Accumulating evidence suggests that aerobic glycolysis is important for colorectal cancer (CRC) development. However, the underlying mechanisms have yet to be elucidated. B7-H3, an immunoregulatory protein, is broadly overexpressed by multiple tumor types and plays a vital role in tumor progression. In this study, we found that overexpression of B7-H3 effectively increased the rate of glucose consumption and lactate production, whereas knockdown of B7-H3 had the opposite effect. Furthermore, we showed that B7-H3 increased glucose consumption and lactate production by promoting hexokinase 2 (HK2) expression in CRC cells, and we also found that HK2 was a key mediator of B7-H3-induced CRC chemoresistance. Depletion of HK2 expression or treating cells with HK2 inhibitors could reverse the B7-H3-induced increase in aerobic glycolysis and B7-H3-endowed chemoresistance of cancer cells. Moreover, we verified a positive correlation between the expression of B7-H3 and HK2 in tumor tissues of CRC patients. Collectively, our findings suggest that B7-H3 may be a novel regulator of glucose metabolism and chemoresistance via controlling HK2 expression in CRC cells, a result that could help develop B7-H3 as a promising therapeutic target for CRC treatment.

Core 1- and core 3-derived mucin-type O-glycans are primary components of the mucus layer in the colon. Reduced mucus thickness and impaired O-glycosylation are observed in human ulcerative colitis. However, how both types of O-glycans maintain mucus barrier function in the colon is unclear. We found that C1galt1 expression, which synthesizes core 1 O-glycans, was detected throughout the colon, whereas C3GnT, which controls core 3 O-glycan formation, was most highly expressed in the proximal colon. Consistent with this, mice lacking intestinal core 1-derived O-glycans (IEC C1galt1−/−) developed spontaneous colitis primarily in the distal colon, whereas mice lacking both intestinal core 1- and core 3-derived O-glycans (DKO) developed spontaneous colitis in both distal and proximal colon. DKO mice showed an early onset and more severe colitis than IEC C1galt1−/− mice. Antibiotic treatment restored the mucus layer and attenuated colitis in DKO mice. Mucins from DKO mice were more susceptible to proteolysis than WT mucins. This study indicates that core 1- and 3-derived O-glycans collectively contribute to the mucus barrier by protecting it from bacterial protease degradation and suggests new therapeutic targets to promote mucus barrier function in colitis patients.

Tumor angiogenesis is a hallmark of cancer and is involved in the tumorigenesis of solid tumors. B7-H3, an immune checkpoint molecule, plays critical roles in proliferation, metastasis and tumorigenesis in diverse tumors; however, little is known about the biological functions and molecular mechanism underlying B7-H3 in regulating colorectal cancer (CRC) angiogenesis. In this study, we first demonstrated that the expression of B7-H3 was significantly upregulated and was positively associated with platelet endothelial cell adhesion molecule-1 (CD31) level in tissue samples from patients with CRC. In addition, a series of in vitro and in vivo experiments showed that conditioned medium from B7-H3 knockdown CRC cells significantly inhibited the migration, invasion, and tube formation of human umbilical vein endothelial cells (HUVECs), whereas overexpression of B7-H3 had the opposite effect. Furthermore, B7-H3 promoted tumor angiogenesis by upregulating VEGFA expression. Recombinant VEGFA abolished the inhibitory effects of conditioned medium from shB7-H3 CRC cells on HUVEC angiogenesis, while VEGFA siRNA or a VEGFA-neutralizing antibody reversed the effects of conditioned medium from B7-H3-overexpressing CRC cells on HUVEC angiogenesis. Moreover, we verified that B7-H3 upregulated VEGFA expression and angiogenesis by activating the NF-κB pathway. Collectively, our findings identify the B7-H3/NF-κB/VEGFA axis in promoting CRC angiogenesis, which serves as a promising approach for CRC treatment. Facts B7-H3 is significantly upregulated and is positively associated with CD31 level in colorectal cancer (CRC) tissue samples. B7-H3 modulates tumor angiogenesis by upregulating vascular endothelial growth factor A (VEGFA) expression in CRC cells. VEGFA is critical for B7-H3-mediated CRC angiogenesis both in vitro and in vivo. B7-H3 promotes VEGFA expression and angiogenesis by activating NF-κB signaling.

BACKGROUND & AIMS Core 1- and 3-derived mucin-type O-linked oligosaccharides (O-glycans) are major components of the colonic mucus layer. Defective forms of colonic O-glycans, such as Tn antigen, are frequently observed in patients with ulcerative colitis and colorectal cancer, but it is not clear if they contribute to pathogenesis. We investigated whether and how impaired O-glycosylation contributes to development of colitis-associated colorectal cancer using mice lacking intestinal core 1- and 3-derived O-glycans. METHODS We generated mice that lack the core 1- and 3-derived intestinal O-glycans (DKO mice) and analyzed them, along with mice that lack the intestinal epithelial core 1 O-glycans (IEC C1galt1−/− mice) or mice that lack core 3 O-glycans (C3Gnt−/− mice). Intestinal tissues were collected at different time points and analyzed for levels of mucin and Tn antigen, development of colitis, and tumor formation using imaging, quantitative PCR, immunoblot, and ELISA techniques. We also used cellular and genetic approaches, as well as intestinal microbiota depletion, to identify inflammatory mediators and pathways that contribute to disease in DKO and wild-type littermates (controls). RESULTS Intestinal tissues from DKO mice contained higher levels of Tn antigen and had more severe spontaneous chronic colitis than tissues from IEC C1galt1−/− mice, whereas spontaneous colitis was absent in C3GnT−/− and control mice. IEC C1galt1−/− mice and DKO mice developed spontaneous colorectal tumors, although the onset of tumors in the DKO mice was earlier (age 8–9 months) than that in IEC C1galt1−/− mice (around age 12 months). Antibiotic depletion of the microbiota did not cause loss of Tn antigen but did reduce the development of colitis and cancer formation in DKO mice. Colon tissues from DKO mice, but not control mice, contained active forms of caspase-1 and increased caspase-11, which were reduced after antibiotic administration. Supernatants from colon tissues of DKO mice contained increased levels of interleukin-1β and interleukin-18, compared to those from control mice. Disruption of the caspase 1 and caspase 11 genes in DKO mice (DKO/Casp1/11−/− mice) decreased development of colitis, characterized by reduced colonic thickening, hyperplasia, and inflammatory infiltrate, compared with DKO mice. CONCLUSIONS Impaired expression of O-glycans causes colonic mucus barrier breach and subsequent microbiota-mediated activation of caspase 1-dependent inflammasomes in colonic epithelial cells of mice. These processes could contribute to colitis-associated colon cancer in humans.