Advanced ovarian cancer (OC) patients have a poor 5-year survival of only 28%, emphasizing the medical need for improved therapies. Adjuvant immunotherapy could be an attractive approach since OC is ...an immunogenic disease and the presence of tumor-infiltrating lymphocytes has shown to positively correlate with patient survival. Among these infiltrating lymphocytes are natural killer (NK) cells, key players involved in tumor targeting, initiated by signaling via activating and inhibitory receptors. Here, we investigated the role of the DNAM-1/TIGIT/CD96 axis in the anti-tumor response of NK cells toward OC. Ascites-derived NK cells from advanced OC patients showed lower expression of activating receptor DNAM-1 compared to healthy donor peripheral blood NK cells, while inhibitory receptor TIGIT and CD96 expression was equal or higher, respectively. This shift to a more inhibitory phenotype could also be induced in vitro by co-culturing healthy donor NK cells with OC tumor spheroids, and in vivo on intraperitoneally infused NK cells in SKOV-3 OC bearing NOD/SCID-IL2Rγnull (NSG) mice. Interestingly, TIGIT blockade enhanced degranulation and interferon gamma (IFNγ) production of healthy donor CD56
dim
NK cells in response to OC tumor cells, especially when DNAM-1/CD155 interactions were in place. Importantly, TIGIT blockade boosted functional responsiveness of CD56
dim
NK cells of OC patients with a baseline reactivity against SKOV-3 cells. Overall, our data show for the first time that checkpoint molecules TIGIT/DNAM-1/CD96 play an important role in NK cell responsiveness against OC, and provides rationale for incorporating TIGIT interference in NK cell-based immunotherapy in OC patients.
Cytotoxic T lymphocyte antigen-4 has become recognized as one of the key negative regulators of adaptive immune responses, having a central role in the maintenance of peripheral tolerance and in ...shaping the repertoire of emergent T cell responses. Concurrent recognition of the potential importance of inhibitory immune regulators in limiting antitumor responses, either as a result of chronic antigenic stimulation or the self-nature of many tumor-selective target antigens, has led to the development of cytotoxic T lymphocyte antigen-4-blocking antibodies as therapeutic anticancer agents. Following extensive preclinical modeling, these agents have entered clinical trials, where they are showing encouraging activity in heavily pretreated patients with advanced-stage disease, particularly with melanoma or renal carcinoma. Finding ways to dissociate antitumor activity from adverse immune events should enable actualization of their therapeutic potential in the coming years.
Identification of cytotoxic T-lymphocyte antigen-4 (CTLA-4) as a key negative regulator of T-cell activity led to development of the fully human, monoclonal antibody ipilimumab to block CTLA-4 and ...potentiate antitumor T-cell responses. Animal studies first provided insight into the ability of an anti-CTLA-4 antibody to cause tumor regression, particularly in combination regimens. Early clinical studies defined ipilimumab pharmacokinetics and possibilities for combinability. Phase II trials of ipilimumab in advanced melanoma showed objective responses, but a greater number of patients had disease stabilization. In a phase III trial, ipilimumab was the first agent to demonstrate an improvement in overall survival in patients with previously treated, advanced melanoma. The adverse event profile associated with ipilimumab was primarily immune-related. Adverse events can be severe and life-threatening, but most were reversible using treatment guidelines. Ipilimumab monotherapy exhibits conventional and new patterns of activity in advanced melanoma, with a delayed separation of Kaplan-Meier survival curves. The observation of some new response patterns with ipilimumab, which are not captured by standard response criteria, led to novel criteria for the evaluation of immunotherapy in solid tumors. Overall, lessons from the development of ipilimumab contributed to a new clinical paradigm for cancer immunotherapy evolved by the Cancer Immunotherapy Consortium.
Novel therapeutic approaches combining immune-checkpoint inhibitors are needed to improve clinical outcomes for patients with cancer. Lymphocyte-activation gene 3 (LAG-3) is an immune-checkpoint ...molecule that inhibits T-cell activity and antitumor immune responses, acting through an independent mechanism from that of programmed death-1 (PD-1) and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4). Here, we describe the development and preclinical characterization of relatlimab, a human antibody that binds to human LAG-3 with high affinity and specificity to block the interaction of LAG-3 with the ligands MHC II and fibrinogen-like protein-1, and to reverse LAG-3-mediated inhibition of T-cell function in vitro. Consistent with previous reports, in mouse models, the combined blockade of LAG-3 and PD-1 with surrogate antibodies resulted in enhanced antitumor activity greater than the individual blockade of either receptor. In toxicity studies in cynomolgus monkeys, relatlimab was generally well tolerated when combined with nivolumab. These results are consistent with findings from the RELATIVITY-047 phase II/III trial showing that relatlimab combined with nivolumab is a well-tolerated regimen that demonstrates superior progression-free survival compared with nivolumab monotherapy in patients with unresectable or metastatic melanoma.
Costimulation via CD137 (4-1BB) enhances antitumor immunity mediated by cytotoxic T lymphocytes. Anti-CD137 agonist antibodies elicit mild liver inflammation in mice, and the maximum tolerated dose ...of Urelumab, an anti-human CD137 agonist monoclonal antibody, in the clinic was defined by liver inflammation-related side effects. A protease-activated prodrug form of the anti-mouse CD137 agonist antibody 1D8 (1D8 Probody therapeutic, Pb-Tx) was constructed and found to be selectively activated in the tumor microenvironment. This construct, which encompasses a protease-cleavable linker holding in place a peptide that masks the antigen binding site, exerted antitumor effects comparable to the unmodified antibody but did not result in liver inflammation. Moreover, it efficaciously synergized with both PD-1 blockade and adoptive T-cell therapy. Surprisingly, minimal active Pb-Tx reached tumor-draining lymph nodes, and regional lymphadenectomy did not abrogate antitumor efficacy. By contrast, S1P receptor-dependent recirculation of T cells was absolutely required for efficacy. The preferential cleavage of the anti-CD137 Pb-Tx by tumor proteases offers multiple therapeutic opportunities, including neoadjuvant therapy, as shown by experiments in which the Pb-Tx is given prior to surgery to avoid spontaneous metastases.
CD4+ Tregs impede T cell responses to tumors. They express multiple inhibitory receptors that support their suppressive functions, including T cell Ig and ITIM domain (TIGIT). In melanoma patients, ...we show that Tregs exhibit increased TIGIT expression and decreased expression of its competing costimulatory receptor CD226 as compared with CD4+ effector T cells, resulting in an increased TIGIT/CD226 ratio. Tregs failed to upregulate CD226 upon T cell activation. TIGIT+ Tregs are highly suppressive, stable, and enriched in tumors. TIGIT and CD226 oppose each other to augment or disrupt, respectively, Treg suppression and stability. A high TIGIT/CD226 ratio in Tregs correlates with increased Treg frequencies in tumors and poor clinical outcome upon immune checkpoint blockade. Altogether, our findings show that a high TIGIT/CD226 ratio in Tregs regulates their suppressive function and stability in melanoma. They provide the rationale for novel immunotherapies to activate CD226 in Tregs together with TIGIT blockade to counteract Treg suppression in cancer patients.
The programmed death-1 (PD-1) pathway limits the function of virus-specific T cells during chronic infection. We previously showed that blockade of the PD-1 pathway increases HIV-1-associated T cell ...function in vitro. However, the effect of PD-1 blockade on HIV-1 disease progression in vivo has not been examined. As in humans, HIV-1-infected humanized BALB/c-Rag2(-/-)γc(-/-) (Rag-hu) mice express elevated levels of PD-1 on T cells during chronic infection. To examine the effect of PD-1 blockade on disease progression, Rag-hu mice with chronic HIV-1 infection were treated with a blocking mAb directed against programmed cell death-1 ligand-1, the ligand for PD-1. Programmed cell death-1 ligand-1-treated Rag-hu mice exhibited a progressive decrease in the HIV-1 plasma viral load, with a 7-fold decrease by day 7, a 20-fold decrease by day 14, a 178-fold decrease by day 21, and a 269-fold decrease by day 28 postinitiation of treatment. By day 7, the percentage of CD4(+) T cells was statistically higher in the treated compared with the untreated group, and this trend was sustained throughout the 28-d treatment period. Moreover, there was a strong inverse correlation between plasma viral load and the percentage of both CD4(+) (r = -0.66; p < 0.0001) and CD8(+) (r = -0.64; p < 0.0001) T cells in the treated mice but not the untreated mice. This study provides "proof of concept" that humanized mice can be used to examine the effects of immunotherapeutic interventions on HIV-1 infection. Furthermore, to our knowledge, these data demonstrate for the first time that blockade of the PD-1 pathway reduces HIV-1 viral loads.
Triple-negative breast cancer (TNBC) is an aggressive and molecularly diverse breast cancer subtype typified by the presence of p53 mutations (∼80%), elevated immune gene signatures and neoantigen ...expression, as well as the presence of tumor infiltrating lymphocytes (TILs). As these factors are hypothesized to be strong immunologic prerequisites for the use of immune checkpoint blockade (ICB) antibodies, multiple clinical trials testing single ICBs have advanced to Phase III, with early indications of heterogeneous response rates of <20% to anti-PD1 and anti-PDL1 ICB. While promising, these modest response rates highlight the need for mechanistic studies to understand how different ICBs function, how their combination impacts functionality and efficacy, as well as what immunologic parameters predict efficacy to different ICBs regimens in TNBC. To address these issues, we tested anti-PD1 and anti-CTLA4 in multiple models of TNBC and found that their combination profoundly enhanced the efficacy of either treatment alone. We demonstrate that this efficacy is due to anti-CTLA4-driven expansion of an individually unique T-cell receptor (TCR) repertoire whose functionality is enhanced by both intratumoral Treg suppression and anti-PD1 blockade of tumor expressed PDL1. Notably, the individuality of the TCR repertoire was observed regardless of whether the tumor cells expressed a nonself antigen (ovalbumin) or if tumor-specific transgenic T-cells were transferred prior to sequencing. However, responsiveness was strongly correlated with systemic measures of tumor-specific T-cell and B-cell responses, which along with systemic assessment of TCR expansion, may serve as the most useful predictors for clinical responsiveness in future clinical trials of TNBC utilizing anti-PD1/anti-CTLA4 ICB.
Recent advances in cancer treatment with checkpoint blockade of receptors such as CTLA-4 and PD-1 have demonstrated that combinations of agents with complementary immunomodulatory effects have the ...potential to enhance antitumor activity as compared to single agents. We investigated the efficacy of immune-modulatory interleukin-21 (IL-21) combined with checkpoint blockade in several syngeneic mouse tumor models. After tumor establishment, mice were administered recombinant mouse IL-21 (mIL-21) alone or in combination with blocking monoclonal antibodies against mouse PD-1 or CTLA-4. In contrast to monotherapy, IL-21 enhanced antitumor activity of mCTLA-4 mAb in four models and anti-PD-1 mAb in two models, with evidence of synergy for one or both of the combination treatments in the EMT-6 and MC38 models. The enhanced efficacy was associated with increased intratumoral CD8+ T cell infiltrates, CD8+ T cell proliferation, and increased effector memory T cells, along with decreased frequency of central memory CD8+ T cells. In vivo depletion of CD8+ T cells abolished the antitumor activities observed for both combination and monotherapy treatments, further supporting a beneficial role for CD8+ T cells. In all studies, the combination therapies were well tolerated. These results support the hypothesis that the combination of recombinant human IL-21 with CTLA-4 or PD-1 monoclonal antibodies could lead to improved outcomes in cancer patients.
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