Triple-negative breast cancer (TNBC) does not express estrogen receptor (ER), progesterone receptor (PR) and Her2 as therapeutic targets and consequently has the worst prognosis among all types of breast cancers. Metabolic reprogramming toward aerobic glycolysis (also called the Warburg effect) is a hallmark of cancer, which is further activated in TNBC. Although aerobic glycolysis may be therapeutically targeted, the druggable pathway has not been identified. p38 is a TNBC oncogene and stimulates glucose uptake and metabolic adaption. This proposal will test the hypothesis that p38 promotes TNBC oncogenesis by stimulating PFKFB3/GLUT1-dependent aerobic glycolysis. This hypothesis is based on the following findings: 1) p38 stimulates glucose transporter 1 (GLUT1) expression and increases glucose uptake; 2) p38 promotes TNBC oncogenesis; 3) TNBCs are highly glycolytic with elevated PFKFB3 and GLUT1 expression; 4) p38 binds PFKFB3, a key glycolytic activator, in TNBC cells, whereas it interacts with much less glycolytic three family members (PFKFB1, 2 and 4) in non- TNBC cells; 5) MS/MS analysis identifies that p38 phosphorylates PFKFB3 at S467 leading to its stabilization, whereas data from public data-base shows that p38 gene expression is correlated with GLUT1 in invasive breast cancer tissues; 6) conditional p38 knockout (KO) inhibits tumorigenesis in a TNBC-like PyMT mouse genetic breast cancer model and decreases PFKFB3/GLUT1 expression; 7) p38 overexpression in TNBC cells increases PFKFB3/GLUT1 abundance, promotes their interaction, and stimulates ECAR (an indicator of glycolysis), indicating its activity of stimulating aerobic glycolysis by forming a ternary complex; and 8) pharmacological p38 and PFKFB3 inhibitors cooperatively decrease p-PFKFB3/PFKFB3/GLUT1 levels and inhibit TNBC growth in a manner dependent on p38. These results together indicate that p38 links aerobic glycolysis and TNBC oncogenesis through activating PFKFB3 and GLUT1. We will test this hypothesis by targeting the following aims: AIM 1 will investigate if p38 links aerobic glycolysis and TNBC oncogenesis through interaction with PFKFB3 and stimulating of PFKFB3 phosphorylation at S467; AIM 2 will determine if p38 cooperates with both PFKFB3 and GLUT1 to promote aerobic glycolysis and TNBC oncogenesis by stimulating a S467-dependent ternary-complex; and AIM 3 will determine if the p38-PFKFB3 kinase cascade is a therapeutic target for TNBC and if the p38/PFKFB3/Glut1 co-upregulation with elevated p-PFKFB3 identifies a subgroup of TNBC with a worse prognosis in clinic. These studies will demonstrate if the p38-PFKFB3 kinase cascade is a novel therapeutic target for TNBC and reveal if a combined application of p38/PFKFB3 pharmacological inhibitors is a potential effective therapeutic strategy that will impact TNBC clinical outcome and veteran health care.