Decursinol Angelate Inhibits Glutamate Dehydrogenase 1 Activity and Induces Intrinsic Apoptosis in MDR-CRC Cells

SIMPLE SUMMARY: Upregulated glutaminolysis is a typical hallmark of malignant tumors across different cancers. In our previous published paper, we proved the ability of decursinol angelate (DA) to bind to glutamate dehydrogenase 1 (GDH1) in the abortive complex of the ADP activation site and inhibit the activity of GDH. We also elucidated the significance of upregulated GDH in colorectal cancer (CRC) patients with a comparison of different subtypes of CRC and the survival probability for CRC across different races. As per the data collected by the Clinical Proteomic Tumor Analysis Consortium (CPTAC), we found that African Americans were highly susceptible to early death for CRC patients that had upregulated GDH. As specified in reports, glutamate dehydrogenase is a novel prognostic marker for predicting metastasis in colorectal cancer patients. The expression of glutaminolysis is highly upregulated in many different types of cancers such as lymph, cervix, lungs, breast and brain. In our present study, we evaluated the role of DA in inducing apoptotic cell death in multidrug-resistant colorectal cancer cells. Inhibiting glutaminolysis can be a good therapeutic approach for developing anti-cancer drugs. ABSTRACT: Colorectal cancer (CRC) was the second most commonly diagnosed cancer worldwide and the second most common cause of cancer-related deaths in Europe in 2020. After CRC patients’ recovery, in many cases a patient’s tumor returns and develops chemoresistance, which has remained a major challenge worldwide. We previously published our novel findings on the role of DA in inhibiting the activity of GDH1 using in silico and enzymatic assays. No studies have been conducted so far to explain the inhibitory role of DA against glutamate dehydrogenase in MDR-CRC cells. We developed a multidrug-resistant colorectal cancer cell line, HCT-116(MDR), after treatment with cisplatin and 5-fluorouracil. We confirmed the MDR phenotype by evaluating the expression of MDR1, ABCB5, extracellular vesicles, polyploidy, DNA damage response markers and GDH1 in comparison with parental HCT-116(WT) (HCT-116 wild type). Following confirmation, we determined the IC(50) and performed clonogenic assay for the efficacy of decursinol angelate (DA) against HCT-116(MDR) (HCT-116 multidrug resistant). Subsequently, we evaluated the novel interactions of DA with GDH1 and the expression of important markers regulating redox homeostasis and cell death. DA treatment markedly downregulated the expression of GDH1 at 50 and 75 μM after 36 h, which directly correlated with reduced expression of the Krebs cycle metabolites α-ketoglutarate and fumarate. We also observed a systematic dose-dependent downregulation of MDR1, ABCB5, TERT, ERCC1 and γH2AX. Similarly, the expression of important antioxidant markers was also downregulated. The markers for intrinsic apoptosis were notably upregulated in a dose-dependent manner. The results were further validated by flow cytometry and TUNEL assay. Additionally, GDH1 knockdown on both HCT-116(WT) and HCT-116(MDR) corresponded to a decreased expression of γH2AX, catalase, SOD1 and Gpx-1, and an eventual increase in apoptosis markers. In conclusion, inhibition of GDH1 increased ROS production, decreased cell proliferation and increased cell death.