Chordoma is a midline neoplasm accounting for approximately 20% of primary spinal tumors. Due to chordoma's locally aggressive nature, patients experience high rates of disease progression and have limited treatment options, highlighting an unmet clinical need. This underscores the importance of exploring novel therapeutic strategies. Oncolytic viral (OV) therapy uses genetically modified viruses to selectively replicate in tumor cells, mediate tumor cell lysis, and initiate a pro-inflammatory response within the infected tumor microenvironment (TME). The aim of the study was to evaluate the anti-chordoma effect of the replicating oncolytic adenovirus Delta-24-RGD. The efficacy of Delta-24-RGD was assessed using in vitro approaches, ex vivo bone scaffolds, and in vivo murine models. Additionally, we explored the underlying mechanism of OV cytotoxicity, immunogenic cell death (ICD), brachyury modulation, and reshaping of the TME towards a pro-inflammatory state. Delta-24-RGD achieved viral infectivity, oncolysis, and ICD across multiple human chordoma cell lines and ex vivo bone scaffold models. In vivo murine xenograft models of human CH22 and U-CH1 chordoma treated with Delta-24-RGD resulted in tumor volume reduction, enhanced overall survival, and microenvironmental transcriptional modulation. Analyzing a human chordoma tissue microarray, the immunosuppressive macrophage marker CD163 was associated with shortened recurrence-free survival. Using macrophage/chordoma co-culture, OV infection achieved a reduction in macrophage CD163 expression and a concomitant pro-inflammatory macrophage polarization. The immunosuppressive chordoma TME is associated with clinical outcomes and our data suggests OV infection reverses this deleterious immunosuppressive profile. These studies provide a framework for future clinical implementation amongst chordoma patients.

Current literature suggests IDH-mutant astrocytoma contains several molecular subgroups. In this study we are interested in determining the connection between different molecular subgroups with grade and/or survival. A cohort of 470 Mayo Clinic adult patients (≥18 years, 56.2% male) with primary IDH-mutant astrocytoma diagnosed by WHO 2021 criteria were examined. Results were validated in an independent cohort of 614 Mayo Clinic Neuropathology consult patients and 235 TCGA patients. The Mayo Clinic Practice cohort confirmed the association of CDKN2A/B deletion with overall survival (OS, homozygous vs hemizygous vs intact, 2.7 vs 9.6 vs 17.2 years, p < 0.001). PTEN deletion was also associated with poor OS (7.3 vs 17.4years, p < 0.001). Increased number of copy number alterations was associated with OS (continuous variable, HR = 1.027, p < 0.001). Carrying one or more copies of the germline risk allele at rs55705857 was associated with earlier age of onset (median age 33 vs 35 yrs, p = 0.01), and a shorter OS after adjusting for age, grade, sex and treatment (HR = 1.81, p = 0.007). The Mayo Clinic Neuropathology Consult cohort and TCGA were utilized to validate age of onset and survival, respectively. Unsupervised clustering of the copy number alterations identified several clinically significant groups that may define pathways to disease progression. Losses of chromosomes 11p, 13q, 1p and 10q were all associated with reduced overall survival in the Mayo Clinic cohort. Patients with hemizygous loss of CDKN2A/B, loss of PTEN, increased number of copy number alterations, specific chromosomal arm losses or rs55705857 germline risk allele have reduced overall survival.

Immunotherapy has yet to make significant gains in glioblastoma (GBM) treatment, due in part to GBM-mediated immune suppression. Increasing evidence points to critical roles for tumor-derived extracellular vesicles (EVs) and immunosuppressive myeloid cells as key factors in this process. Immunophenotyping of the tumor-immune microenvironment was performed using ultrasonic aspirate collected during GBM resection by high-dimensional flow cytometry. EVs collected from patient-derived GBM cell lines were used to condition myeloid cells collected from healthy donors to generate immunosuppressive myeloid cells. siRNA was used to knockdown TIGIT and/or NLRP3 expression prior to EV conditioning. T cell co-culture studies were performed with donor-matched T cells. Immune phenotyping of the tumor microenvironment and EV-conditioned myeloid cells revealed similar immunomodulatory protein expression across myeloid cell populations, with particularly elevated TIGIT expression. Knockdown of TIGIT reduced the immunosuppressive polarization of myeloid cells, resulting in improved T cell function. This finding proceeded in an NLRP3-dependent manner, with substantial co-expression of TIGIT and NLRP3 expression prior to knockdown, and concomitant knockdown of NLRP3 abrogating the effect of TIGIT knockdown. TIGIT expression correlated with increased IL-13 expression, and IL-13 blockade unmasked a pro-inflammatory myeloid cell phenotype. TIGIT expression in myeloid cells in the GBM microenvironment is a functional marker of immunosuppressive activity, with TIGIT knockdown reducing IL-13 expression and unmasking the pro-inflammatory activity of NLRP3. This study bolsters our understanding of the immunosuppressive complexities of the GBM microenvironment, and supports attenuation of immunosuppressive myeloid cell activity as a strategy to restore immune function in GBM.