Efficacy of Enzyme-T cells for long-term restoration of function in Mucopolysaccharidosis Type 1 - ABSTRACT Lysosomal Storage Disorders (LSDs) are a diverse group of over 70 inherited metabolic diseases caused by loss-of-function mutations in lysosomal enzymes, leading to progressive accumulation of undegraded substrates in cells and tissues. Many LSDs, including Mucopolysaccharidosis Type I (MPS I, Hurler Syndrome), are life- limiting conditions with multisystem involvement and central nervous system (CNS) pathology. Current treatments such as enzyme replacement therapy (ERT) and hematopoietic stem cell transplantation (HSCT) offer only partial benefit. ERT requires lifelong infusions, does not cross the blood-brain barrier (BBB), and is costly. HSCT can slow neurological decline but carries risks including graft-versus-host disease, infection, and mortality. Moreover, over 80% of LSDs still have no approved therapies, representing a significant unmet need. To overcome these limitations, we are developing a transformative platform using genetically engineered human T cells, termed Enzyme-T cells, designed to systemically and durably deliver functional lysosomal enzymes. T cells persist long-term, traffic to peripheral tissues and the CNS, and are well suited for scalable manufacturing. We engineer Enzyme-T cells using the TcBuster (TcB) transposon system, a clinical-stage non-viral gene delivery method, to stably express α-L-iduronidase (IDUA), the enzyme deficient in MPS I. Our published preclinical studies show that IDUA-T cells secrete active enzymes, cross-correct deficient cells, and are well tolerated. In the R61 phase, we will (1) engineer T cells from MPS I patients to stably express IDUA and a safety marker (EGFRt); (2) characterize enzyme secretion, immune phenotype, and genomic safety including transposon copy number and integration site analysis; (3) assess in vitro cross-correction using patient-derived cells and iPSC-derived IDUA-deficient lines; and (4) develop a quantitative systems pharmacology (QSP) model to predict biodistribution, enzyme kinetics, and dosing. In the R33 phase, we will evaluate the in vivo efficacy of IDUA-T cells in an MPS I mouse model, assessing exposure-response, tissue enzyme levels, cognitive function, and toxic glycosaminoglycan (GAG) reduction. Histopathological analysis will assess biodistribution, persistence, and safety. This multidisciplinary project combines MPS clinical expertise, advanced genome engineering, and computational modeling to develop a novel, durable, and CNS-penetrant therapy. If successful, our Enzyme-T cell platform will represent a paradigm shift in the treatment of MPS I and other LSDs with few or no effective options.
