Biology subjects

Washam, C. L.

Publications and source records attributed to Washam, C. L..

4 recordsLinked to original sources

bioRxiv2024

HIF-alpha signaling mediates the macrophage inflammatory response during Leishmania major infection

Cutaneous leishmaniasis (CL) contributes significantly to the global burden of neglected tropical diseases, with 12 million people currently infected with Leishmania parasites. CL encompasses a range of disease manifestations, from self-healing skin lesions to permanent disfigurations. Currently there is no vaccine available, and many patients are refractory to treatment, emphasizing the need for new therapeutic targets. Previous work demonstrated macrophage HIF--mediated lymphangiogenesis is necessary to achieve efficient wound resolution during murine L. major infection. Here, we investigate the role of macrophage HIF- signaling independent of lymphangiogenesis. We sought to determine the relative contributions of the parasite and the host-mediated inflammation in the lesional microenvironment to myeloid HIF- signaling. Because HIF- activation can be detected in infected and bystander macrophages in leishmanial lesions, we hypothesize it is the hosts inflammatory response and microenvironment, rather than the parasite, that triggers HIF- activation. To address this, macrophages from mice with intact HIF- signaling (LysMCreARNTf/+) or mice with deleted HIF- signaling (LysMCreARNTf/f) were subjected to RNASequencing after L. major infection and under pro-inflammatory stimulus. We report that L. major infection alone is enough to induce some minor HIF--dependent transcriptomic changes, while infection with L. major in combincation with pro-inflammatory stimuli induces numerous transcriptomic changes that are both dependent and independent of HIF- signaling. Additionally, by coupling transcriptomic analysis with several pathway analyses, we found HIF- suppresses pathways involved in protein translation during L. major infection in a pro-inflammatory environment. Together these findings show L. major induces a HIF--dependent transcriptomic program, but HIF- only suppresses protein translation in a pro-inflammatory environment. Thus, this work indicates the host inflammatory response, rather than the parasite, largely contributes to myeloid HIF- signaling during Leishmania infection.

microbiology↗

bioRxiv2022

In vivo Safety and Immunoactivity of Oncolytic Jurona Virus in Hepatocellular Carcinoma: A Comprehensive Proteogenomic Analysis

Oncolytic viruses can effectively unwrap a multimodal anti-tumor activity, encompassing a selective tumor cell killing and promoting a systemic anti-tumor immunity, making them a formidable foe against cancer. Among these, several members of the Rhabdoviridae family are particularly attractive as oncolytic agents due to their natural tumor selectivity and non-pathogenicity in humans. In this study, we demonstrated that intratumorally (IT) administration of Jurona virus (JURV), a novel oncolytic Rhabdovirus, induces dynamic tumor regression in human HCC xenograft and syngeneic models. Our data shows that IT injections of JURV trigger the recruitment and activation of cytotoxic T (CTLs) and decrease the tumor-associated macrophages (TAM) infiltration leading to tumor growth delay in both local and distant murine HCC tumors in a syngeneic model. Moreover, when administered concomitantly, JURV and anti-PD-1 therapy profoundly modulate the tumor microenvironment (TME) via enhanced infiltration of CTLs, suggesting that immune checkpoint blockade therapy could potentiate the immunomodulatory effect of JURV and potentially provide durable anti-tumor immunity. Our analysis of the molecular and cellular mechanism of JURV-medicated anti-cancer activity unveiled that JURV and anti-PD-1 antibodies activate different effectors of the immune system but have complementary anti-tumor activities. Furthermore, our results indicate that the abscopal effect induced by JURV is likely mediated by the mechanism regulating the T helper cell responses. Our work supports the further development of JURV as a novel immunovirotherapy platform for hepatocellular carcinoma.

immunology↗

bioRxiv2022

Proteogenomics analysis to identify acquired resistance-specific alterations in melanoma PDXs on MAPKi therapy

Therapeutic approaches to treat melanoma include small molecule drugs that target activating protein mutations in pro-growth signaling pathways like the MAPK pathway. While beneficial to the approximately 50% of patients with activating BRAFV600 mutation, mono- and combination therapy with MAPK inhibitors is ultimately associated with acquired resistance. To better characterize the mechanisms of MAPK inhibitor resistance in melanoma, we utilize patient-derived xenografts and apply proteogenomic approaches leveraging genomic, transcriptomic, and proteomic technologies that permit the identification of resistance-specific alterations and therapeutic vulnerabilities. A specific challenge for proteogenomic applications comes at the level of data curation to enable multi-omics data integration. Here, we present a proteogenomic approach that uses custom curated databases to identify unique resistance-specific alternations in melanoma PDX models of acquired MAPK inhibitor resistance. We demonstrate this approach with a NRASQ61L melanoma PDX model from which resistant tumors were developed following treatment with a MEK inhibitor. Our multi-omics strategy addresses current challenges in bioinformatics by leveraging development of custom curated proteogenomics databases derived from individual resistant melanoma that evolves following MEK inhibitor treatment and is scalable to comprehensively characterize acquired MAPK inhibitor resistance across patient-specific models and genomic subtypes of melanoma.

bioinformatics↗

bioRxiv2021

In vivo reprogramming of murine host immune response genes following Leishmania major infection.

Leishmania parasites cause cutaneous leishmaniasis (CL), a pathologic disease characterized by disfiguring, ulcerative skin lesions. Both parasite and host gene expression following infection with various Leishmania species has been investigated in vitro, but global transcriptional analysis following L. major infection in vivo is lacking. Thus, we conducted a comprehensive transcriptomic profiling study combining bulk RNA sequencing (RNA-Seq) and single-cell RNA sequencing (scRNA-Seq) to identify global changes in gene expression in vivo following L. major infection. Bulk RNA-Seq analysis revealed that host immune response pathways like the antigen processing and presentation pathway were significantly enriched amongst differentially expressed genes (DEGs) upon infection, while ribosomal pathways were significantly downregulated in infected mice compared to naive controls. scRNA-Seq analyses revealed cellular heterogeneity including distinct resident and recruited cell types in the skin following murine L. major infection. Within the individual immune cell types, several DEGs indicative of many interferon induced GTPases and antigen presentation molecules were significantly enhanced in the infected ears including macrophages (Gbp2, H2-K1, H2-Aa, H2-Ab1), resident macrophages (H2-K1, H2-D1, Gbp4, Gbp8, Gbp2), and inflammatory monocytes (Gbp2, Gbp5, Gbp7, Gbp3). Ingenuity Pathway Analysis of scRNA-Seq data indicated the antigen presentation pathway was increased with infection, while EIF2 signaling is the top downregulated pathway followed by eIF4/p70S6k and mTOR signaling in multiple cell types including macrophages, BECs, and LECs. Altogether, this transcriptomic profile highlights known recruitment of myeloid cells to lesions and recognizes a previously undefined role for EIF2 signaling in murine L. major infection in vivo. Author summaryLeishmania major cause cutaneous leishmaniasis, which is characterized by disfiguring, ulcerative skin lesions. Here, we show murine L. major-directed reprogramming of the host transcriptome in vivo. Our bulk RNA-Seq analyses revealed upregulation of antigen processing and presentation pathway, while the host ribosomal pathway was downregulated following L. major infection. Similarly, scRNA-Seq analyses revealed the upregulation of transcripts responsible for antigen presentation and host defense proteins like guanylate binding proteins (GBPs) alongside the downregulation of EIF2 signalling at the site of L. major infection. Overall, our transcriptomic dataset not only provides the comprehensive list of gene expression at the single-cell resolution, and highlights a previously undefined role for EIF2 signalling during murine L. major infection in vivo.

immunology↗