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1Tuberous Sclerosis Alliance, Silver Spring, MD, USA; 2Translational EEG, PsychoGenics, Tarrytown, NY, USA;
3PsychoGenics, Montvale, NJ, USA;
4Dept. of Neurol., Washington Univ. Sch. of Med., St. Louis, MO, USA;

Creating a preclinical consortium was among the highest priorities arising from a 2015 NIH-sponsored workshop to update the TSC research strategy. A consortium was needed to enable testing candidate molecules or pharmacological tools in a rigorous, standardized way using translatable animal models.

The TS Alliance gathered input from researchers on various animals and assays used to model features of TSC that most impact patients: epilepsy, tumors, and neuropsychiatric issues. The TS Alliance designed a consortium structure, raised funds, and put contracts in place to get the work done.

The TS Alliance established a consortium structure in which the TS Alliance funds much of the work and serves as the administrative center. The TSC Preclinical Consortium Steering Committee guides overall priorities. Much of the study design of the consortium is done at the level of Working Groups, composed primarily of academic researchers who can evaluate data to recommend which models and endpoints are best suited for preclinical translational studies.

To begin, consortium members evaluated data from several TSC genetic mouse models that develop epilepsy. Among these, Tsc1flox / flox;GFAP-Cre+ (Tsc1GFAPCKO) mice have a robust, fully penetrant seizure phenotype, with seizures typically beginning between 4-7 weeks of age. The TS Alliance obtained licenses to these mice to enable testing of drugs for academic researchers and for-profit companies, ensuring the Preclinical Consortium can be used both to advance publishable TSC research and to accelerate the development of new treatments by commercial entities. As a result, six pharma and biotech companies have paid a membership fee to join the consortium since August 2016. The TS Alliance also established agreements with a contract research organization to do the laboratory work.

To demonstrate reproducibility of the model and build upon published data, we evaluated incidence of seizures using EEG and changes in mRNA and protein biomarkers in Tsc1GFAPCKO mice. Vehicle-treated Tsc1GFAPCKO mice exhibited robust electrographic seizures and significantly increased mortality. Treatment with rapamycin beginning at 3 weeks of age completely prevented the development of seizures and mortality during the study period. Tsc1GFAPCKO mice demonstrated increased expression in brain of transcripts of relevant genes including IBA1, CD68, ICAM1, and VEGF-D. Tsc1GFAPCKO mice also had increased phosphorylation in brain of mTOR (Ser2448) and S6 (Ser235/236), consistent with activation of the mTOR pathway in these mice.

Our initial experiments successfully demonstrated that we can independently measure seizures in Tsc1GFAPCKO mice consistent with the published phenotype. Additionally, we found that rapamycin was effective at the same doses as those previously published to prevent seizures and premature death in these mice. Having demonstrated this model is highly reproducible and robust, our findings show the clear utility of using these mice to screen new potential anti-epileptic therapeutics for TSC. Furthermore, our identification of mRNA and phospho-protein biomarkers in these mice opens the potential for research to ask whether similar biomarkers are altered and measurable in blood from individuals with TSC and may be useful for translational studies. The consortium is also establishing models for other TSC phenotypes, including tumor growth and neuropsychiatric disorders.

Establishing this consortium addressed one of the highest priorities from the 2015 TSC research strategy workshop. The consortium accelerates the testing of ideas for new treatments by obviating the need for researchers to establish an animal model and assay in their own laboratory. If the data will be published, the consortium also saves researchers from finding the funding, as the TS Alliance will pay for testing molecules with sufficient rationale and drug-like properties. Academic or industry investigators with hypotheses about the therapeutic potential of new mechanisms can nominate compounds for testing.

Additionally, the ability of industry researchers to access robust and reproducible models has attracted the interest and investment of companies. Industry members can pay for confidential testing of proprietary compounds and retain ownership of the data. This encourages companies to test their new or repurposed compounds in models relevant to TSC, which we anticipate will increase the number of future clinical trials and, hopefully, the number of approved treatments for individuals affected by TSC.