Cancer-intrinsic Cxcl5 orchestrates global metabolic reprogramming of three dimensional cancers by upregulating Hif1a and Myc expression

The tumor microenvironment (TME) of pancreatic cancer is characterized by deposition of extracellular matrix (ECM). In 3D TME comprising cellular and non-cellular components, physical and chemical restriction induce oxygen and nutrients shortage in tumor site. Therefore, it is inevitable for cancer cells to manipulate the biological processes and metabolism for survival. However, it has not been fully explored the metablic phenotype coupling to the components existed in 3D and how the cancer cells exploit the metabolic reprogramming. In this study, we establisehd in vitro 3D tumor spheroid-in-Matrigel culture system recapitulating the 3D TME. We discovered the significant upregulation of Cxcl5 expression in murine pancreatic cancer cell line Panc02 by 3D culture with M1 macrophage conditioned media treatment. The loss of Cxcl5 in 3D Panc02 cells showed dramatical reduction of cancer growth. Importantly, the Cxcl5-deficient 3D cancer cells showed global alteration in metabolism. Additionally, Cxcl5 loss failed to control oxidative stress and in turn, increased ferroptotic cell death. Overexpression of Hif1a and Myc in Cxcl5-deficient cancer cells rescued the impaired metabolic processes, ferroptotic cell death, and cancer growth. Overall design: 1) To investigate the impact of macrophage soluble factors on Panc02 spheroid growth, we obtained unstimulated or stimulated macrophage conditioned media and treated to Panc02 spheroid. 6.5% FBS containing DMEM was treated to Panc02 spheroid as a control. 2) To investigate the role of Cxcl5 in 3D cancer growth, Cxcl5 was knockout using CRISPR/Cas9 system and cultured in 2D- or 3D-condition 3) To investigate the role of Hif1a and Myc, we overexpressed Hif1a and Myc in Cxcl5-deficient cells respectively.