Eccleston, M. | bioRxiv Science

bioRxiv2024

EHZ2 inhibition enhances the activity of platinum chemotherapy in aggressive variant prostate cancer

BackgroundEZH2 promotes aggressive-variant prostate cancer (AVPC) progression via histone H3-Lysine-27 tri-methylation (H3K27me3). We hypothesize that epigenetic reprogramming via EZH2 inhibitors (EZH2i) improves the efficacy of chemotherapy in AVPC. MethodsWe studied the expression of EZH2 in clinical prostate cancer cohorts (bioinformatics). We determined the effect of EZH2i on both cellular- and cell-free-H3K27me3 levels. We measured effects of carboplatin with/without EZH2i on AVPC cell viability (IC50). We studied how EZH2i modulate gene expression (RNA Seq). ResultsEZH2 was significantly up-regulated in AVPC vs other prostate cancer types. EZH2i reduced both cellular and cell free-H3K27me3 levels. EZH2i significantly reduced carboplatin IC50. EZH2i reduced the expression of DNA repair and increased the expression of pro-apoptotic genes. Article HighlightsO_LIPolycomb-mediated gene silencing promotes prostate cancer progression C_LIO_LIAggressive-variant prostate cancers (AVPCs) are characterized by increased activity of the Polycomb-Repressive Complex 2 (PRC2) C_LIO_LIHere we show that PRC2 inhibitors are scarcely effective as monotherapy in ACPC cells C_LIO_LIHowever the combination of PRC2 inhibitors and carboplatin is highly synergistic C_LIO_LIRNA Seq studies revealed that PRC2 inhibitors enhance carboplatin activity by modulating several key pathways, including DNA repair and apoptosis. C_LI

pharmacology and toxicology↗

bioRxiv2023

Novel rapid high-throughput method of NETosis Induction and Inhibition with physiological triggers and inhibitors

Neutrophils, the most abundant white blood cells in humans, play pivotal roles in innate immunity, rapidly migrating to sites of infection and inflammation to phagocytose, neutralize, and eliminate invading pathogens. Neutrophil Extracellular Trap (NET) formation in response to pathogens is increasingly recognized as an essential rapid innate immune response, but when dysregulated contributes to pathogenesis of sepsis and immunothrombotic disease. Current models of NETosis are limited, routinely employing non-physiological triggers that can bypass natural NET regulatory pathways. Models utilizing isolated neutrophils and immortalized cell lines, do not reflect the complex biology underlying neutrophil activation and NETosis, that occurs in whole-blood. Here we describe a novel, high-throughput ex-vivo whole blood induced NETosis model using combinatorial pooling of native NETosis inducing factors in a more biologically relevant Synthetic-Sepsis model. We found different combinations of factors evoked distinct neutrophil responses in the rate of NET generation and/or magnitude of NETosis. Despite inter-donor variability, similar sets of pro-inflammatory molecules induced consistent responses across donors. We found at least three biological triggers were necessary to induce NETosis in our system including either TNF- or LT-. To our knowledge, we report the first human ex-vivo model utilizing naturally occurring molecules to induce NETosis in whole blood. This approach could be used for drug screening and, importantly, inadvertent activators of NETosis. These findings emphasize the importance of investigating neutrophil physiology in a biologically relevant context to enable a better understanding of disease pathology, risk factors, and therapeutic targets, potentially, providing novel strategies for disease intervention and treatment. EssentialsO_LINETosis is a vital immune response, but dysregulation leads to disastrous health outcomes. C_LIO_LICurrent NETosis models dont reflect the complex endogenous signaling that occurs in whole blood C_LIO_LINetosis induction stimuli differs between isolated neutrophils and whole blood. C_LIO_LIA minimum of three physiological factors are required to induce NETosis in whole blood. C_LI

immunology↗