IFNγ is a central node of cancer immune equilibrium.

Tumors in immune equilibrium are held in balance between outgrowth and destruction by the immune system. The equilibrium phase defines the duration of clinical remission and stable disease, and escape from equilibrium remains a major clinical problem. Using a non-replicating HSV-1 vector expressing interleukin-12 (d106S-IL12), we developed a mouse model of therapy-induced immune equilibrium, a phenomenon previously seen only in humans. This immune equilibrium was centrally reliant on interferon-γ (IFNγ). CD8+ T cell direct recognition of MHC class I, perforin/granzyme-mediated cytotoxicity, and extrinsic death receptor signaling such as Fas/FasL were all individually dispensable for equilibrium. IFNγ was critically important and played redundant roles in host and tumor cells such that IFNγ sensing in either compartment was sufficient for immune equilibrium. We propose that these redundant mechanisms of action are integrated by IFNγ to protect from oncogenic or chronic viral threats and establish IFNγ as a central node in therapy-induced immune equilibrium.

cIAP1/2 Antagonism Induces Antigen-Specific T Cell-Dependent Immunity.

Checkpoint blockade immunotherapy has failed in pancreatic cancer and other poorly responsive tumor types in part due to inadequate T cell priming. Naive T cells can receive costimulation not only via CD28 but also through TNF superfamily receptors that signal via NF-κB. Antagonists of the ubiquitin ligases cellular inhibitor of apoptosis protein (cIAP)1/2, also called second mitochondria-derived activator of caspases (SMAC) mimetics, induce degradation of cIAP1/2 proteins, allowing for the accumulation of NIK and constitutive, ligand-independent activation of alternate NF-κB signaling that mimics costimulation in T cells. In tumor cells, cIAP1/2 antagonists can increase TNF production and TNF-mediated apoptosis; however, pancreatic cancer cells are resistant to cytokine-mediated apoptosis, even in the presence of cIAP1/2 antagonism. Dendritic cell activation is enhanced by cIAP1/2 antagonism in vitro, and intratumoral dendritic cells show higher expression of MHC class II in tumors from cIAP1/2 antagonism-treated mice. In this study, we use in vivo mouse models of syngeneic pancreatic cancer that generate endogenous T cell responses ranging from moderate to poor. Across multiple models, cIAP1/2 antagonism has pleiotropic beneficial effects on antitumor immunity, including direct effects on tumor-specific T cells leading to overall increased activation, increased control of tumor growth in vivo, synergy with multiple immunotherapy modalities, and immunologic memory. In contrast to checkpoint blockade, cIAP1/2 antagonism does not increase intratumoral T cell frequencies. Furthermore, we confirm our previous findings that even poorly immunogenic tumors with a paucity of T cells can experience T cell-dependent antitumor immunity, and we provide transcriptional clues into how these rare T cells coordinate downstream immune responses.

PD-1 blockade and CDK4/6 inhibition augment nonoverlapping features of T cell activation in cancer.

We performed single-cell RNA-sequencing and T cell receptor clonotype tracking of breast and ovarian cancer patients treated with the CDK4/6 inhibitor ribociclib and PD-1 blockade. We highlight evidence of two orthogonal treatment-associated phenomena: expansion of T cell effector populations and promotion of T cell memory formation. Augmentation of the antitumor memory pool by ribociclib boosts the efficacy of subsequent PD-1 blockade in mouse models of melanoma and breast cancer, pointing toward sequential therapy as a potentially safe and synergistic strategy in patients.

Radiation combines with immune checkpoint blockade to enhance T cell priming in a murine model of poorly immunogenic pancreatic cancer.

Radiation has been a pillar of cancer therapy for decades. The effects of radiation on the anti-tumour immune response are variable across studies and have not been explicitly defined in poorly immunogenic tumour types. Here, we employed combination checkpoint blockade immunotherapy with stereotactic body radiation therapy and examined the effect on tumour growth and immune infiltrates in subcutaneous and orthotopic mouse models of pancreatic cancer. Although immune checkpoint blockade and radiation were ineffective alone, their combination produced a modest growth delay in both irradiated and non-irradiated tumours that corresponded with significant increases in CD8+ T cells, CD4+ T cells and tumour-specific T cells as identified by IFNγ ELISpot. We conclude that radiation enhances priming of tumour-specific T cells in poorly immunogenic tumours and that the frequency of these T cells can be further increased by combination with immune checkpoint blockade.

cIAP1/2 antagonism eliminates MHC class I-negative tumors through T cell-dependent reprogramming of mononuclear phagocytes.

Loss of major histocompatibility complex (MHC) class I and interferon-γ (IFN-γ) sensing are major causes of primary and acquired resistance to checkpoint blockade immunotherapy. Thus, additional treatment options are needed for tumors that lose expression of MHC class I. The cellular inhibitor of apoptosis proteins 1 and 2 (cIAP1/2) regulate classical and alternative nuclear factor κB (NF-κB) signaling. Induction of noncanonical NF-κB signaling with cIAP1/2 antagonists mimics costimulatory signaling, augmenting antitumor immunity. We show that induction of noncanonical NF-κB signaling induces T cell-dependent immune responses, even in ß2-microglobulin (ß2M)-deficient tumors, demonstrating that direct CD8 T cell recognition of tumor cell-expressed MHC class I is not required. Instead, T cell-produced lymphotoxin reprograms both mouse and human macrophages to be tumoricidal. In wild-type mice, but not mice incapable of antigen-specific T cell responses, cIAP1/2 antagonism reduces tumor burden by increasing phagocytosis of live tumor cells. Efficacy is augmented by combination with CD47 blockade. Thus, activation of noncanonical NF-κB stimulates a T cell-macrophage axis that curtails growth of tumors that are resistant to checkpoint blockade because of loss of MHC class I or IFN-γ sensing. These findings provide a potential mechanism for controlling checkpoint blockade refractory tumors.

Remission-Stage Ovarian Cancer Cell Vaccine with Cowpea Mosaic Virus Adjuvant Prevents Tumor Growth.

Ovarian cancer is the deadliest gynecological malignancy. Though most patients enter remission following initial interventions, relapse is common and often fatal. Accordingly, there is a substantial need for ovarian cancer therapies that prevent relapse. Following remission generated by surgical debulking and chemotherapy, but prior to relapse, resected and inactivated tumor tissue could be used as a personalized vaccine antigen source. The patient's own tumor contains relevant antigens and, when combined with the appropriate adjuvant, could generate systemic antitumor immunity to prevent relapse. Here, we model this process in mice to investigate the optimal tumor preparation and vaccine adjuvant. Cowpea mosaic virus (CPMV) has shown remarkable efficacy as an immunostimulatory cancer therapy in ovarian cancer mouse models, so we use CPMV as an adjuvant in a prophylactic vaccine against a murine ovarian cancer model. Compared to its codelivery with tumor antigens prepared in three other ways, we show that CPMV co-delivered with irradiated ovarian cancer cells constitutes an effective prophylactic vaccine against a syngeneic model of ovarian cancer in C57BL/6J mice. Following two vaccinations, 72% of vaccinated mice reject tumor challenges, and all those mice survived subsequent rechallenges, demonstrating immunologic memory formation. This study supports remission-stage vaccines using irradiated patient tumor tissue as a promising option for treating ovarian cancer, and validates CPMV as an antitumor vaccine adjuvant for that purpose.