SYST-13 AAV VECTOR ENCODING FC MODIFIED HER2 ANTIBODIES REDUCES TUMOR BURDEN WITH MINIMIZED PERIPHERAL EXPOSURE IN MODELS OF BREAST CANCER BRAIN METASTASIS

Abstract

Abstract We engineered a HER2 Ab that inhibited HER2+ breast cancer cell growth in vitro and led to a profound increase in Ab-dependent CD16A receptor stimulation relative to trastuzumab, the standard of care. Blood-brain barrier penetrant AAV vectors were administered intravenously to mice orthotopically engrafted with HER2+ cancer cells. In both prophylactic AAV administration and a treatment model where AAV was dosed post-engraftment, the vectorized Ab significantly reduced tumor burden and prolonged survival. We employed single-cell RNA-Seq to characterize immune responses to these vectorized Abs. Upon dissociation of brains from tumor engrafted mice that underwent intravenous AAV treatment, we performed transcriptomic profiling of CD45+ immune cells. Our results indicate a pronounced innate immune response to the HER2 Ab that increased tumor-proximal natural killer cells and proliferating microglia. Response to gene therapy also included dendritic cells that are mediators of adaptive immune activation. Therefore, the adaptive immune system could augment the efficacy of the therapy when non-exhausted, tumor-associated T lymphocytes are present. To minimize potential neuronal toxicity, we evaluated the use of a GFAP promoter for astrocyte specific expression. Furthermore, to reduce patient risk of cardiac toxicity induced by antibody exposure, we engineered the Fc domain of the antibody so that it would not bind to FcRN, a receptor which promotes antibody recycling and subsequent increased antibody levels in the periphery. In mouse studies, treatment with a BBB-penetrant capsid with a GFAP driven construct and with binding to FcRN eliminated resulted in a 96% reduction in serum antibody levels, to 84% below an estimated cardiac toxicity threshold. Additionally, there was a 78% reduction in antibody levels in the heart, to 98% below an estimated cardiac toxicity threshold. Remarkably, brain levels were maintained. Our animal data indicate that vectorized HER2 antibody gene transfer may address unmet medical needs of patients.