SUMMARY It remains unclear how cancer cells coordinate glycolysis and biosynthesis to support rapidly growing tumors. We found that glycolytic enzyme phosphoglycerate mutase 1 (PGAM1), commonly upregulated in human cancers due to loss of TP53, contributes to biosynthesis regulation in part by controlling intracellular levels of its substrate 3-phosphoglycerate (3-PG) and product 2-phosphoglycerate (2-PG). 3-PG binds to and inhibits 6-phosphogluconate dehydrogenase in the oxidative pentose phosphate pathway (PPP), while 2-PG activates 3-phosphoglycerate dehydrogenase to provide feedback control of 3-PG levels. Inhibition of PGAM1 by shRNA or a small molecule inhibitor PGMI-004A results in increased 3-PG and decreased 2-PG levels in cancer cells, leading to significantly decreased glycolysis, PPP flux and biosynthesis, as well as attenuated cell proliferation and tumor growth.

SUMMARY How mitochondrial glutaminolysis contributes to redox homeostasis in cancer cells remains unclear. Here we report that the mitochondrial enzyme glutamate dehydrogenase 1 (GDH1) is commonly upregulated in human cancers. GDH1 is important for redox homeostasis in cancer cells by controlling the intracellular levels of its product alpha-ketoglutarate (α-KG) and subsequent metabolite fumarate. Mechanistically, fumarate binds to and activates a ROS scavenging enzyme glutathione peroxidase 1 (GPx1). Targeting GDH1 by shRNA or a small molecule inhibitor R162 resulted in imbalanced redox homeostasis, leading to attenuated cancer cell proliferation and tumor growth.

SUMMARY Mitochondrial pyruvate dehydrogenase complex (PDC) is crucial for glucose homoeostasis in mammalian cells. The current understanding of PDC regulation involves inhibitory serine phosphorylation of pyruvate dehydrogenase (PDH) by PDH kinase (PDK), whereas dephosphorylation of PDH by PDH phosphatase (PDP) activates PDC. Here we report that lysine acetylation of PDHA1 and PDP1 is common in EGF-stimulated cells and diverse human cancer cells. K321 acetylation inhibits PDHA1 by recruiting PDK1 and K202 acetylation inhibits PDP1 by dissociating its substrate PDHA1, both of which are important to promote glycolysis in cancer cells and consequent tumor growth. Moreover, we identified mitochondrial ACAT1 and SIRT3 as the upstream acetyltransferase and deacetylase, respectively, of PDHA1 and PDP1, while knockdown of ACAT1 attenuates tumor growth. Furthermore, Y381 phosphorylation of PDP1 dissociates SIRT3 and recruits ACAT1 to PDC. Together, hierarchical, distinct post-translational modifications act in concert to control molecular composition of PDC and contribute to the Warburg effect.

The oxidative pentose phosphate pathway (PPP) contributes to tumor growth, but the precise contribution of 6-phosphogluconate dehydrogenase (6PGD), the third enzyme in this pathway, to tumorigenesis remains unclear. We found that suppression of 6PGD decreased lipogenesis and RNA biosynthesis and elevated ROS levels in cancer cells, attenuating cell proliferation and tumor growth. 6PGD-mediated production of ribulose-5-phosphate (Ru-5-P) inhibits AMPK activation by disrupting the active LKB1 complex, thereby activating acetyl-CoA carboxylase 1 and lipogenesis. Ru-5-P and NADPH are thought to be precursors in RNA biosynthesis and lipogenesis, respectively; thus, our findings provide an additional link between oxidative PPP and lipogenesis through Ru-5-P-dependent inhibition of LKB1-AMPK signaling. Moreover, we identified and developed 6PGD inhibitors, Physcion and its derivative S3, that effectively inhibited 6PGD, cancer cell proliferation and tumor growth in nude mice xenografts without obvious toxicity, suggesting that 6PGD could be an anticancer target.

Because the p300/CBP-mediated hyperacetylation of RelA (p65) is critical for nuclear factor-KB (NF-KB) activation, the attenuation of p65 acetylation is a potential molecular target for the prevention of chronic inflammation. During our ongoing screening study to identify natural compounds with histone acetyltransferase inhibitor (HATi) activity, we identified epigallocatechin-3-gallate (EGCG) as a novel HATi with global specificity for the majority of HAT enzymes but with no activity toward epigenetic enzymes including HDAC, SIRT1, and HMTase. At a dose of 100 Mmol/L, EGCG abrogates p300-induced p65 acetylation in vitro and in vivo, increases the level of cytosolic IKBA, and suppresses tumor necrosis factor A (TNFA)-induced NF-KB activation. We also showed that EGCG prevents TNFA-induced p65 translocation to the nucleus, confirming that hyperacetylation is critical for NF-KB translocation as well as activity. Furthermore, EGCG treatment inhibited the acetylation of p65 and the expression of NF-KB target genes in response to diverse stimuli. Finally, EGCG reduced the binding of p300 to the promoter region of interleukin-6 gene with an increased recruitment of HDAC3, which highlights the importance of the balance between HATs and histone deacetylases in the NF-KB-mediated inflammatory signaling pathway. Importantly, EGCG at 50 Mmol/L dose completely blocks EBV infection-induced cytokine expression and subsequently the EBV-induced B lymphocyte transformation. These results show the crucial role of acetylation in the development of inflammatory-related diseases. [Cancer Res 2009;69(2):583-92]