Biology subjects

Byrne, K. T.

Publications and source records attributed to Byrne, K. T..

3 recordsLinked to original sources

bioRxiv2025

Pancreatic cancer-intrinsic HuR regulates the pro-tumorigenic properties of extracellular vesicles

Pancreatic ductal adenocarcinoma (PDAC) tumors contain chaotic vasculature that limits immune surveillance and promotes early events in the metastatic cascade. However, current antiangiogenic therapies have failed in PDAC, and thus, it remains important to uncover mechanisms by which cancer cells signal to endothelial cells to increase angiogenesis. Our lab has shown that the tumor-intrinsic RNA-binding protein HuR (ELAVL1) plays an important role re-shaping the tumor microenvironment (TME) by regulating the stability and translation of cytokine encoding transcripts. Herein, we demonstrate that PDAC-intrinsic HuR influences endothelial cell function in the TME via extracellular vesicle (EV) signaling, an underexplored signaling axis in tumor progression. We found that HuR knockout (KO) tumors have impaired growth in an immunocompetent mouse model, and that administering purified wildtype (WT) EVs can increase tumor growth. Further, we observed that PDAC EVs contain HuR-dependent mRNA and protein cargoes relating to endothelial cell function and angiogenesis. Treatment of endothelial cells with HuR WT EVs strongly increased the expression of genes involved in barrier function and endothelial cell development, and directly increased their migratory and tube forming functions. In an immunocompetent orthotopic mouse model of PDAC, we showed that HuR increases endothelial cell presence and sprouting, while decreasing ICAM-1 expression. Importantly, we found utilizing a genetic EV reporter, that decreased ICAM-1 within WT tumors occurs in endothelial cells that have imported PDAC EVs, suggesting that this signaling axis is directly modulating endothelial cell behavior in vivo. Collectively, our data reveal a new role of HuR in EV signaling to endothelial cells, promoting angiogenesis while restricting endothelial cell leukocyte trafficking behavior.

cancer biology↗

bioRxiv2022

Agonistic anti-CD40 converts Tregs into Type 1 effectors within the tumor micro-environment

Multiple cell types, molecules, and processes contribute to inhibition of anti-tumor effector responses, often frustrating effective immunotherapy. Among these, Foxp3+ CD4+ cells (Tregs) are well-recognized to play an immunosuppressive role in the tumor microenvironment. The first clinically successful checkpoint inhibitor, anti-CTLA-4 antibody, may deplete Tregs at least in part by antibody-dependent cellular cytotoxicity (ADCC), but this effect is unreliable in mice, including in a genetically engineered mouse model of pancreatic ductal adenocarcinoma (PDAC). In contrast, agonistic CD40 antibody, which serves as an effective therapy, is associated with notable Treg disappearance in the PDAC model. The mechanism of CD40-mediated Treg loss is poorly understood, as Tregs are CD40-negative. Here we have explored the mechanistic basis for the loss of Foxp3 T cells upon anti-CD40 treatment and find, using tissue-level multiplex immunostaining and orthogonal dissociated cell analyses, that Tregs are not depleted but converted into interferon-{gamma} (IFN-{gamma}) producing, Type I CD4+ T effector cells. This process depends on IL-12 and IFN-{gamma} signaling evoked by action of the anti-CD40 antibody on dendritic cells (DCs), especially BATF3-dependent cDC1s. These findings provide insight into a previously unappreciated mechanism of CD40 agonism as a potent anti-tumor intervention that promotes the re-programming of Tregs into tumor-reactive CD4+ effector T cells, markedly augmenting the anti-tumor response.

immunology↗

bioRxiv2022

An open protocol for modeling T Cell Clonotype repertoires using TCRβ CDR3 sequences

T cell receptor (TCR) repertoires can be profiled using next generation sequencing (NGS) to monitor dynamical changes in response to disease and other perturbations. Several strategies for profiling TCRs have been recently developed with different benefits and drawbacks. Genomic DNA-based bulk sequencing, however, remains the most cost-effective method to profile TCRs. The major disadvantage of this method is the need for multiplex target amplification with a large set of primer pairs with potentially very different amplification efficiencies. One approach addressing this problem is by iteratively adjusting the concentrations of the primers based on their efficiencies, and then computationally correcting any remaining bias. Yet there are no standard, publicly available protocols to process and analyze raw sequencing data generated by this method. Here, we utilize an equimolar primer mixture and propose a single statistical normalization step that efficiently corrects for amplification bias post sequencing. Using samples analyzed by both approaches, we show that the concordance between bulk clonality metrics obtained from using the commercial kits and that developed herein is high. Therefore, we suggest the method presented here as an inexpensive and non-commercial alternative for measuring and monitoring adaptive dynamics in TCR clonotype repertoire.

bioinformatics↗