Targeting glycosylation pathway as a novel therapeutic intervention

About This Grant

PROJECT SUMMARY Metabolic reprogramming is a hallmark of cancer and potential opportunity for therapeutic intervention. The study of cancer metabolism, to its detriment, has largely focused on central carbon metabolism, such as glycolysis, the pentose phosphate pathway, and the citric acid cycle. Accordingly, less attention has been given to investigating the functional roles of nitrogen metabolism, especially how carbon and nitrogen metabolism are interconnected, and its impact on tumorigenesis. Glycosylation presents an excellent model to broaden the field by studying how the interplay of carbon (glucose) and nitrogen metabolism (amino acids and nucleotides) contributes to tumor aggressiveness through the production of amino-sugar/nucleotide sugar products. Accumulation of these products facilitates the abnormal glycosylation of cancer cells, often leading to inhibition of the immune process. Thus, targeting aberrant glycosylation may synergize with immune checkpoint blockers to improve therapeutic efficacy. To identify gene(s) critical for survival of human non-small cell lung cancer (NSCLC) with KRAS/LKB1 co- mutations (KL), a highly aggressive molecular subtype of NSCLC, compared to those with KRAS mutations (K), we performed a genome-wide CRISPR knockout screening using an isogenic pair of K cells with or without LKB1 loss. Based on integrative analysis using MAGeCK ranking algorithms and subsequent validation assays, a gene encoding ALG5 dolichyl-phosphate beta-glucosyltransferase (ALG5) in the N-glycosylation biosynthesis pathway emerged as the top candidate responsible for KL NSCLC survival and proliferation. By establishing both molecular and metabolic platforms to measure metabolites and glycan species involved in KL proliferation, and utilizing clinically relevant mouse models for in vivo studies, we are now poised to define the oncogenic role of ALG5 during lung tumorigenesis and aggressiveness. In Aim 1, we will interrogate the mechanistic basis of ALG5 dependence in KL NSCLC cells. In Aim 2, we will investigate the molecular mechanism by which LKB1 regulates ALG5. In Aim 3, we will examine 1) whether ALG5 suppression reduces tumor growth in vivo and 2) whether ALG5 suppression-mediated alteration in N- glycans can perturb tumor cell-immune cell interaction, enhancing immune cell infiltration and create ‘immune- hot’ conditions. While the role of O-GlcNAcylation in cancer growth, another arm downstream of amino- sugar/nucleotide sugar metabolism pathway, has been reported, the importance of N-glycosylation metabolism in KL NSCLC has yet to be elucidated. By combining cell biology, state-of-the art glycan mapping techniques, spatial single cell transcriptomics and in vivo studies, this work will provide comprehensive insight into the vulnerability of KL NSCLC tumors to inhibition of ALG5 and potentially illuminate novel and selective treatment strategies.