Phosphoenolpyruvate carboxykinase-2 (PCK2) is a therapeutic target in triple-negative breast cancer

Metabolic rewiring in malignant transformation is often accompanied by altered expression of metabolic isozymes. Phosphoenolpyruvate carboxykinase-2 (PCK2) catalyzes the rate-limiting step of gluconeogenesis and is the dominant isoform in many cancers including triple-negative breast cancer (TNBC)....

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Published in:Breast cancer research and treatment 2024-08
Main Authors: Gunasekharan, Vignesh, Lin, Hao-Kuen, Marczyk, Michal, Rios-Hoyo, Alejandro, Campos, Gerson Espinoza, Shan, Naing Lin, Ahmed, Mostafa, Umlauf, Sheila, Gareiss, Peter, Raaisa, Raaisa, Williams, Richard, Cardone, Rebecca, Siebel, Stephan, Kibbey, Richard, Surovtseva, Yulia V, Pusztai, Lajos
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Language:English
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Summary:Metabolic rewiring in malignant transformation is often accompanied by altered expression of metabolic isozymes. Phosphoenolpyruvate carboxykinase-2 (PCK2) catalyzes the rate-limiting step of gluconeogenesis and is the dominant isoform in many cancers including triple-negative breast cancer (TNBC). Our goal was to identify small molecule inhibitors of PCK2 enzyme activity.PURPOSEMetabolic rewiring in malignant transformation is often accompanied by altered expression of metabolic isozymes. Phosphoenolpyruvate carboxykinase-2 (PCK2) catalyzes the rate-limiting step of gluconeogenesis and is the dominant isoform in many cancers including triple-negative breast cancer (TNBC). Our goal was to identify small molecule inhibitors of PCK2 enzyme activity.We assessed the impact of PCK2 down regulation with shRNA on TNBC cell growth in vitro and used AtomNet® deep convolutional neural network software to identify potential small molecule inhibitors of PCK2-based structure. We iteratively tested candidate compounds in an in vitro PCK-2 enzyme assay. The impact of the top hit on metabolic flux and cell viability was also assessed.METHODSWe assessed the impact of PCK2 down regulation with shRNA on TNBC cell growth in vitro and used AtomNet® deep convolutional neural network software to identify potential small molecule inhibitors of PCK2-based structure. We iteratively tested candidate compounds in an in vitro PCK-2 enzyme assay. The impact of the top hit on metabolic flux and cell viability was also assessed.PCK2 downregulation decreased growth of BT-549 and MDA-MB-231 cells and reduced metabolic flux through pyruvate carboxylase. The first AtomNet® in silico structural screen of 7 million compounds yielded 86 structures that were tested in PCK2 enzyme assay in vitro. The top hit (IC50 = 2.4 µM) was used to refine a second round of in silico screen that yielded 82 candidates to be tested in vitro, which resulted in 45 molecules with inhibition > 20%. In the second in vitro screen we also included 3-(3,4-dihydroxyphenyl)-2-hydroxypropanoate, previously suggested to be PCK2 inhibitor based on structure, which emerged as the top hit. The specificity of this compound was tested in PCK1 and PCK2 enzymatic assays and showed IC50 of 500 nM and 3.5-27 nM for PCK1 and PCK2, respectively.RESULTSPCK2 downregulation decreased growth of BT-549 and MDA-MB-231 cells and reduced metabolic flux through pyruvate carboxylase. The first AtomNet® in silico structural screen of 7 million c
ISSN:1573-7217
1573-7217
DOI:10.1007/s10549-024-07462-z