Overall Research Objectives

Our research focuses on developing stem cell and gene therapy strategies for degenerative multi-systemic disorders, and to understand the mechanisms by which hematopoietic stem and progenitor cells (HSPC) can lead to tissue repair in non-hematopoietic genetic diseases.

Summary of the main Research Projects in the lab

Hematopoietic Stem Cell Gene Therapy 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. We showed that Hematopoietic Stem and Progenitor Cell (HSPC) transplant in the mouse model of cystinosis, the Ctns-/- mice, led to the abundant tissue integration of bone marrow-derived cells, significant decrease of tissue cystine accumulation, and long-term kidney, eye and thyroid preservation. However, allogeneic transplants are associated with high risks of mortality and morbidity. Thus, our long-term goal is to develop 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. Transduced cells were capable of reducing cystine in all tissues and of improving kidney function. We are currently performing the preclinical pharmacological and toxicological testing following the FDA requirements and assembling an Investigational New Drug (IND) application for a phase ½ clinical trial for cystinosis. 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.

Mechanism of Hematopoietic Stem Cell-mediated Therapy in Cystinosis

The extent of efficacy of HSPCs to rescue cystinosis was surprising especially considering that cystinosin is a transmembrane lysosomal protein. The study of the mechanism by which Ctns-expressing HSPCs leads to tissue repair in the Ctns-/- mice showed that a large subset of HSPCs differentiated into macrophages that can transfer cystinosin-bearing lysosomes to the deficient host cells via long tubular extensions known as tunneling nanotubes (TNTs). Conversely, diseased cells also exploited the same route to transfer cystine-loaded lysosomes to the macrophages, providing a bidirectional correction mechanism. Cellular stress from the Ctns-deficient cells stimulated the formation of the TNTs. While cross-correction, either upon secretion-recapture after bone marrow transplantation was shown in several lysosomal storage disorders caused by defective soluble hydrolases, our study is the first demonstration of cross-correction in the context of a lysosomal transmembrane protein and of TNTs as key cellular device in the transfer. We also showed for the first time that TNTs could cross the thick, dense and stiff renal tubular basement membrane in vivo and transfer cystinosin-bearing lysosomes to the proximal tubular cells, providing a mechanism underlying the long-term kidney preservation after HSPC transplantation in the Ctns-/- mice. We also showed that the mechanism of HSPC-mediated therapeutic action was similar for the ocular defects and for the thyroid rescue in the Ctns-/- mice. Our objectives are now to investigate the molecular protagonists and activating signal(s) involved in the TNT formation as well as the phenotype of the macrophages involved in tissue repair.

Hematopoietic Stem Cell Gene Therapy for Friedreich’s Ataxia

Because mitochondria can also be transferred via TNTs, we hypothesized that HSPC transplantation could also treat mitochondrial diseases. We thus tested this hypothesis in a mouse model of Friedreich’s Ataxia (FRDA), the YG8R mice. FRDA is an autosomal recessive mitochondrial disease characterized by neurodegeneration, cardiomyopathy and muscle weakness caused by reduced expression of the mitochondrial protein frataxin. There is no treatment for this debilitating and lethal disorder. We showed that HSPC transplantation in the YG8R mice completely prevented the development of the locomotor deficits and muscle weakness. Degeneration of the large sensory neurons in the dorsal root ganglia (DRG) was fully prevented in the HSPC-transplanted YG8R mice as well as mitochondrial dysfunction in brain, skeletal muscle and heart. Abundant GFP+ HSPC-derived cells were observed in tissues as differentiated phagocytic cells such as microglial cells in the brain and spinal cord, and macrophages in DRGs, heart and skeletal muscle. Finally, we observed in vivo transfer of frataxin-GFP and cox8-GFP mitochondrial proteins from HSPC-derived microglia/macrophages to diseased neurons and cardiac/muscular myocytes. Our objective is now to develop an HSPC gene therapy approach for this disease.

Kidney-targeted Gene Delivery Using AAV

A wide range of monogenic kidney disorders has been identified, and so far, no gene therapy approach has been developed to target specifically the kidney whereas renal transplantation is associated with significant morbidity and mortality. Moreover, due to the severe shortage of donor organs, patients may wait three to six years for transplantation. The main goal of our project is to develop an efficient and minimally invasive kidney-targeted gene delivery system using recombinant Adeno-Associated Viruses (rAAV). We recently optimized this strategy via retrograde renal vein injection of rAAV serotype 9 that have the potential of transducing a wide range of renal cells. We now propose to test this approach based on renal vein injection of rAAV-CTNS as a minimally invasive procedure for treating the renal dysfunction in cystinosis. If successful, this strategy may be used in many monogenic hereditary nephropathies.