Aspulvinone O, a natural inhibitor of GOT1 suppresses pancreatic ductal adenocarcinoma cells growth by interfering glutamine metabolism

Abstract

Background Distinctive from their normal counterparts, cancer cells exhibit unique metabolic dependencies on glutamine to fuel anabolic processes. Specifically, pancreatic ductal adenocarcinoma (PDAC) cells rely on an unconventional metabolic pathway catalyzed by aspartate transaminase 1 (GOT1) to rewire glutamine metabolism and support nicotinamide adenine dinucleotide phosphate (NADPH) production. Thus, the important role of GOT1 in energy metabolism and Reactive Oxygen Species (ROS) balance demonstrates that targeting GOT1 may serve as an important therapeutic target in PDAC. Methods To assay the binding affinity between Aspulvinone O (AO) and GOT1 proteins, the virtual docking, microscale thermophoresis (MST), cellular thermal shift assay (CETSA) and drug affinity responsive target stability (DARTS) methods were employed. GOT1 was silenced in several PDAC cell lines. The level of OCR and ECR were assayed by seahorse. To evaluate the in vivo anti-tumor efficacy of AO, the xenograft model was built in CB17/scid mouse. Results Screening of an in-house natural compound library identified the AO as a novel inhibitor of GOT1 and repressed glutamine metabolism, which sensitizes PDAC cells to oxidative stress and suppresses cell proliferation. Virtual docking analysis suggested that AO could bind to the active site of GOT1 and form obvious hydrophobic interaction with Trp141 together with hydrogen bonds with Thr110 and Ser256. Further in vitro validation, including MST, CETSA and DARTS, further demonstrated the specific combining capacity of AO. We also show that the selective inhibition of GOT1 by AO significantly reduces proliferation of PDAC in vitro and in vivo. Conclusions Taken together, our findings identify AO as a potent bioactive inhibitor of GOT1 and a novel anti-tumour agent for PDAC therapy. Electronic supplementary material The online version of this article (10.1186/s12964-019-0425-4) contains supplementary material, which is available to authorized users.

GOT1 Inhibition Primes Pancreatic Cancer for Ferroptosis through the Autophagic Release of Labile Iron

“…To test the specificity of GOT1 against PDA, we extended our cell panel to non-transformed human lines. We found human pancreatic stellate cells (hPSC), human lung fibroblasts (IMR-90), and human non-transformed pancreatic exocrine cells (hPNE) were minimally affected upon GOT1 knockdown, in agreement with previous results, suggesting that this pathway may be dispensable in non-transformed cells (Figures 1E and S1G) 8,12 . Together, these data demonstrate many PDA cell lines require GOT1 for growth while non-transformed cell lines do not, highlighting a potential therapeutic window.…”

“…This could also account for how GOT1 inhibition sensitizes PDA to ferroptosis (Figures 3-5) . These results would be consistent with previous studies demonstrating GOT1 inhibition can induce ROS 8,12 and radiosensitize PDA both in vitro and in vivo 18 . Future studies will be required to examine the role of FSP1 and potential interactions with the GOT1 pathway in PDA.…”

“…This points to GOT1 inhibition as a logical combination therapy to enhance therapeutic efficacy of BSO. To this end, we and others engaged in drug discovery campaigns to develop small inhibitors of GOT1 10,11,12,13 . Among these, we found that the KATII inhibitor PF-04859989 also has potent GOT1 inhibitory activity 13 , and we apply that herein as a tool compound.…”

“…Thus, in an effort to target this rewired metabolic pathway, we have placed our focus on GOT1. And, notably, we and others recently identified drug scaffolds that may serve as leads in the development of a clinical grade GOT1 inhibitor 10,11,12,13 .…”

Cysteine Aminotransferase (CAT): A Pivotal Sponsor in Metabolic Remodeling and an Ally of 3-Mercaptopyruvate Sulfurtransferase (MST) in Cancer

“…Several studies reported inhibitors towards CAT, particularly the cCAT form, which led to anti-tumoral effects. In pancreatic cancer, Yoshida and colleagues tested an inhibitory compound (PF-04859989) that covalently bonded to cCAT, promoting inhibitory effects in a time- and PLP-dependent fashion, further showing selective anti-proliferative effects towards pancreas cancer cell lines [ 163 ]. Sun et al reported the tumor suppressive effects of Aspulvinone O (AO), a natural compound isolated from Aspergillus terreus and found it to be a selective inhibitor for cCAT in PDAC cells but not healthy cells, both in vitro and in vivo.…”