Cancer stem-like cells (CSC) play key roles in long-term tumor propagation and metastasis, but their dynamics during disease progression are not understood. Tumor relapse in patients with initially excised skin squamous cell carcinomas (SCC) is characterized by increased metastatic potential, and SCC progression is associated with an expansion of CSC. Here, we used genetically and chemically-induced mouse models of skin SCC to investigate the signaling pathways contributing to CSC function during disease progression. We found that CSC regulatory mechanisms change in advanced SCC, correlating with aggressive tumor growth and enhanced metastasis. β-catenin and EGFR signaling, induced in early SCC CSC, were downregulated in advanced SCC. Instead, autocrine FGFR1 and PDGFRα signaling, which have not been previously associated with skin SCC CSC, were upregulated in late CSC and promoted tumor growth and metastasis, respectively. Finally, high-grade and recurrent human skin SCC recapitulated the signaling changes observed in advanced mouse SCC. Collectively, our findings suggest a stage-specific switch in CSC regulation during disease progression that could be therapeutically exploited by targeting the PDGFR and FGFR1 pathways to block relapse and metastasis of advanced human skin SCC.

Malignant brain tumors remain incurable diseases. Although much effort has been devoted to improving patient outcome, multiple factors such as the high tumor heterogeneity, the strong tumor-induced immunosuppressive microenvironment, and the low mutational burden make the treatment of these tumors especially challenging. Thus, novel therapeutic strategies are urgent. Oncolytic viruses (OVs) are biotherapeutics that have been selected or engineered to infect and selectively kill cancer cells. Increasingly, preclinical and clinical studies demonstrate the ability of OVs to recruit T cells and induce durable immune responses against both virus and tumor, transforming a “cold” tumor microenvironment into a “hot” environment. Besides promising clinical results as a monotherapy, OVs can be powerfully combined with other cancer therapies, helping to overcome critical barriers through the creation of synergistic effects in the fight against brain cancer. Although many questions remain to be answered to fully exploit the therapeutic potential of OVs, oncolytic virotherapy will clearly be part of future treatments for patients with malignant brain tumors.

Background: Glioblastoma (GBM) is an aggressive brain tumor associated with poor outcome and limited treatment options. Chimeric antigen receptor (CAR) T cells targeting cell surface antigens were shown to induce tumor regression in patients with GBM, although efficacy was transient. To broaden the range of tumor-restricted antigens, we developed CAR T cells targeting Tenascin-C (TNC), a secreted extracellular matrix protein that is overexpressed in GBM and plays a critical role in tumor progression. Methods: Second-generation CAR T cells were engineered to target the alternatively spliced fibronectin type III (FNIII)-D domain of TNC using a single-chain variable fragment isolated from the R6N antibody and coupled to a CD28 costimulatory domain. TNC-CAR T cells were evaluated in vitro for antigen specificity, activation, and cell proliferation using TNC-expressing patient-derived GBM cell lines cultured as adherent cells or as neurospheres. Reactivity toward purified TNC protein, tumor supernatant, and ex vivo patient tumor samples was also assessed. Cytotoxic CAR T-cell activity was tested against TNC-positive and TNC-negative GBM cell lines, including bystander effects mediated by secreted TNC. In vivo efficacy and safety were determined in NOD scid gamma mice bearing patient-derived GBM tumors. Results: TNC-CAR T cells demonstrated activation when exposed to TNC-positive GBM cells, cell-derived supernatants, or purified TNC protein. They exhibited potent cytotoxicity against TNC-expressing, GBM-derived adherent cells and neurospheres, and induced bystander killing of TNC-negative cells in the presence of either TNC-secreting cells or purified TNC. In vivo, TNC-CAR T cells efficiently infiltrated tumors, triggered cancer cell apoptosis, and significantly extended survival of mice bearing patient-derived GBM, with no evidence of off-tumor toxicity. Notably, TNC-CAR T cells were activated exclusively in the presence of tumor samples and showed no reactivity toward patient-derived non-tumor tissues. Conclusions: Targeting the alternatively spliced FNIII-D domain of TNC with CAR T cells offers a promising therapeutic approach for GBM. TNC-CAR T cells demonstrated specific tumor recognition, robust antitumor activity and the ability to induce bystander effects mediated by secreted TNC. Their efficacy in preclinical models, combined with a favorable safety profile, underscores their potential for clinical translation.

Oncolytic adenoviruses (OAds) present limited efficacy in clinics. The insertion of therapeutic transgenes into OAds genomes, known as “arming OAds”, has been the main strategy to improve their therapeutic potential. Different approaches were published in the decade of the 2000s, but with few comparisons. Most armed OAds have complete or partial E3 deletions, leading to a shorter half-life in vivo. We generated E3+ OAds using two insertion sites, After-fiber and After-E4, and two different splice acceptors linked to the major late promoter, either the Ad5 protein IIIa acceptor (IIIaSA) or the Ad40 long fiber acceptor (40SA). The highest transgene levels were obtained with the After-fiber location and 40SA. However, the set of codons of the transgene affected viral fitness, highlighting the relevance of transgene codon usage when arming OAds using the major late promoter.