Abstract 4880: A novel heparan-sulfate (HS) oligosaccharide on tumor-blood vessels is a target for anti-immune vascular therapy in renal cell carcinoma (RCC)

Abstract In ccRCC, tumor cells are arranged in nests surrounded by capillary endothelial cells (EC) abundant in HS polysaccharide proteoglycan receptors (HSPGs). HS structure is highly heterogeneous, due to HS sulfotransferases (HSST) that randomly sulfate at different positions of the core disaccharides. Specific sulfation pattern determines the avidity of HS to soluble factors, which regulates the interaction between EC and leukocytes. We identified 6-O-highly sulfated HS 8-mer saccharides (6O-HS) to be the ligand of GPNMB receptor that inhibits T cell activation and induces angiogenesis. Using our 6O-HS-specific 1A7 mAb, we examined the expression of 1A7 epitope in RCC. IHC staining of RCC specimens (n=4) showed 1A7 mAb stained some but not all EC and nearly no staining in tumor cells. We examined this EC selectivity by identifying 6O-HS+ HSPGs using mouse SVEC line expressing HSPGs (syndecans (SD), CD44). Surprisingly FACS analysis showed very little 1A7 expression on these cells. To ask whether this is due to low expression of HS6ST1 (for 6-O-sulfation), HS6ST1-transfected SVEC cells were stained with 1A7 and HSPG Ab and found all HSPGs positive for 1A7. To examine whether binding of GPNMB to 1A7+ HSPG induces specific signaling, SD4/HS6ST1-transfected 293T cells were treated with GPNMB-Fc and measured Tyr-phosphorylation (p-Tyr) on SD4 protein. Western blotting showed GPNMB-Fc induced p-Tyr on SD4 in 10 min post-incubation and the signaling was completely blocked by pretreatment of transfectants with 1A7 mAb. We assayed the effects of 1A7 mAb on T cells and EC. For T cells, we assayed in vitro suppressor function of GPNMB+ myeloid-derived suppressor cells (MDSC from RCC patients, n=3) to autologous T cells. In coculture of T cells/MDSC, 1A7 mAb inhibited IFN-γ T cell response by 50-84% (p>0.01) and 40-60% T cells were 1A7+. For EC, we used in vivo angiogenesis assays: 1A7 mAb was injected into GPNMB-Fc-matrigel plugs in mice and 2 dpf zebrafish embryos. This mAb (n=5, 10 μg/ml) reversed GPNMB’s angiogenic activity completely in mice and inhibited growth of the subintestinal vessels (SIV) (n=9, 50% reduction in SIV # by 2 μg/ml, p<0.001). To evaluate the efficacy of 1A7 mAb to treat RCC, BALB/c mice bearing RenCa tumor were treated with 1A7 or PD1 mAb; 200 μg i.p./mouse, 2x/wk. 1A7 mAb inhibited tumor growth as greatly as PD1 Ab (64% inhibition by 1A7 vs. 61% by PD1 Ab, p<0.01, n=6) with a reduction in tumor density of microvessels and myeloid cells (50-60% reduction) and a rise in T cells (∼5-fold). Similarly, 1A7 mAb inhibited human RCC growth in Caki-2-derived xenograft NSG mice injected with human PBMC (49% inhibition by 1A7 vs. 37% by Nivolumab, p=0.005). 6O-HS+ renal EC may regulate T cell activation/ trafficking into RCC and facilitates tumor angiogenesis via the high avidity to signaling factors. Thus, 6O-HS-targeting may be a promising anti-angiogenic immunotherapy for RCC. Citation Format: Jin-Sung Chung, Nilambari Pawar, Ponciano D. Cruz, Hans Hammers, Kiyoshi Ariizumi. A novel heparan-sulfate (HS) oligosaccharide on tumor-blood vessels is a target for anti-immune vascular therapy in renal cell carcinoma (RCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4880.