Prostate Cancer (PCa) is the second leading diagnosed cancer and the fifth cause of cancer mortality in men worldwide. The combination of chemotherapy with phytochemicals have been widely studied in different types of cancer including PCa. Andrographolide, a labdane diterpenoid that is the main bioactive component of the medicinal plant Andrographis paniculata, have shown a wide range of biological activities including anticarcinogenic properties. Previous studies from our laboratory has shown that Andrographolide induces a DNA damage response involving cell grown suppression, cell cycle arrest and apoptosis in PCa in vitro. In this study we aim to evaluate the role of Andrographolide in carbohydrate metabolism and mitochondrial function in PCa. Tumors from a mouse xenograft model in which the anterior prostate lobes of SCID mice were injected with 22RV1 cells and treated with Andrographolide during 4 weeks, were used for microarray analysis. The global gene expression profile was identified and analyzed using the Affymetrix GeneChip® Human Gene 2.0 array. Ingenuity Pathway Analysis (IPA) revealed alterations in carbohydrate metabolism and predicted the upregulation of mitochondrial associated genes DAP3 and HIFa. The microarray results were confirmed by real-time quantitative PCR (RT-qPCR) for some representative genes. Mitochondrial function was assessed using the Seahorse Mito Stress assay in androgen-dependent (22RV1) and androgen-independent (PC3) cell lines treated with Andrographolide (25µM). RT-qPCR showed that ZNF676, ZNF766, TXNL4 were up-regulated in PC3 PCa cells treated with Andrographolide, and ZNF676, TXNL4B, SASS6 and FBXL4 in 22Rv1 PCa cells treated with Andrographolide. The Seahorse Mito Stress assay showed a reduction of ATP Production, Basal Respiration and Spare Respiratory Capacity in 22RV1 PCa cells when treated with Andrographolide 25mM at 24hrs and 48hrs. ATP production was reduced in PC3 PCa cells treated with 25mM Andrographolide for 24hrs and 48hrs. Our data confirmed that Andrographolide alters the regulation of genes associated with carbohydrate metabolism, and the mitochondrial function. Our ongoing studies are evaluating the effect of Andrographolide on mtDNA damage and mtDNA expression in PCa cells. This data suggests a novel mechanism in which Andrographolide up-regulates nuclear carbohydrate metabolism's genes and suppress the mitochondrial activity in PCa. Citation Format: Jesus M. Sosa Rivera, Andres Lopez Rivas, Yarelis Roque Reyes, Desiree Arroyo Villegas, Maria Sanchez Velazquez, Humberto Ortiz Zuazaga, Carlos Torres Ramos, Magaly Martinez Ferrer. Andrographolide induces metabolic changes and suppresses mitochondrial activity in prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4784.