Rosario, C. J. | Search bioRxiv

bioRxiv2025

Overcoming Immune Checkpoint Inhibitor Resistance via Potent and Selective Dual αvβ6/8 Inhibitors Based on Engineered Lasso Peptides

Integrins v{beta}6 and v{beta}8 in the tumor microenvironment (TME) have been shown to activate immunosuppressive TGF-{beta}, which serves as an important mechanism for immune checkpoint inhibitor resistance in a range of tumors. In this study, we demonstrate the utility of lasso peptides as versatile scaffolds for designing new therapeutics. A series of highly potent and selective dual v{beta}6/8 inhibitors were engineered through a combination of epitope scanning, computational design, and directed evolution. Several analogs, such as lassotides 36 and 47, were fully characterized and physicochemical, in vitro pharmacological, and in vivo data are reported. Lassotide 47, a half-life extended derivative of 36, was shown to strongly sensitize anti-mPD-1-resistant tumors in mice when dosed in combination with the checkpoint inhibitor. The 47/anti-mPD-1 combination was shown to halt tumor growth and regress tumors in mouse models of triple negative breast and ovarian cancers. Dual inhibition of v{beta}6/8 integrins expressed in the TME thus represents a promising tumor-specific strategy to overcome TGF-{beta}-driven resistance and enhance the anti-tumor efficacy of immune checkpoint inhibitors.

cancer biology↗

bioRxiv2022

Regulation of Chlamydia RsbU phosphatase activity by the enolase product, phosphoenolpyruvate

The intracellular pathogen Chlamydia temporally regulates the expression of its genes but the upstream signals that control transcription are not known. The best studied regulatory pathway is a partner switching mechanism that involves an anti-sigma factor RsbW, which inhibits transcription by binding and sequestering the sigma subunit of RNA polymerase. RsbW is itself regulated by an anti-anti-sigma factor RsbV whose phosyphorylation state is controlled by the phosphatase RsbU. In this study, we showed that Chlamydia trachomatis RsbU requires manganese or magnesium as a cofactor and dephosphorylates RsbV1 and RsbV2, which are the two chlamydial paralogs of RsbV. The gene for RsbU is adjacent to the enolase gene in a number of Chlamydia genomes, and we showed that eno and rsbU are co-transcribed from the same operon. In other bacteria, there is no known functional connection between the Rsb pathway and enolase, which is an enzyme in the glycolytic pathway. We found, however, that Chlamydia RsbU phosphatase activity was inhibited by phosphoenolpyruvate (PEP), the product of the enolase reaction, but not by 2-phosphoglycerate (2PGA), which is the substrate. These findings suggest that the enolase reaction, and more generally glucose metabolism, may provide an upstream signal that regulates transcription in Chlamydia through the RsbW pathway. IMPORTANCEThe RsbW pathway is a phosphorelay that regulates gene expression in Chlamydia but its upsteam signal has not been identified. We showed that RsbU, a phosphatase in this pathway is inhibited by phosphoenolpyruvate, which is the product of the enolase reaction. As enolase is an enzyme in the glycolytic pathway, these results reveal an unrecognized link between glucose metabolism and gene regulation in chlamydiae. Moreover, as these intracellular bacteria acquire gluose from the infected host cell, our findings suggest that glucose availability may be an external signal that controls chlamydial gene expression.

microbiology↗