Errington-Mais, F. | Search bioRxiv

bioRxiv2024

Enhancing oncolytic virotherapy by exosome-mediated microRNA reprograming of the tumour microenvironment.

BackgroundThere has been limited success of cancer immunotherapies in the treatment of ovarian cancer (OvCa) to date, largely due to the immunosuppressive tumour microenvironment (TME). Tumour-associated macrophages (TAMs) are a major component of both the primary tumour and malignant ascites, promoting tumour growth, angiogenesis, metastasis, chemotherapy resistance and immunosuppression. Differential microRNA (miRNA) profiles have been implicated in the plasticity of TAMs. Therefore, delivering miRNA to TAMs to promote an anti-tumour phenotype is a novel approach to reverse their pro-tumour activity and enhance the efficacy of cancer immunotherapies. Oncolytic viruses (OVs) preferentially replicate in tumour cells making them ideal vehicles to deliver miRNA mimetics to the TME. Importantly, miRNA expressed by OVs get packaged within tumour-derived extracellular vesicles (TDEVs), and release of TDEV is augmented by OV infection, thus enhancing the dissemination of miRNA throughout the TME. MethodSmall RNAseq was used to identify differentially expressed miRNA during TAM generation and following LPS/IFN{gamma} stimulation to induce an anti-tumour phenotype. Two differentially expressed miRNA identified, miR-155 and miR-19a, were cloned into oncolytic rhabdovirus (ORV), and anti-tumour efficacy was investigated using both in vitro and in vivo models of OvCa. ResultsThis study demonstrates that ORV infection enhances TDEV production in OvCa cell lines both in vitro and in vivo and that TDEV are preferentially taken up by myeloid cells, including TAMs. Small RNAseq identified 23 miRNAs that were significantly upregulated in anti-tumour TAMs, including miR-155-5p. While 101 miRNAs were downregulated during pro-tumour TAM differentiation, including miR-19a-3p. Culturing TDEV expressing miR-155 or miR-19a with TAMs reversed their immunosuppressive activity, as measured by T cell proliferation. While ORV-miR-155 enhanced the generation of anti-tumour T cells, only ORV-miR19a significantly improved survival of mice bearing ovarian tumours. ConclusionThis study demonstrates (i) that arming ORVs with immunomodulatory miRNA is an effective approach to deliver miRNA to myeloid cells within the TME and (ii) that miRNA have the capacity to reverse the tumour promoting properties of TAMs and improve the efficacy of cancer immunotherapies, such as OV.

cancer biology↗

bioRxiv2024

Combination of oncolytic Maraba virus with immune checkpoint blockade overcomes therapy resistance in an immunologically cold model of advanced melanoma with dysfunctional T cell receptor signalling.

BackgroundOver the past decade, cancer immunotherapies have revolutionised the treatment of melanoma; however, responses vary across patient populations. Recently, baseline tumour size has been identified as an independent prognostic factor for overall survival in melanoma patients receiving immune checkpoint inhibitors (ICIs). MG1 is a novel oncolytic agent with broad tumour tropism that has recently entered early phase clinical trials. The aim of this study was to characterise T cell responses in human and mouse melanoma models following MG1 treatment and to establish if features of the tumour immune microenvironment (TIME) at two distinct tumour burdens would impact the efficacy of oncolytic virotherapy. MethodsHuman 3D in vitro priming assays were performed to measure anti-tumour and anti-viral T cell responses following MG1 infection. TCR sequencing, T2 killing assay, and peptide recall assays were used to assess the evolution of the TCR repertoire, and measure specific T cell responses, respectively. In vivo, subcutaneous 4434 melanomas were characterised using RNAseq, immunohistochemistry (IHC), and flow cytometry. The effectiveness of intra-tumoural MG1 was assessed in advancing 4434 tumours and the generation of anti-tumour and anti-viral T cells measured by splenocyte recall assays. Finally, combination MG1 and -PD-1 therapy was investigated in advanced 4434 tumours. ResultsMG1 effectively primed functional cytotoxic T cells (CTLs) against tumour associated antigens (TAA) as well as virus-derived peptides, as assessed using peptide recall and T2 killing assays, respectively. TCR sequencing revealed that MG1-primed CTL comprised larger clusters of similar CDR3 amino acid sequences compared to controls. In vivo testing of MG1 demonstrated that MG1 monotherapy was highly effective at treating early disease, resulting in 90% cures; however, the efficacy of MG1 reduced as the disease burden (local tumour size) increased, and the addition of -PD-1 was required to overcome resistance in more advanced disease. Differential gene expression profiles revealed that increased tumour burden was associated with an immunologically colder TIME. Furthermore, analysis of TCR signalling in advancing tumours demonstrated a different dynamic of TCR engagement compared to smaller tumours, in particular a shift in antigen recognition by CD4+ cells, from conventional to regulatory subset. ConclusionCombination of MG1 with PD-1 overcomes therapy resistance in an immunologically cold model of advanced melanoma.

cancer biology↗