Baeri, A. | bioRxiv Science

bioRxiv2021

Blockade of pro-fibrotic response mediated by the miR-143/-145 cluster prevents targeted therapy-induced phenotypic plasticity and resistance in melanoma

Lineage dedifferentiation towards a mesenchymal-like state is a common mechanism of adaptive response and resistance to targeted therapy in melanoma. Yet, the transcriptional network driving this phenotypic plasticity remains elusive. Remarkably, this cellular state displays myofibroblast and fibrotic features and escapes MAPK inhibitors (MAPKi) through extracellular matrix (ECM) remodeling activities. Here we show that the anti-fibrotic drug Nintedanib/BIBF1120 is active to normalize the fibrous ECM network, enhance the efficacy of MAPK-targeted therapy and delay tumor relapse in a pre-clinical model of melanoma. We also uncovered the molecular networks that regulate the acquisition of this resistant phenotype and its reversion by Nintedanib, pointing the miR-143/-145 pro-fibrotic cluster as a driver of the therapy-resistant mesenchymal-like phenotype. Upregulation of the miR-143/-145 cluster under BRAFi/MAPKi therapy was observed in melanoma cells in vitro and in vivo and was associated with an invasive/undifferentiated profile of resistant cells. The 2 mature miRNAs generated from this cluster, miR-143-3p and miR-145-5p collaborated to mediate phenotypic transition towards a drug resistant undifferentiated mesenchymal-like state by targeting Fascin actin-bundling protein 1 (FSCN1), modulating the dynamic crosstalk between the actin cytoskeleton and the ECM through the regulation of focal adhesion dynamics as well as contributing to a fine-tuning of mechanotransduction pathways. Our study brings insights into a novel miRNA-mediated regulatory network that contributes to non-genetic adaptive drug resistance and provides proof-of-principle that preventing MAPKi-induced pro-fibrotic stromal response is a viable therapeutic opportunity for patients on targeted therapy.

cancer biology↗

bioRxiv2020

Identification of oncolytic vaccinia restriction factors in canine high-grade mammary tumor cells using single-cell transcriptomics

Mammary carcinoma, including triple-negative breast carcinomas (TNBC) are tumor-types for which human and canine pathologies are closely related at the molecular level. Low-passage, primary carcinoma cells from TNBC versus non-TNBC were used to compare the efficacy of an oncolytic vaccinia virus (VV). We show that non-TNBC cells are 28 times more sensitive to VV than TNBC cells in which VV replication is impaired. Single-cell RNA-seq performed on two different TNBC cell samples infected or not with VV highlighted three distinct populations: naive cells, bystander cells, defined as cells exposed to the virus but not infected and infected cells. The transcriptome of these three populations showed striking variations in the modulation of pathways regulated by cytokines and growth factors. We hypothesized that the pool of genes expressed in the bystander populations was enriched in antiviral genes. Bio-informatic analysis suggested that the reduced activity of the virus was associated with a higher mesenchymal status of the cells. In addition, we demonstrate experimentally that high expression of one gene, DDIT4, is detrimental to VV production. Considering that DDIT4 is associated with a poor prognosis in various cancers including TNBC, out data highlight DDIT4 as a candidate resistance marker for oncolytic poxvirus therapy. This information could be used to design new generations of oncolytic poxviruses. Beyond the field of gene therapy, this study demonstrate that single-cell transcriptomics can be used to identify cellular factors influencing viral replication. Author summaryThe identification of cellular genes influencing viral replication/propagation have been studied using hypothesis-driven approaches and/or high-throughput RNA interference screens. In the present report, we propose a methodology based on single-cell transcriptomic. We have studied, in the context of oncolytic virothepary, the susc eptibility of primary, low-passage mammary carcinoma cells of canine origin from different grades to an oncolytic vaccinia virus (VV). We highlight a fault in replication of VV in cells originated from high-grade triple-negative breast carcinomas (TNBC). Single-cell RNA-seq performed on TNBC cell samples infected with VV suggested that the reduced activity of the virus was associated with a higher mesenchymal status of the cells. We also demonstrate that high expression of one gene, DDIT4, is detrimental to VV production. Considering that DDIT4 is associated with a poor prognosis in various cancers including TNBC, out data highlight DDIT4 as a candidate resistance marker for oncolytic poxvirus therapy. Beyond the field of cancer gene therapy, we demonstrate here that single-cell transcriptomics increases the arsenal of tools available to identify cellular factors influencing viral replication.

cancer biology↗