Munuganti, R. | Search bioRxiv

bioRxiv2022

Discovery and characterization of a first-in-field transcription factor BRN2 inhibitor for the treatment of neuroendocrine prostate cancer.

The increased incidence of treatment-emergent neuroendocrine prostate cancer (NEPC) is particularly alarming as this diagnosis is associated with poor prognosis. Despite initial responses to platinum-based chemotherapy, relapses are common and there is no effective second line therapy for NEPC. We previously identified that neuronal transcription factor BRN2 (POU3F2) is a potent driver of neuroendocrine differentiation and an attractive target for NEPC. Utilizing a combination of in silico modeling and X-ray crystallography followed by structure-based lead optimization, we have developed the first potent, specific and orally bioavailable BRN2 inhibitor (B18-94), which inhibits the interaction between BRN2 and DNA. This loss of BRN2 on the chromatin drastically reduces its transcriptional output resulting in downregulation of several known targets in NEPC such as SOX2, ASCL1 and PEG10. Additionally, B18-94 reduces specifically cell proliferation specifically in multiple NEPC models with no effect on adenocarcinoma and other BRN2 negative prostate cancer models. Importantly, the consistency in the transcriptomic changes driven by B18-94 and or CRISPR/Cas9 mediated BRN2 knockout confirmed the on-target specificity, with both methods of BRN2 inhibition downregulating pathways involved in cellular plasticity and proliferation. Finally, we have demonstrated that B18-94, the first-in-field POU-domain transcription factor inhibitor, significantly reduced tumor growth in several NEPC xenograft models with no observable toxicity, suggesting potential for therapeutic intervention of NEPC.

cancer biology↗

bioRxiv2021

A novel small molecule that induces cytotoxicity in lung cancer cells inhibits disulfide reductases GSR and TXNRD1

High-throughput phenotype-based screening of large libraries of novel compounds without known targets can identify small molecules that elicit a desired cellular response, but additional approaches are required to find and characterize their targets and mechanisms of action. Here we show that a compound termed lung cancer screen 3 (LCS3), previously selected for its ability to impair the growth of human lung adenocarcinoma (LUAD) cell lines, but not normal lung cells, induces oxidative stress and activates the NRF2 signaling pathway by generating reactive oxygen species (ROS) in sensitive LUAD cell lines. To identify the target that mediates this effect, we applied thermal proteome profiling (TPP) and uncovered the disulfide reductases GSR and TXNRD1 as LCS3 targets. Through enzymatic assays using purified protein, we confirmed that LCS3 inhibits disulfide reductase activity through a reversible, uncompetitive mechanism. Further, we demonstrate that LCS3-sensitive LUAD cells are correspondingly sensitive to the synergistic inhibition of glutathione and thioredoxin pathways. Lastly, a genome-wide CRISPR knockout screen identified the loss of NQO1 as a mechanism of LCS3 resistance. This work highlights the ability of TPP to uncover targets of small molecules identified by high-throughput screens and demonstrates the potential utility of inhibiting disulfide reductases as a therapeutic strategy for LUAD.

cancer biology↗