Cytosine deaminase base editing to restore COL7A1 in dystrophic epidermolysis bullosa human:murine skin model

Recessive dystrophic epidermolysis bullosa (RDEB) is a debilitating blistering skin disorder caused by loss-of-function mutations in COL7A1 encoding type VII collagen (C7), the main component of anchoring fibrils (AFs) at the dermal-epidermal junction (DEJ). Although conventional gene therapy approaches through viral vectors have been tested in pre-clinical and clinical trials, they are limited by transgene size constraints and only support unregulated gene expression. Genome editing could potentially overcome some of these limitations, and CRISPR/Cas9 has already been applied in research studies to restore COL7A1 expression. Delivery of suitable repair templates for repair of DNA cleaved by Cas9 is still major challenge, and alternative base editing strategies may offer corrective solutions for certain mutations. We demonstrate highly targeted and efficient cytidine deamination and molecular correction of a defined RDEB mutation (c.425A>G) leading to restoration of full-length C7 protein expression in primary human fibroblasts and iPSCs. C7 basement membrane expression and skin architecture were restored with de novo AFs identified by electron microscopy in base edited human RDEB grafts recovered from immunodeficient mice. The results demonstrate the potential and promise of emerging base editing technologies in tackling inherited disorders with well-defined single nucleotide mutations

Castration-mediated IL-8 promotes myeloid infiltration and prostate cancer progression

Unlike several other tumor types, prostate cancer (PCa) rarely responds to immune checkpoint blockade (ICB). To define tumor-cell intrinsic factors that contribute to PCa progression and resistance to ICB, we analyzed PCa epithelial cells from castration-sensitive and castration-resistant samples using implanted tumors, cell lines, transgenic models, and human tissue. We found that castration resulted in increased expression of Interleukin-8 (IL-8) and its likely murine homolog Cxcl15 in prostate epithelial cells. We showed that these chemokines drove subsequent intra-tumoral infiltration of tumor-promoting polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs), which was largely abrogated when IL-8 signaling was blocked genetically or pharmacologically. Targeting IL-8 signaling in combination with ICB delayed the onset of castration-resistance and increased the density of polyfunctional CD8 T cells in tumors. Our findings establish a novel mechanism by which castration mediates IL-8 secretion and subsequent PMN-MDSC infiltration and highlight blockade of the IL8/CXCR2 axis as a potential therapeutic intervention