Abstract 6967: Role of CXCL7 in colon cancer cell proliferation
Authors: Aldhowayan Hadeel , Nipa Jannatul F. , Jun Lauren , Odeniyi Ifeoluwa , Jones Parker L. , Jeganathan Ramesh , Lipke Elizabeth A. , Greene Michael W. 
Published: March 2024

Chemokine (C-X-C motif) ligand 7 (CXCL7) - a protein product of the pro platelet basic protein (PPBP) gene - is a potential biomarker for colorectal cancer (CRC) diagnosis. However, its role in CRC progression is not known. Hence, we examined the role of CXCL7 in colon cancer cell proliferation through enhanced aerobic glycolysis. We first examined the effect of CXCL7 on cellular proliferation and glycolytic flux using human colon cancer (HT-29) cells transfected with CXCL7 or an empty vector as a control. Stable CXCL7- and vector-expressing cells were selected using hygromycin B. In addition, we treated HT-29 cells with CXCL7 obtained from the conditioned media (CM) from CXCL7-transfected HEK-293T cells or CM from vector-transfected (control) cells. CXCL7 expression was confirmed by analyzing PPBP expression using RT-qPCR, and CXCL7 level in the conditioned media was quantified by ELISA. The specificity of CXCL7 actions were examined using a CXCL7 neutralizing antibody (R&D Systems) and an antagonist (SB225002) to C-X-C motif chemokine receptor 2 (CXCR2). Cancer cell proliferation was assessed by measuring DNA synthesis (EdU incorporation). Glycolytic flux was monitored by measuring glucose uptake using a fluorescent derivative of 2-deoxyglucose (2-NBDG), Seahorse glycolysis stress test analysis, lactate accumulation in the cultured cell’s media, and Western blotting for glycolytic enzyme protein expression. To further investigate CXCL7-mediated proliferation, we examined EdU incorporation and colony size in 3D engineered tissues and tumor xenograft growth in nude mice injected with CXCL7- and vector-expressing HT29 cells. The proliferation rate in CXCL7-expressing HT-29 cells and HT-29 cells treated with the CXCL7-CM were significantly higher (1.2-fold and 1.8-fold, respectively) compared to controls (p&lt;0.01). Consistently, the proliferation rate in CXCL7-expressing HT-29 3D engineered tissues was significantly enhanced compared to vector-expressing tissues (p&lt;0.05). Importantly, blocking the CXCL7/CXCR2 axis using a CXCL7 neutralizing antibody and a CXCR2 inhibitor abrogated CXCL7-stimulated proliferation. Lactate levels were two-fold higher in the culture media from both CXCL7-expressing HT-29 cells and HT-29 cells treated with CXCL7-CM compared to the controls (p&lt;0.05). Glucose uptake was also greater in CXCL7-overexpressing cells and CXCL7-CM treated cells compared to vector controls. Data from Seahorse assay and Western blots confirmed that CXCL7-expressing HT-29 cells and HT-29 cells treated with CXCL7-CM have higher glycolytic capacity and increased glycolytic enzymes’ protein expression compared to the controls, respectively. In vivo, CXCL7-expressing HT-29 xenograft tumor weight and PCNA protein levels were 100% and 60% greater, respectively, than vector-expressing tumors (p&lt;0.05). Our study for the first time showed that CXCL7 stimulates colon cancer cell proliferation and enhances aerobic glycolysis.

Citation Format: Hadeel Aldhowayan, Jannatul F. Nipa, Lauren Jun, Ifeoluwa Odeniyi, Parker L. Jones, Ramesh Jeganathan, Elizabeth A. Lipke, Michael W. Greene. Role of CXCL7 in colon cancer cell proliferation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6967.