PURPOSE: Paclitaxel is an integral component of primary therapy for breast and epithelial ovarian cancers, but less than half of these cancers respond to the drug. Enhancing the response to primary therapy with paclitaxel could improve outcomes for women with both diseases.Experimental Design: Twelve kinases that regulate metabolism were depleted in multiple ovarian and breast cancer cell lines to determine whether they regulate sensitivity to paclitaxel in Sulforhodamine B assays. The effects of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 2 (PFKFB2) depletion on cell metabolomics, extracellular acidification rate, nicotinamide adenine dinucleotide phosphate, reactive oxygen species (ROS), and apoptosis were studied in multiple ovarian and breast cancer cell lines. Four breast and ovarian human xenografts and a breast cancer patient-derived xenograft (PDX) were used to examine the knockdown effect of PFKFB2 on tumor cell growth in vivo. RESULTS: Knockdown of PFKFB2 inhibited clonogenic growth and enhanced paclitaxel sensitivity in ovarian and breast cancer cell lines with wild-type TP53 (wtTP53). Silencing PFKFB2 significantly inhibited tumor growth and enhanced paclitaxel sensitivity in four xenografts derived from two ovarian and two breast cancer cell lines, and prolonged survival in a triple-negative breast cancer PDX. Transfection of siPFKFB2 increased the glycolysis rate, but decreased the flow of intermediates through the pentose-phosphate pathway in cancer cells with wtTP53, decreasing NADPH. ROS accumulated after PFKFB2 knockdown, which stimulated Jun N-terminal kinase and p53 phosphorylation, and induced apoptosis that depended upon upregulation of p21 and Puma. CONCLUSIONS: PFKFB2 is a novel target whose inhibition can enhance the effect of paclitaxel-based primary chemotherapy upon ovarian and breast cancers retaining wtTP53.