InVivoMAb anti-mouse PD-1 (CD279)

Immune checkpoint inhibitor (ICI) therapy is effective and in routine clinical use for various cancers, but accurately identifying which patients will respond remains a significant challenge. The PET agent 18F-FDG has uptake by cancer cells as well as inflammation induced by ICI therapy, complicating and often limiting the utility of 18F-FDG for early response assessment during ICI therapy. An imaging agent that accurately distinguishes responders from nonresponders early in the course of ICI therapy could enable intensification or change of therapy for nonresponders. In this study, the 18F-labeled amino acid 18F-MeFAMP, a fluorinated analog selectively targeting system A amino acid transport, was compared with 18F-FDG in the MC38 syngeneic mouse model of ICI therapy. 18F-MeFAMP was chosen because of the relatively low uptake of system A substrates in inflammatory tissues combined with growing evidence suggesting system A transporters are involved in immunotherapy. Methods: PET/CT imaging was used to compare tumor uptake of 18F-MeFAMP with tumor uptake of 18F-FDG before and 6 d after starting dual ICIs in MC38 tumor-bearing female C57BL/6 mice. SUVs, biologic tumor volumes, and total lesion activity were measured along with selected tumor-to-organ ratios. Histogram analysis of tracer uptake was performed to assess differences in tumor activity distribution between responders and nonresponders. Results:18F-FDG showed no significant differences at baseline or after ICI regardless of response. In contrast, 18F-MeFAMP SUVs defined using a 40% of SUVmax threshold (SUV40%) decreased significantly in responders (-60.0% ± 15.6%, P < 0.0001), whereas nonresponders showed no significant change (+45.5% ± 51.2%, P = 0.09). Similar patterns were observed with SUVmax, biologic tumor volume, and total lesion activity measures with 18F-MeFAMP. Histogram analysis revealed significant 18F-MeFAMP uptake differences between groups before and after imaging (P < 0.05). 18F-MeFAMP demonstrated low uptake in common metastatic sites, including liver, lungs, and brain. Conclusion:18F-MeFAMP better detected early ICI response than 18F-FDG with favorable whole-body imaging properties. These findings support further investigation of 18F-MeFAMP for early evaluation of response to ICI and the role of system A substrates in cancer and immune cells before and during ICI.

Immune checkpoint inhibitors (ICIs) have transformed the treatment of advanced melanoma, yet their efficacy is limited by high-grade immune-related adverse events that often require treatment with systemic corticosteroids. Although corticosteroids are widely used, their impact on anti-tumor immunity remains poorly defined. Using an ICI-responsive murine melanoma model, we show that tapered systemic prednisolone administered after three cycles of combined anti-CTLA4 and anti-PD1 therapy compromises ICI-mediated tumor control, leading to delayed progression in one-third of initially responding animals. Mechanistically, prednisolone selectively suppressed CD8+ effector T-cell activation in tumor-draining lymph nodes and in the circulation, while expanding activated regulatory T-cells. These changes increased the Treg:CD8+ effector ratio, reduced cytotoxic T-cell function and blocked the early ICI-mediated induction of cytokines, including IL-2, IFNγ, VEGF, CCL3/4, IL-13, IL-3, and GM-CSF. Importantly, despite these early immunosuppressive effects, long-term tumor-specific memory responses were preserved. Autologous melanoma:T-cell cocultures validated these findings. Overall, systemic prednisolone disrupts early CD8+ T-cell-mediated anti-tumor activity but spares durable immunity, highlighting the critical importance of timing and context in the introduction of corticosteroids during ICI therapy.

Tumor cells are highly dependent on branched-chain amino acids, which can activate mechanistic target of rapamycin complex 1, but the downstream catabolite branched-chain α-keto acids (BCKAs) are not well studied in this context. Here, using clinical samples and genetically engineered mouse tumor models, we showed that tumor-derived BCKAs are secreted actively into the tumor microenvironment (TME) where they reprogram tumor-associated macrophages (TAMs) to promote tumor progression. Through genome-wide CRISPR screening, we identified Notch2 as a direct molecular target of BCKAs. BCKAs activate Notch signaling by binding to and stabilizing cleaved Notch2, functionally reprogramming TAMs and fostering an immunosuppressive TME. Mutation of the BCKA-binding site in Notch2 abolishes this effect in vivo. Together, these findings identify BCKAs as signaling metabolites that mediate tumor immunosuppression through direct sensing by Notch2.

Necroptosis is a form of programmed cell death that promotes tumor immunogenicity. To identify druggable regulators of necroptosis, we performed a small-molecule inhibitor screen and identified mouse double minute 2 (MDM2) as a suppressor of tumor necrosis factor α (TNF-α)-induced necroptosis. Genetic deletion or pharmacologic inhibition of MDM2 markedly enhanced necroptosis in a receptor-interacting protein kinase 1 (RIPK1)-dependent and p53-independent manner. Mechanistically, MDM2 interacted with RIPK3 and promoted its proteasome-mediated degradation, thereby limiting RIPK3 abundance and restraining pathway activation. In vivo, MDM2 deficiency increased tumor cell necroptosis, promoted inflammatory remodeling of the tumor microenvironment (TME), and enhanced CD8+ T cell infiltration, leading to improved tumor control. In immunologically "cold" tumor models, combining MDM2 inhibition with anti-PD-1 blockade converted tumors to a T cell-inflamed state and significantly improved therapeutic efficacy, even in p53-deficient settings. These findings identify MDM2 as a regulator of TNF-α-induced necroptosis and highlight its potential as a therapeutic target for cancer immunotherapy.