Targeting post-translational modifications of B7-H4 in carcinogenesis and therapy the goal of this project is to determine the impact of interplay between glycosylation and ubiquitination of B7-H4…

Targeting post-translational modifications of B7-H4 in carcinogenesis and therapy the goal of this project is to determine the impact of interplay between glycosylation and ubiquitination of B7-H4 in breast carcinogenesis and anti-breast cancer therapy. B7-H4, also known as v-set domain containing t cell activation inhibitor 1 (VTCN1), acts as an immune checkpoint protein whose local abnormal accumulation is correlated with poor prognosis of various types of cancers including triple negative breast cancer (tnbc). While a current tcga study suggests the critical role of B7-H4 in conferring tumor sensitivity to chemotherapy drugs and immune checkpoint inhibitors, how B7-H4 is regulated and how its deregulation contributes to breast carcinogenesis and tumor drug response remain unknown. Results from our recent purification of the B7-H4 protein complex revealed that 2 critical enzymes, amfr and STT3 complex, tightly interact with and regulate B7-H7. While the E3 ubiquitin ligase amfr catalyzes ubiquitination of B7-H4 followed by degradation, we observed that glycosylation of B7-H4 protein by the STT3 complex (a glycosyltransferase) results in the stabilization of B7-H4 and inhibition of doxorubicin-induced immunogenic cell death (ICD). Elevation of B7-H4 levels due to enhanced B7-H4 glycosylation counteracts B7-H4 ubiquitination, which in turn suppresses endoplasmic reticulum stress-mediated phagocytosis in response to chemotherapy drugs, a critical step in triggering mass tumor eradication by ICD. We further demonstrated that blockade of B7-H4 glycosylation by NGI-1, a novel protein glycosylation inhibitor, significantly enhances B7-H4 ubiquitination and subsequent degradation, resulting in promotion of tumor killing efficacy due to increased phagocytosis by dendritic cells and their capacity to elicit CD8+ interferon – producing t cell responses. In this project, we plan to test the hypothesis that deregulation of B7-H4 by the crosstalk between glycosylation and ubiquitination promotes tnbc tumor evasion from chemotherapy and blockade of B7-H4 glycosylation by NGI-1 could be a new strategy for anti-tnbc treatment. We propose the following specific aims to pursue this goal: (1) to determine the mechanism by which interplay between glycosylation and ubiquitination regulates B7-H4-orchestrated phagocytosis; (2) to determine the physiological relevance of STT3-mediated glycosylation and amfr-facilitated ubiquitination in B7-H4-mediated ICD in response to chemotherapy drugs; and (3) to determine the clinical relevance of blockade of B7-H4 glycosylation by NGI-1 in anti-tnbc treatment using various preclinical murine models.