Biology subjects

Fraile-Bethencourt, E.

Publications and source records attributed to Fraile-Bethencourt, E..

3 recordsLinked to original sources

bioRxiv2022

Tumor cell intrinsic RIG-I activation is sufficient to drive immune mediated tumor rejection

Targeting cytosolic nucleic acid sensors is a potent approach to drive type I interferon responses and anti-tumor immunity. Recent evidence suggests that activation of retinoic acid inducible gene-I (RIG-I) using synthetic hairpin RNA agonists decreases tumor progression in multiple preclinical models. However, the role of tumor cell intrinsic RIG-I in shaping tumor cell fates and the host immune microenvironment remains unclear. Here, we show that RIG-I expression is correlated with better overall survival and a distinct immune gene signature in specific human cancers including colorectal cancer. Activation of RIG-I in breast and colorectal cancer cells is sufficient to drive tumor cell death in vitro and significantly delay tumor growth in vivo in multiple preclinical models. Importantly, the efficacy of tumor cell RIG-I activation is lost in immune deficient mice suggesting the requirement of immune responses for this effect. We observe that tumor cell intrinsic RIG-I activation elicits a robust cellular and molecular immune response. We show that tumor cell RIG-I activation also leads to induction of specific immune checkpoints including PD-L1. Using a publicly available database, we found that RIG-I expression serves as an excellent prognostic marker for responders to checkpoint immunotherapy, particularly PD-L1/PD-1 across cancers. Finally, combination of tumor cell intrinsic RIG-I activation with anti-PD-L1 led to a synergistic decrease in tumor growth in a colorectal tumor model. Our findings suggest that tumor cell intrinsic RIG-I can be targeted to enhance anti-tumor immune responses and highlights a potential strategy for anti-cancer vaccines that can invigorate the immune system.

cancer biology↗

bioRxiv2022

DNA damage-induced lncRNA MEG9 impacts angiogenesis

Endothelial cells are highly responsive to environmental changes that allow them to adapt to intrinsic and extrinsic stimuli and switch their transcriptome accordingly to go back to vascular homeostasis. Our previous data demonstrated that small non-coding-RNAs respond quickly to genotoxic stressors and determined endothelial cell fate and DNA damage response. To further understand the contribution of non-coding-RNAs, we profiled differentially expressed long non-coding RNAs in response to genotoxic stress and compared them to pro-angiogenic growth factor signaling. We identified the Maternally expressed gene 9 (MEG9) as a cytoprotective lncRNA in the endothelium. Gain and Loss-of-function studies indicate that MEG9 prevents endothelial cells from cell death, suggesting that MEG9 responses to genotoxic stress can be an adaptive and protective mechanism. Consistent with this phenotype, the knockdown of MEG9 decreases growth factor-dependent angiogenesis in a 3D fibrin gel angiogenesis assay. Deletion of the MEG9 ortholog, Mirg, in mice results in increased vascular leak in Matrigel plugs and a sex and age-dependent decrease in platelets. Mechanistically, we observed that both MEG9 knockdown in vitro and Mirg-deleted mice in vivo activated common pathways, including apoptosis, clotting, and inflammation. Indeed, the proinflammatory adhesion molecule ICAM1 was significantly increased in human and mouse endothelial cells in a MEG9-dependent manner, supporting the increased vascular permeability observed on MEG9 deficient cells. Taken together, our findings illustrate how genotoxic stress responses through dynamic modulation of lncRNAs, such as MEG9, trigger adaptive mechanisms to maintain endothelial function, while loss of these molecules contributes to maladaptive responses and endothelial cell dysfunction.

cell biology↗

bioRxiv2021

A cell-based screen identifies HDAC inhibitors and PLK inhibitor as activators of RIG-I signaling

Enhancing the immune microenvironment in cancer by targeting the nucleic acid sensors is becoming a potent therapeutic strategy. Among the nucleic acid sensors, activation of the RNA sensor Retinoic Acid-inducible Gene (RIG-I) using small hairpin RNAs has been shown to elicit powerful innate and adaptive immune responses. Given the challenges inherent in pharmacokinetics and delivery of RNA based agonists, we set out to discover small molecule agonists of RIG-I using a cell-based assay. To this end, we established and validated a robust high throughput screening assay based on a commercially available HEK293 reporter cell line with a luciferase reporter downstream of tandem interferon stimulated gene 54 (ISG54) promoter elements. We first confirmed that the luminescence in this cell line is dependent on RIG-I and the interferon receptor using a hairpin RNA RIG-I agonist. We established a 96-well and a 384-well format HTS based on this cell line and performed a proof-of-concept screen using an FDA approved drug library of 1200 compounds. Surprisingly, we found two HDAC inhibitors Entinostat, Mocetinostat and the PLK1 inhibitor Volasertib significantly enhanced ISG-luciferase activity. This luminescence was substantially diminished in the null reporter cell line indicating the increase in signaling was dependent on RIG-I expression. Combination treatment of tumor cell lines with Entinostat increased RIG-I induced cell death in a mammary carcinoma cell line that is resistant to either Entinostat or RIG-I agonist alone. Taken together, our data indicates an unexpected role for HDAC1,-3 inhibitors in enhancing RIG-I signaling and highlight potential opportunities for therapeutic combinations.

cancer biology↗