40points by this timepoint.
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Below is an additional analysis of the mean change from baseline in MFM32 scorefor the therapeutic dose cohorts compared to natural history at patients' lastvisit. As shown, TSHA-120 demonstrated increasing improvement in the mean changein MFM32 score from baseline over time.
Sensory nerve action potential, or SNAP, was assessed through nerve conductionstudies in patients with GAN. Natural history data from the NIH suggest rapidand irreversible decline in sensory function early in life in patients with GAN.SNAPs are within normal limits early in life and rapid reduction in SNAPamplitude occurs around the age of symptom presentation. As demonstrated below,all patients with classic GAN have an abnormally low SNAP by the age of 4,reflective of compromised sensory neuronal function. By age 9, all patients hadan irreversibly absent SNAP. The results from these nerve conduction studiesreflect the clinical progression of patients with GAN.
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TSHA-120-treated patients demonstrated a durable improvement in SNAP responsecompared to natural history. Five of the twelve patients treated demonstrated aresponse. One patient demonstrated near complete recoverability to normal fromzero at the time of treatment.
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Once SNAP reaches zero, natural history suggests sensory function is presumednon-recoverable. Among patients treated with 1.2E14 total vg or greater ofTSHA-120, the three patients with a positive value at baseline maintained apositive SNAP at last study visit with the longest span of 3 years to date andcontinue to improve.
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Below are individual patient SNAP change from baseline from treated patients whoshowed a positive response including their run-in natural history.
Biopsies of TSHA-120-treated patients confirmed presence of regenerative nerveclusters. Below is pathology data from biopsies of the superficial radialsensory nerve in 11 out of 11 patient samples analyzed. The remaining twosamples were unable to be assessed due to biopsy limitations. Peripheral nervebiopsies from the superficial radial sensory nerve were obtained at baseline andat 1 year post gene therapy transfer. Data consistently generated an increase inthe number of regenerative clusters observed at Year 1 compared to baseline,indicating active regeneration of nerve fibers following treatment withTSHA-120. Data also indicated improvement in disease pathology, providingevidence that the peripheral nervous system can respond to treatment.
Loss of vision has been frequently cited by patients and caregivers as a symptomthey find particularly debilitating and would like to see improvement infollowing treatment. Patients were analyzed for visual acuity using a standardLogarithm of the Minimum Angle of Resolution, or LogMAR. An increase in LogMARscore represents a decrease in visual acuity. A LogMAR score of 0 means normalvision, approximately 0.3 reflects the need for eyeglasses, and a score value of1.0 reflects blindness. Based on natural history,
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individuals with GAN experience a progressive loss in visual function asindicated by an increase in the LogMAR score. Ophthalmologic assessmentsfollowing treatment with TSHA-120 demonstrated preservation of visual acuityover time compared to the loss of visual acuity observed in natural history.Stabilization of visual acuity was observed following treatment with TSHA-120 asdemonstrated below.
The thickness of the retinal nerve fiber layer or RNFL was also examined as anobjective biomarker of visual system involvement and overall nervous systemdegeneration in GAN. Treatment with TSHA-120 resulted in stabilization of RNFLthickness and prevention of axonal nerve degeneration compared to diffusethinning of RNFL observed in natural history as measured by optical coherencetomography, or OCT. Analysis by individual dose groups, as seen on the graphbelow, demonstrated relatively stable RNFL thickness which is in contrast to thenatural history of GAN, where RNFL decreases. Overall, these data provide newevidence of TSHA-120's ability to generate nerve fibers and preserve visualacuity.
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As of January 2022, there were 53 patient-years of clinical data to supportTSHA-120's favorable safety and tolerability profile. TSHA-120 has beenwell-tolerated at multiple doses with no signs of significant acute or subacuteinflammation, no sudden sensory changes and no drug-related or persistingtransaminitis. Adverse events related to immunosuppression or study procedureswere similar to what has been seen with other gene therapies and transient innature. There was no increase in incidence of adverse events with increaseddose. Importantly, TSHA-120 was safely dosed in the presence of neutralizingantibodies as a result of the combination of route of administration, dosing andimmunosuppression regimen.
We currently have up to six years of longitudinal data in individual patientswith GAN and collectively 53-patient years of clinical safety and efficacy datafrom our ongoing clinical study. Treatment with TSHA-120 was well-tolerated withno significant safety issues. There was no increase in incidence of adverseevents with increased dose, no dose-limiting toxicity, no signs of acute orsubacute inflammation, no sudden sensory changes and no drug-related orpersistent elevation of transaminases. Adverse events related toimmunosuppression or study procedures were similar to what was seen with othergene therapies and transient in nature.
We believe the comprehensive set of evidence generated across diseasemanifestations, depicted in the table below, support a robust clinical packagefor TSHA-120 in GAN.
In order to deliver a robust chemistry, manufacturing, and controls, or CMC,data package to support licensure discussions, we have successfully completedsix development and GMP lots of TSHA-120 with our contract development andmanufacturing organization, or CDMO, partner. We have also completed acomprehensive side-by-side biochemical and biophysical analysis of
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current and previous clinical lots. Our CDMO utilizes the same Pro10TMmanufacturing platform used to produce the original GAN lots, therefore reducingwhich is intended to reduce comparability risk. Five development lots rangingfrom 2L to 250L scale and one full-scale 500L GMP lot were analyzed side-by-sidewith the current TSHA-120 clinical lot using a comprehensive analytical panelthat meets current regulatory requirements including assays for criticalattributes such as product and process residuals, empty/full ratio, geneticintegrity, potency and strength.
The side-by-side analysis demonstrated that the newly produced TSHA-120 lotswere generally comparable to the original clinical trial material in impurityprofile including host cell contaminants, residual plasmid, empty particlecontent, aggregate content and genomic integrity. These results supported ourbiophysical and biochemical comparability of the newly produced lots.Furthermore, we developed product-specific GAN potency methods which have alsodemonstrated that the previous and current clinical lots were functionallyindistinguishable. Validation of our potency release assay is now underway.
We have applied our panel of release assays for side-by-side testing of theoriginal clinical trial material and our commercial grade lots. Shown below areeight of the most critical attributes of TSHA-120.
First, all results demonstrated that both the clinical and commercial grade lotswere of a high purity and lacked significant levels of host cell or processcontaminants such as protein and, DNA or and aggregated species. Vector puritywas in excess of 95% for all three lots and host cell protein contamination wasbelow detection. In addition, and aggregation of all lots was very low. Hostcell and plasmid DNA contamination are also important attributes to discuss withregulatory agencies since carryover represents a theoretical immunogenicity oroncogenicity risk. Residual plasmid and host cell DNA were similar for all lots,indicating a similar safety profile for both products. Empty capsids are a keyattribute for AAV vectors since empty capsids can stimulate immune responses tothe vector and reduce potency. All three lots were highly enriched in fullparticles. Potency of AAV vectors is a key measure that correlates with clinicalefficacy. We developed a number of product-specific potency assays to measurethe functional activity of our product which is reported relative to a referencestandard. These assays recapitulated the biological activity of TSHA-120starting with transduction of GAN knockout cell lines. Activity is measured byquantitation of transgene RNA or protein expression as two independent andcomplimentary readouts. We observed good agreement with both readouts and highactivity of all three lots against our reference suggesting that the lots are ofhigh and comparable activity.
Overall, these results support that our early clinical and pivotal lots arebiochemically and biophysically similar and based on these results we believethey should perform identically in a clinical study.
Recently, regulators have encouraged sponsors to conduct deeper analysis ofproduct contaminants not covered by standard release assays to better assessproduct safety and comparability. To comply with this guidance, we have addedPac-Bio next generation sequencing to our product characterization panel tobetter understand the nature of nucleic acid contaminants in our products. Thismethod not only allows us to identify the source of the nucleic acid, but alsothe fragment size, and sequence variability, which also needs to be consideredwhen assessing AAV safety and efficacy. Our analysis of the clinical trial lotand commercial grade pivotal batches demonstrated that the source andcomposition of transgene and contaminating host and plasmid
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DNA are nearly identical and provided further support that for a conclusion thatthe nature of our product is unchanged between our early clinical and pivotalbatches as noted in the below pie charts.
The TSHA-120 pivotal lot, which yielded over 50 patient doses of TSHA-120 at thehighest dose cohort of 3.5E14 total vg, is expected to complete quality releasetesting by end of the third quarter of 2022. This material positions us forfuture BLA-enabling activities and commercial production. These lots were alsoplaced on stability to provide critical shelf-life data in support of our BLAfiling.
In September 2021, we submitted a request for a Scientific Advice meeting forTSHA-120 to the United Kingdom's Medicines and Healthcare products RegulatoryAgency, or MHRA, and were granted a meeting in January 2022. MHRA agreed on ourcommercial manufacturing and release assay testing strategy including potencyassays and we plan to dose a few additional patients with commercial gradematerial, which will be released in September 2022. Finally, MHRA was supportiveof our proposal to perform validation work on MFM32 for GAN as a key clinicalendpoint and for us to explore the MFM32 items with patients and families aspart of this process. Given the positive comparability data for TSHA-120 that werecently received, additional discussions with Health Authorities to discussthese data and potential registration pathway are planned with regulatoryfeedback anticipated by year-end 2022.
TSHA-102 for Rett Syndrome
TSHA-102, a neurodevelopmental disorder product candidate, is being developedfor the treatment of Rett syndrome, one of the most common genetic causes ofsevere intellectual disability, characterized by rapid developmental regressionand in many cases caused by heterozygous loss of function mutations in MECP2, agene essential for neuronal and synaptic function in the brain. The estimatedprevalence of Rett syndrome is 350,000 patients worldwide and the disease occursin 1 of every 10,000 female births worldwide. We designed TSHA-102 to preventgene overexpression-related toxicity by inserting microRNA, or miRNA, targetbinding sites into the 3' untranslated region of viral genomes. Thisoverexpression of MECP2 is seen in the clinic in patients with a condition knownas MECP2 duplication syndrome, where elevated levels of MECP2 result in aclinical phenotype similar to Rett syndrome both in terms of symptoms andseverity. TSHA-102 is constructed from a neuronal specific promoter, MeP426,coupled with the miniMECP2 transgene, a truncated version of MECP2, andmiRNA-Responsive Auto-Regulatory Element, or miRARE, our novel miRNA targetpanel, packaged in self-complementary AAV9. Currently, there are no approvedtherapies for the treatment of Rett syndrome, which affects more than 350,000patients worldwide, according to the Rett Syndrome Research Trust.
In May 2021, preclinical data from the ongoing natural history study forTSHA-102 were published online in Brain, a highly esteemed neurological sciencepeer-reviewed journal. The preclinical study was conducted by the UTSouthwestern Medical Center laboratory of Sarah Sinnett, Ph.D., and evaluatedthe safety and efficacy of regulated miniMECP2 gene transfer, TSHA-102(AAV9/miniMECP2-miRARE), via IT administration in adolescent mice between fourand five weeks of age. TSHA-102 was compared to unregulated full length MECP2(AAV9/MECP2) and unregulated miniMECP2 (AAV9/miniMECP2).
TSHA-102 extended knockout survival by 56% via IT delivery. In contrast, theunregulated miniMECP2 gene transfer failed to significantly extend knockoutsurvival at either dose tested. Additionally, the unregulated full-length MECP2construct did not demonstrate a significant extension in survival and wasassociated with an unacceptable toxicity profile in wild type mice.
In addition to survival, behavioral side effects were explored. Mice weresubjected to phenotypic scoring and a battery of tests including gait, hindlimbclasping, tremor and others to comprise an aggregate behavioral score. miRAREattenuated
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miniMECP2-mediated aggravation in wild type aggregate phenotype severity scores.Mice were scored on an aggregate severity scale using an established protocol.AAV9/MECP2- and AAV9/miniMECP2-treated wild type mice had a significantly highermean (worse) aggregate behavioral severity score versus that observed forsaline-treated mice (p <0.05; at 6-30 and 7-27 weeks of age, respectively).TSHA-102-treated wild type mice had a significantly lower (better) meanaggregate severity score versus those of AAV9/MECP2- and AAV9/miniMECP2-treatedmice at most timepoints from 11-19 and 9-20 weeks of age, respectively. Nosignificant difference was observed between saline- and TSHA-102-treated wildtype mice.
miRARE-mediated genotype-dependent gene regulation was demonstrated by analyzingtissue sections from wild type and knockout mice treated with AAV9 vectors givenintrathecally. When knockout mice were injected with a vector expressing themini-MECP2 transgene with and without the miRARE element, miRARE reduced overallminiMECP2 transgene expression compared to unregulated miniMECP2 in wild typemice as shown below.
TSHA-102 demonstrated regulated expression in different regions of the brain. Asshown in the graph and photos below, in the pons and midbrain, miRARE inhibitedmean MECP2 gene expression in a genotype-dependent manner as indicated bysignificantly fewer myc(+) cells observed in wild type mice compared to knockoutmice (p<0.05), thereby demonstrating that TSHA-102 achieved MECP2 expressionlevels similar to normal physiological parameters.
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In preclinical animal models, intrathecal myc-tagged TSHA-102 was not associatedwith early death and did not cause adverse behavioral side effects in wild typemice demonstrating appropriate downregulation of miniMECP2 protein expression ascompared to unregulated MECP2 gene therapy constructs. In addition, preclinicaldata demonstrated that miRARE reduced overall expression of miniMECP2 transgeneexpression and regulated genotype-dependent myc-tagged miniMECP2 expressionacross different brain regions on a cell-by-cell basis and improved the safetyof TSHA-102 without compromising efficacy in juvenile mice. Pharmacologicactivity of TSHA-102 following IT administration was assessed in the MECP2knockout mouse model of Rett syndrome across three dose levels and three agegroups (n=252). A one-time IT injection of TSHA-102 significantly increasedsurvival at all dose levels, with the mid to high doses improving survivalacross all age groups compared to vehicle-treated controls. Treatment withTSHA-102 significantly improved body weight, motor function and respiratoryassessments in MECP2 knockout mice. An additional study in neonatal mice isongoing, and preliminary data suggest normalization of survival. Finally, anIND/CTA-enabling 6-month Good Laboratory Practice, or GLP, toxicology study(n=24) examined the biodistribution, toxicological effects and mechanism ofaction of TSHA-102 when intrathecally administered to Non-Human Primates, orNHPs, across three dose levels. Biodistribution, as reflected by DNA copynumber, was observed in multiple areas of the brain, sections of spinal cord andthe DRG. Importantly, mRNA levels across multiple tissues were low, indicatingmiRARE regulation is minimizing transgene expression from the construct in thepresence of endogenous MECP2 as expected, despite the high levels of DNA thatwere delivered. No toxicity from
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transgene overexpression was observed, confirmed by functional andhistopathologic evaluations demonstrating no detrimental change inneurobehavioral assessments and no adverse tissue findings on necropsy.
In neonatal knockout Rett mice, treatment with TSHA-102 resulted in nearnormalization of survival as shown below. A single intracerebroventricular, orICV, injection of TSHA-102 at a dose of 8.8E10 vg/mouse (Human Equivalent Doseof 2.86E14 vg/participant) within 48 hours after birth in Mecp2-/Y male micesignificantly extended the survival of the animals as shown below. All cohorts,including vehicle, were sacrificed at 34 weeks. Preliminary data demonstratedapproximately 70% of the treated Mecp2-/Y males survived to 34 weeks of agecompared to 9 weeks in the vehicle-treated Mecp2-/Y male.
In addition, neonatal knockout Rett mice demonstrated normalization of behaviorfollowing treatment with TSHA-102 as assessed by the Bird Score, a compositemeasure of six different phenotypic abilities. Knockout animals were initiallyassessed at 4 weeks of age with a mean Bird Score of 4. Over the course of thestudy, TSHA-102 improved the behaviors (as assessed by the Bird aggregate score)of TSHA-102 treated mice as shown below.
In summary, we believe the totality of preclinical data generated to date,specifically including the mouse pharmacology study to ascertain minimallyeffective dose, the two toxicology studies (wild type rat and wild type NHP) andthe recent mouse neonatal data, represents the most robust package supportingclinical advancement of TSHA-102 in Rett syndrome as shown below.
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Safety and biodistribution assessments in NHPs were presented in May 2022 at theInternational Rett Syndrome Foundation (IRSF) meeting along with the caregiverperspective on Rett syndrome in adulthood. At the ASCEND National Summit, therewas an oral presentation on "Putting Patients at the Center." Finally, mousepharmacology, rat and NHP toxicology data were presented at the 25th AnnualMeeting of the American Society of Gene & Cell Therapy (ASGCT).
We submitted a CTA for TSHA-102 in November 2021 and announced initiation ofclinical development under a CTA approved by Health Canada in March 2022. We areadvancing TSHA-102 in the REVEAL Phase 1/2 clinical trial which is anopen-label, dose escalation, randomized, multicenter study that will examine thesafety and efficacy of TSHA-102 in adult female patients with Rett syndrome. Upto 18 patients will be enrolled. In the first cohort, a single 5E14 total vgdose of TSHA-102 will be given intrathecally. The second cohort will be given a1E15 total vg dose of TSHA-102. Key assessments will include Rett-specific andglobal assessments, quality of life, biomarkers, and neurophysiology and imagingassessments. Sainte-Justine Mother and Child University Hospital Center inMontreal, Quebec, Canada has been selected as the initial clinical trial siteunder the direction of Dr. Elsa Rossignol, Assistant Professor Neuroscience andPediatrics, and Principal Investigator. We expect to report preliminary clinicaldata for TSHA-102 in Rett syndrome by year-end 2022.
We have received orphan drug designation and rare pediatric disease designationfrom the FDA and orphan drug designation from the European Commission forTSHA-102 for the treatment of Rett syndrome.
TSHA-121 for CLN7 Disease
Recommendation and review posted by G. Smith
