84. Pharmacology/Toxicology Studies for Gene-Modified Hematopoietic Stem Cell Transplantation for Cystinosis

Cystinosis is an autosomal metabolic disease belonging to the family of lysosomal storage disorders. Mutations in the CTNS gene, encoding a lysosomal cystine transporter, lead to cystine accumulation and multi-organ failure such as blindness, myopathy, diabetes and central nervous system defects. Affected individuals also develop proximal tubulopathy and eventually progress to end stage renal failure. Treatment is available with the drug cysteamine to reduce intracellular cystine content. However, cysteamine only delays the progression of the disease.In April 2012, the first clinical trial using stem cells for cystinosis was approved as an allogeneic Hematopoietic Stem and Progenitor Cell (HSPC) transplant to be conducted at University of California, Los Angeles. This trial was based on our preclinical studies in the mouse model of cystinosis, Ctns-/- mice, which showed that transplantation of HSPCs expressing a functional Ctns gene led to the abundant tissue integration of bone marrow-derived cells, a significant decrease of cystine accumulation and kidney preservation. In order to avoid the high mortality and morbidity associated with allogeneic transplants, however, we have developed an autologous transplantation strategy of HSPCs genetically modified ex vivo to express a functional CTNS gene as a treatment for cystinosis. Preclinical studies using a SIN-lentivirus vector containing CTNS to transduce Ctns-/- HSPCs and transplanted in Ctns-/- mice were promising leading to cystine reduction in all tissues and kidney function improvement.We are currently performing pharmacological and toxicological studies using a batch of pCCL-CTNS lentiviral vector preparation produced under comparable Good Manufacturing Practice, the results of which will be included in an IND for a phase 1 clinical trial for cystinosis. We have now completed the in vitro studies, establishing the optimal conditions to transduce human CD34+ hematopoietic stem cells with our lentiviral vector to obtain a Vector Copy Number (VCN) included between 1 and 3. We performed Colony Forming Unit (CFU) assays using human CD34+ PBSC isolated from five healthy donors and four cystinotic patients and neither showed aberrant proliferation or differentiation potential with the lentivirus compared to negative controls. Additionally, the In Vitro Immortalization (IVIM) assay, a genotoxicity test, did not produce immortalized clones, thus demonstrating an excellent safety profile. Lastly, we have in progress in vivo serial transplantation experiments of transduced murine Ctns-/- HSPCs to assess the safety of this strategy in vivo by comprehensive molecular, clinical and histological analyses. This work represents the first stem cell and gene therapy treatment strategy for cystinosis. If successful, this treatment could be a proof of concept for other degenerative multi-systemic disorders.