Usher Syndrome (USH) Missense mutations can lead to proteopathies which result in the misfolding of the protein and targeted removal by endoplasmic reticulum associated degradation (ERAD) facilitated by ubiquitin. One example is the Ashkenazi form of Usher Type 3A, a recessive rare genetic disease of progressive hearing and vision loss caused by a single point mutation in the Clarin1 gene. This mutation causes a single amino acid substitution in the Clarin1 protein leading to improper protein folding and ERAD. Another example of Usher Type 3A is the Finnish mutation (Y176X), which leads to a truncation of Clarin1 and no protein production.
A small molecule has been identified which binds heat shock protein (HSP), blocks the ERAD of the Clarin1 protein, prevents degradation of the misfolded protein, and allows insertion of the misfolded Clarin1 protein in the plasma membrane.
In a reported mouse model of Usher Type 3A, a small molecule has been shown to prevent hearing loss. However, this model does not present with a vision phenotype suggesting the mouse model is not appropriate to study Usher Type 3A vision loss. We have created a rabbit model of Usher Type 3A which may be phenotypically closer to human Usher Type 3A with both hearing and vision loss. We have generated homozygous offspring and we will begin to evaluate the natural history of the age-dependent changes in hearing and vision in this rabbit model. We plan to evaluate small molecules and gene-editing approaches for effects on hearing and vision. We have also created a rabbit model of Usher Type 2A and plan to conduct similar activities as for Usher Syndrome Type 3A.
Cystic Fibrosis (CF) is an autosomal recessive disease affecting 70,000 to 100,000 people worldwide and approximately 30,000 cases each exist in the US and Europe. There are more than 2,000 known mutations in the cystic fibrosis trans-membrane conductance regulator (CFTR) gene and there are six classifications of the disease causing mutations (Class I-VI) which vary by severity of disease. CFTR regulates chloride ion transport across membrane apical epithelium cells of the airways, intestine, pancreas, kidney, sweat gland, and the male reproductive tract. CFTR also affects bicarbonate secretion, a regulator of surface liquid pH and inhibition of the epithelial sodium channel which plays a role in the hydration of secretions and mucins. CFTR mutations disrupts function of mucus and sweat producing cells affecting multiple organs and can lead to organ failure and death. Affected individuals are either homozygous (carry the same mutation on their pair of CFTR genes) or compound heterozygous (carry separate mutations on each inherited copy of the CFTR gene). Carriers are heterozygous with one mutated gene and one normal gene and are unaffected. The probability of two carriers having an affected child is 1 in 4 (25%). Gene therapy to correct one of the two affected genes, given sufficient penetration, may be an effective treatment. We have created CF rabbit models that carry the most common point mutations (the CFTR 508 mutation), as well as null mutations, where the CFTR protein is not expressed. Heterozygous carriers are unaffected, however homozygous offspring, if untreated, rapidly present with severe CF symptoms and have only a 50% chance of living beyond 44 days. We have identified a specific gene in rabbit CF that is up regulated and is the target of a small molecular entity under investigation at GeneToBe. We have also discovered that this gene is over expressed in all human CF conditions, including point and null mutations that we have studied to date, but this gene is not up-regulated in normal human or rabbit subjects. We have discovered a registered pharmaceutical small molecule for an unrelated condition that can be re-purposed to alleviate symptoms in our CF rabbits, and markedly prolongs life expectancy. We plan to conduct additional studies with this pharmaceutical agent in additional CF rabbits. Furthermore, we plan to evaluate this pharmaceutical agent in CF patients in a randomized, double-blind, placebo-controlled study. We also plan to evaluate the safety and efficacy of our gene editing Mi-Cas9 vectors as a potential cure in the CF rabbit models.
Creation of Animal Models of Rare Genetic Diseases
GeneToBe considers animal models for developing therapies for common and rare genetic diseases. GeneToBe scientists have a rich history in the creation of mouse models, including the insertion of genes (Transgenic models), removing a gene (Knockout models) and replacing genes (Knockout/Knockin models). Using appropriate chemically sensitive promotor elements, expression or repression of transgenic genes can be regulated chemically and are useful in some models. Using appropriate promotors, ectopic expression of genes, which is useful to study certain conditions, can also be achieved. Although mouse models can be created rapidly, colonies quickly expanded, and do not require a large amount of space to house, they may not be translational for all diseases. we have created rabbit models for our current programs as they better translate to the human disease condition. Although rabbit models are less economical and take longer to develop, they are translational, making them appropriate for therapeutic discovery and development for the current programs.