Design and validation of a GMP stem cell manufacturing protocol for MPSII hematopoietic stem cell gene therapy

Hematopoietic stem cell gene therapy (HSCGT) is a promising therapeutic strategy for the treatment of neurodegenerative, metabolic disorders. The approach involves the ex vivo introduction of a missing gene into patients’ own stem cells via lentiviral-mediated transduction. Once transplanted back into a fully conditioned patient, these genetically modified HSCs can repopulate the blood system and produce the functional protein, previously absent or non-functional in the patient, which can then cross-correct other affected cells in somatic organs and the central nervous system. We previously developed a HSCGT approach for the treatment of Mucopolysaccharidosis type-II (MPSII, Hunter Syndrome), a debilitating paediatric lysosomal disorder caused by mutations in the iduronate-2-sulphatase (IDS) gene, leading to accumulation of heparan and dermatan sulphate which causes severe neurodegeneration, skeletal abnormalities and cardiorespiratory disease. In HSCGT proof-of-concept studies using lentiviral IDS fused to a brain-targeting peptide ApoEII (IDS.ApoEII), we were able to normalise brain pathology and behaviour of MPSII mice. Here we present an optimised and validated good manufacturing practice (GMP) hematopoietic stem cell transduction protocol for MPSII in preparation for first-in-man studies. Inclusion of transduction enhancers LentiBOOST™ and protamine sulphate significantly improved transduction efficiency by at least 3-fold without causing adverse toxicity, thereby reducing vector quantity required.