PDF file - 106K, Supplemental Table 1 Summary of NY-ESO-1 Immune responses.

PDF file - 144K, Supplemental Figure 1 NY-ESO-1 antigen-specific CD4+ T cell lines were CD8 negative.

PDF file - 428K, Supplementary Figure S1. Depigmentation after combination therapy. Supplementary Figure S2. Tumor immune infiltrate after immunomodulation. Supplementary Figure S3. PD-1 expression on T cell subsets. Supplementary Figure S4. Effect of anti-CTLA4 on TILs.

Novel strategies to reprogram tumor-infiltrating myeloid cells for cancer immunotherapy are urgently needed, given that the primary and acquired resistance to immune checkpoint blockade (ICB) therapy has hindered the overall success of immunotherapy. Modified vaccinia virus Ankara (MVA) is a highly attenuated, non-replicative vaccinia virus and an approved vaccine against smallpox and monkeypox. Here we report rational engineering of recombinant MVA, MQ833, by removing three immune suppressive genes, E5R, E3L, and WR199, from the MVA genome and inserting three transgenes encoding Flt3L, OX40L, and IL-12. Intratumoral (IT) delivery of MQ833 generates potent antitumor responses dependent on CD8 + T cells, neutrophils, and M1-like macrophages, the nucleic acid-sensing pathways mediated by MDA5/STING, and interferon feedback loop. IT MQ833 promotes the recruitment and activation of neutrophils and inflammatory monocytes into the injected tumors, depletion of M2-like macrophages, and expansion of M1-like macrophages, generating potent antitumor immunity against tumors resistant to ICB.

Immune checkpoint blockade (ICB) therapy has brought hope to many cancer patients, but the response rate is low in many cancer types, and acquired resistance to ICB can develop over time. Oncolytic viruses are promising therapeutic agents for advanced cancers. Modified vaccinia virus Ankara (MVA) is an attenuated, replication-deficient poxvirus safe for human use, making it a favorable platform for cancer immunotherapy. Our first-generation recombinant MVA has shown promising antitumor efficacy in multiple murine tumor models due to the deletion of the E5R gene (encoding an inhibitor of the DNA sensor cGAS) from the MVA genome and the insertion of two membrane-anchored transgenes – Flt3L and OX40L, which leads to the activation of the host innate and adaptive antitumor immunity. Here in this study, we engineered our second-generation recombinant MVA (MQ833) with the deletion of two more viral immune evasion genes – E3L and WR199, and the insertion of IL12 anchored to the extracellular matrix to mitigate toxicity. Intratumoral (IT) delivery of MQ833 resulted in an 80-100% cure in the mouse B16-F10 melanoma model, which is dependent on nucleic acid-sensing and IFN signaling pathways. Single-cell RNA sequencing analysis revealed that IT MQ833 injection reprogrammed the tumor microenvironment into an immune-stimulating state, by activating CD8+ and CD4+ T cells, depleting regulatory T cells, recruiting and activating neutrophils, and polarizing M1 macrophages. Interestingly, MQ833 treatment cured 70% of B2m knock-out melanomas likely due to combined effects of IL-12 and type I and II IFN. Loss of MHC-I is the most common mechanism of tumor resistance to ICB. Hence, our results support the use of MQ833 for ICB-resistant tumors.

Background CD4 and CD8 T cells are genetically and functionally distinct cell subsets of the adaptive immune system that play pivotal roles in immune surveillance and disease control. During development in the thymus, transcription factors ThPOK and Runx3 regulate the differentiation and maturation of these two lineages into single positive T cells that enter the periphery with mutually exclusive expression of either the CD4 or CD8 co-receptor.1–2 Despite our expectation that these two cell fates are fixed, mature CD4+CD8+ double positive (DP) T cells have been described in the context of numerous immunological responses, including cancer, but their molecular and functional properties and therapeutic relevance remain controversial and largely unknown.3–5 Methods Our lab has identified and characterized a heterogenous DP T cell population in murine and human melanoma tumors comprised of CD4 and CD8 T cells re-expressing the opposite co-receptor and a parallel uptake in the opposite cell type’s phenotype and function. Using CD4 (Trp1) and CD8 (Pmel) transgenic TCR T cells specific to B16 melanoma antigens gp75 and gp100 respectively, we demonstrate the re-expression of the opposite co-receptor following adoptive T cell transfer in B16 melanoma tumor bearing mice. Results Specifically, up to 50% of transferred CD4 Trp1 T cells will re-express CD8 to become a DP T cell in the tumor microenvironment. Further, these CD4 derived DP T cells upregulate CD8 lineage regulator Runx3 and cytolytic genes Gzmb, Gzmk, and Prf1 to become potent cytotoxic T cells. Alternatively, a subset of CD8 Pmel T cells differentiate into DP T cells characterized by the increased expression of CD4, ThPOK, and regulatory marker FoxP3 (figure 1). In addition, we utilized 10x single cell and ATAC sequencing to further characterize these divergent DP T cell populations among open repertoire T cells isolated from murine and human melanoma tumors. Conclusions Our findings highlight the capability of single positive T cells to differentiate in response to antigen and local stimuli into novel T cell subsets with polyfunctional characteristics. The resulting cell subsets will potentially affect the tumor microenvironment in distinct ways. Our studies may inform therapeutic approaches to identify antigen specific T cells as well as innovative signaling pathways to target when genetically engineering T cells to optimize cytotoxic function in the setting of adoptive cell therapy. Ethics Approval The human biospecimen analyses were approved by Memorial Sloan Kettering Cancer Center IRB #06-107 References Ellmeier W, Haust L & Tschismarov R. Transcriptional control of CD4 and CD8 coreceptor expression during T cell development. Cell Mol Life Sci 2013;70:4537–4553. Luckey MA, et al. The transcription factor ThPOK suppresses Runx3 and imposes CD4+ lineage fate by inducing the SOCS suppressors of cytokine signaling. Nature Immunology 2014; 15, 638–645. Bohner P, et al. Double positive CD4(+)CD8(+) T Cells are enriched in urological cancers and favor T Helper-2 polarization. Front Immunol 2019; 10, 622. Nascimbeni M, Shin E-C, Chiriboga L, Kleiner DE & Rehermann B. Peripheral CD4(+)CD8(+) T cells are differentiated effector memory cells with antiviral functions. Blood 2004;104:478–486. Nishida K, et al. Clinical importance of the expression of CD4+CD8+ T cells in renal cell carcinoma. Int Immunol 2020;32:347–357.

Adoptive cell therapy has emerged as a viable strategy to treat cancer. T cells that recognize tumor antigens can be reinvigorated ex-vivo or autologous T cells can be genetically modified to express anti-tumor T cell receptors (TCRs) or chimeric antigen receptors (CARs). However, once re-infused into patients, these tumor specific T cells are subjected to immunosuppressive signals within the tumor. A critical immune checkpoint within tumors is phosphatidylserine (PS), a phospholipid that is exposed on apoptotic cells and tumor cells. Innate cells exposed to PS secrete suppressive cytokines that can significantly impair the function of tumor specific T cells. Antibodies that target PS can reactivate anti-tumor immunity by reducing the number of MDSCs in tumors and promoting the maturation of functional APCs. Our lab has shown that the mouse chimeric version of PS Targeting monoclonal antibody Bavituximab (1N11), in combination with transgenic CD4+ T cells that recognize melanoma antigen Trp1, can regress advanced melanoma tumors in mice. Here, we demonstrate a 2nd generation CAR T cell, that binds Trp1 on the surface of B16 melanoma, in combination with 1N11 can improve anti-tumor activity and survival in B16 tumor bearing mice. Flow cytometry analysis of immune responses in the tumor of mice treated with tumor specific T cells and 1N11 shows a decrease in M2 macrophages and FoxP3+ regulatory T cells. These findings highlight that diminishing suppressive mechanisms locally with PS targeting can enhance the efficacy of transgenic TCR and CAR T cells to improve the outcome in patients with advanced-stage melanoma. Our studies may inform the design of clinical trials combining PS Targeting antibodies with CAR T cell therapy in solid tumors.

609 Objectives: The adoptive transfer of tumor-targeting T cells offers a promising approach to treating tumors; however, clinical success has been limited. Activation and proliferation are key events after contact is made between the T cells and the tumor antigen. OX40 and GITR are co-stimulatory molecules, whose signaling is important for differential immune responses mediated by CD4 or CD8 T cells. In our preliminary testing, surface expression of OX40 and GITR on tumor antigen-specific T cells increased markedly following co-culture with tumor cells. The aim of this study was to investigate whether immune-PET imaging of OX40 and GITR enables non-invasive in vivo monitoring of functionally active T cells and, thus, whether it may assist in predicting therapeutic response in real time at an early stage. Methods: We conjugated antibodies targeting OX40 and GITR via their lysine amino acids to 89Zr-DFO using isothiocyanate-bearing derivative of DFO. We utilized our established Rag1-/- mouse model lacking mature B and T cells engrafted with B16 melanoma, and injected melanoma-specific CD4-Trp1 T cells via the tail vein. Melanoma mice not receiving CD4-Trp1 T cells served as control groups. The mice underwent serial PET imaging following intravenous administration of 89Zr-DFO-OX40, -GITR and -IgG (isotype). We quantified the tracer uptake on PET and performed autoradiography and immunohistochemistry on the harvested tumor tissues. Results: 89Zr-DFO-conjugated antibodies specifically targeted their ligands OX40 and GITR in vivo as detected by immuno-PET and enabled non-invasive serial monitoring of the T cell surface activation markers at the tumor site. Tumor/T cell-to-normal organ ratios were favorable, resulting in high image contrast (Fig. 1, Table 1). Ex vivo autoradiography confirmed the specific binding of the radiolabeled antibodies. In contrast, uptake of 89Zr-DFO-OX40 and -GITR at the tumor site in control mice not injected with CD4-Trp1 T cells was minimal. Conclusions: Preliminary results indicate that OX40 and GITR are up-regulated on CD4-Trp1 T cells at the tumor site, enabling specific binding of corresponding 89Zr-DFO-labelled antibodies in vivo. This approach is now being further developed to characterize the pharmacokinetics (PK) of marker expression over time and to investigate the association between OX40 and GITR expression at the tumor site and treatment efficacy. Monitoring the activation state of the T cells may also predict therapy success, individualize immunosuppressive therapy regimen, and, ultimately, optimize the design of new clinical trials.

A viable strategy to treat advanced cancers includes transferring of tumor-specific T cells. T cells that recognize tumor antigens can be expanded and reinvigorated ex-vivo. Furthermore, autologous T cells can be genetically modified to express antitumor T cell receptors or chimeric antigen receptors (CARs). Although the potency and specificity of tumor-specific T cells can be manipulated ex vivo, once reinfused into patients, the T cells are subjected to immunosuppressive mechanisms established by the tumor. An important immune checkpoint regulator within tumors is phosphatidylserine (PS), a phospholipid that is exposed on apoptotic cells, tumor cells and tumor endothelium. Innate immune cells exposed to PS secrete suppressive cytokines and chemokines that can significantly impair the function and activation of antitumor T cells. Antibodies that target PS have been shown to reactivate antitumor immunity by polarizing tumor-associated macrophages into a proinflammatory M1 phenotype, reducing the number of MDSCs in tumors and promoting the maturation of dendritic cells into functional APCs. Our lab has previously shown that a PS targeting monoclonal antibody (mch1N11), in combination with transgenic CD4+ T cells that recognize the melanoma antigen Trp1, can regress very advanced melanomas in all treated mice. Here, we further those studies with data showing that a 2nd-generation CAR T cell that binds Trp1 on the surface of B16 melanoma, in combination with mch1N11 can improve antitumor activity and overall survival in B16 tumor-bearing mice. Additionally, in vitro killing assays with antigen-specific T cells sorted from the tumor reveal that mch1N11 enhances the cytolytic function of these T cells against B16 melanoma. Flow cytometry analysis of local immune responses in the tumors of animals treated with tumor-specific T cells and mch1N11 showed a decrease in anti-inflammatory (M2) macrophages and FoxP3+ regulatory T cells. These findings highlight that diminishing suppressive mechanisms locally with mch1N11 can enhance the efficacy of transgenic TCR and CAR T cells to improve the outcome in patients with advanced-stage melanoma. Our studies may inform the design of clinical trials combining PS targeting antibodies with CAR T cell therapy in solid tumors. Citation Format: Sara Schad, Daniel Hirschhorn-Cymerman, Sadna Budhu, Hong Zhong, Xia Yang, Joseph Shan, Steven King, Taha Merghoub, Jedd Wolchok. Phosphatidylserine targeting antibody enhances antitumor activity of CAR T cells in mouse melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3568.

A significant proportion of cancer patients do not respond to immune checkpoint blockade. To better understand the molecular mechanisms underlying these treatments, we explored the role of CD4+Foxp3- T cells expressing PD-1 (4PD1hi) and observed that 4PD1hi accumulate intratumorally as a function of tumor burden. Interestingly, CTLA-4 blockade promotes intratumoral and peripheral 4PD1hi increases in a dose-dependent manner, while combination with PD-1 blockade mitigates this effect and improves anti-tumor activity. We found that lack of effective 4PD1hi reduction after anti-PD-1 correlates with poor prognosis. Mechanistically, we provide evidence that mouse and human circulating and intra-tumor 4PD1hi inhibit T cell functions in a PD-1/PD-L1 dependent fashion and resemble follicular helper T cell (TFH)-like cells. Accordingly, anti-CTLA-4 activity is improved in TFH deficient mice.

A significant proportion of cancer patients do not respond to immune checkpoint blockade therapy. To deepen our understanding of the mechanisms of resistance to immunotherapy, we studied a population of CD4+Foxp3− T cells expressing PD-1 (4PD1hi), which we found to be up-regulated in B16-melanoma bearing mice after CTLA-4 blockade in association with limited response to treatment. We observed that 4PD1hi accumulate intratumorally as a function of tumor burden in untreated tumor-bearing hosts. Interestingly, CTLA-4 blockade promotes intratumoral and peripheral 4PD1hiincreases in a dose-dependent manner, while combination with PD-1 blockade mitigates this effect in both B16-bearing mice and melanoma patients and significantly improves anti-tumor activity. In addition, we found that persistence of high levels of 4PD1hi after PD-1 blockade correlates with poor prognosis in melanoma patients. Mechanistically, we show that mouse and human circulating and intra-tumor 4PD1hi inhibit T-cell functions in a PD-1/PD-L1 dependent fashion. In addition, we found that mouse and human 4PD1hi resemble follicular-helper-T-cell(TFH)-like cells and that CTLA-4 blockade activity is improved in TFH deficient mice. These findings broaden our understanding of the mechanisms that limit anti-tumor immunity, providing an additional explanation for the incremental activity of combined CTLA-4 and PD-1 blockade. Our study also defines 4PD1hi as a new prognostic and pharmacodynamic biomarker for the design of optimal checkpoint blockade combination schedules and dosage.

OX40 is a T cell co-stimulatory receptor that can enhance the magnitude and durability of T cell immune responses. Anti-OX40 agonist antibodies have shown significant single agent tumoricidal activity in preclinical models, and can combine effectively with other immunomodulatory antibodies, targeted therapies and vaccines. OX40 agonists are able to counteract the immunosuppressive tumor microenvironment and promote tumor-specific cellular immunity via at least two distinct mechanisms: 1) promoting OX40 forward signaling in tumor-specific T cells; and 2) co-engaging Fcγ receptors expressed by tumor-associated effector cells, and facilitating the selective elimination of OX40high intratumoral regulatory T cells. INCAGN1949, an anti-OX40 human IgG1 antibody, was selected based on its ability to optimally enhance T cell responsiveness under conditions of suboptimal T cell receptor stimulation. INCAGN1949 was shown to mediate effective apical OX40 clustering that is translated into effective downstream activation of the NFκB pathway. Notably, INCAGN1949 was shown to maintain a sigmoidal dose response curve across a broad range of antibody concentrations. This suggests a wide therapeutic window and may be advantageous for dosing considerations. By contrast, evaluation of reference OX40 antibodies indicated an inverted U-shaped dose response curve, leading to impaired T cell responses at high concentrations. INCAGN1949 was selected for clinical development based on its optimal agonist profile, further reinforced by its ability to combine with other co-inhibitory and co-stimulatory antibodies to augment T cell responsiveness. Prior to human testing, the pharmacology and tolerability of INCAGN1949 was evaluated in non-human primates (NHPs). Pharmacokinetic (PK) and pharmacodynamic (PD) parameters were evaluated including longitudinal measurements of serum cytokines, immune cell populations, activation state and T cell-mediated immune responses to reporter vaccine antigens. INCAGN1949 exhibited a linear PK profile and was well tolerated at all doses tested, with no maximum tolerated dose established. Co-administration of INCAGN1949 and vaccines in NHPs showed an immune-based PD signature across a broad exposure range. These studies were in line with in vitro findings and support a wide PD range for INCAGN1949 in patients. An important secondary mechanism of INCAGN1949 is the ability of its IgG1 Fc region to mediate selective depletion of OX40high intratumoral regulatory T cells. Immunohistochemistry and flow cytometry analyses support the validity of this regulatory T cell depletion mechanism in a range of tumors. The functional in vitro and in vivo attributes of INCAGN1949 make it suitable for clinical development. It is currently under evaluation in a Phase 1/2 study in subjects with advanced or metastatic tumors (NCT02923349). Citation Format: Ana M. Gonzalez, Mariana L. Manrique, Lukasz Swiech, Thomas Horn, Ekaterina Breous, Jeremy Waight, David Savitsky, Yuqi Liu, Shiwen Lin, Christopher Clarke, Taha Merghoub, Daniel Hirschhorn-Cymerman, David Schaer, Gerd Ritter, Jennifer Pulini, Kevin Heller, Peggy Scherle, Gregory Hollis, Reid Huber, Marc van Dijk, Jennifer Buell, Robert Stein, Nicholas Wilson. INCAGN1949, an anti-OX40 antibody with an optimal agonistic profile and the ability to selectively deplete intratumoral regulatory T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4703. doi:10.1158/1538-7445.AM2017-4703

Abstract A viable strategy to treat advanced cancers includes transferring of tumor-specific T cells. T cells that recognize tumor antigens can be expanded and reinvigorated ex-vivo. Furthermore, autologous T cells can be genetically modified to express anti-tumor T cell receptors or chimeric antigen receptors. Although the potency and specificity of tumor-specific T cells can be manipulated ex-vivo, once re-infused into patients, the T cells are subjected to immunosuppressive mechanisms established by the tumor. An important immune checkpoint regulator within tumors is phosphatidylserine (PS). Innate immune cells exposed to PS secrete suppressive cytokines and chemokines that can significantly impair the function and activation of anti-tumor T cells. Therefore, monoclonal antibodies that block PS activity can increase the anti-tumor potency of transferred T cells to treat aggressive cancers. Here we show that a PS targeting monoclonal antibody in combination with CD4+ T cells that recognize the melanoma antigen Trp1 can regress very advanced melanomas in all treated mice. Combination of anti-Trp1 CD4+ T cells with other immunomodulatory modalities such as anti-OX40 antibodies, can achieve equivalent treatment rates but these are typically accompanied by severe immune related adverse events. In contrast, in this setting, PS blockade did not show any off-target toxicities. Flow cytometry analysis revealed lower levels of CD206 expression concomitant with higher activation markers in macrophages and neutrophils in tumors from anti-PS treated mice. These results suggest that diminishing suppressive mechanisms locally in adoptive transfer protocols is a highly desirable strategy that can eliminate tumors while minimizing related adverse events. Citation Format: Daniel Hirschhorn-Cymerman, Sara Sara Schad, Sadna Budhu, Zhong Hong, Xia Yang, Hutchins T. Jeff, Bruce D. Freimark, Michael J. Gray, Jedd Wolchok, Taha Merghoub. Targeting phosphatidylserine in combination with adoptive T cell transfer eliminates advanced tumors without off-target toxicities in a melanoma preclinical model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1651. doi:10.1158/1538-7445.AM2017-1651

Abstract OX40 (CD134, TNFRSF4) is a T cell co-stimulatory receptor that potentiates T cell receptor (TCR) signaling during CD4+ and CD8+ T cell priming, effector cell differentiation and memory T cell recall responses. In preclinical mouse tumor models, surrogate anti-OX40 agonist antibodies have shown remarkable single agent anti-tumor efficacy, as well as the ability to combine effectively with other immunomodulatory antibodies and immune education strategies, such as therapeutic cancer vaccines. Agonistic antibodies targeting OX40 are predicted to counteract the immunosuppressive tumor microenvironment and promote tumor-specific T cell immunity via two primary mechanisms: 1) binding and activating OX40 signaling in tumor-specific effector and memory T cells, thereby enhancing their responsiveness to tumor-associated antigens, and 2) co-engaging Fcγ receptors expressed by tumor-associated effector cells, and facilitating the selective depletion of intratumoral regulatory T cells. INCAGN01949 is a novel fully human IgG1 monoclonal antibody identified using the Retrocyte Display™ platform that is being developed for the treatment of advanced malignancies. INCAGN01949 recognizes human and cynomolgus monkey OX40 with comparable binding affinity. INCAGN01949 has been optimized to potently mediate receptor forward signaling under conditions of suboptimal TCR stimulation, leading to features like enhanced production of TNFα and IFNγ, and concomitant suppression of IL-10. INCAGN01949 achieves this functionality through OX40 clustering and downstream activation of the NFκB pathway in T cells, which is sustained across a broad range of antibody concentrations. Consistent with mouse preclinical tumor models, OX40 was found to be selectively overexpressed by intratumoral regulatory T cells from a variety of primary human tumor samples. Commensurate with its human IgG1 Fc region, INCAGN01949 can effectively co-engage activating Fcγ receptors on immune effector cells, including natural killer cells and macrophages. Therefore INCAGN01949 has the potential to mediate selective effector cell activity toward intratumoral populations of regulatory T cells. The biophysical and functional attributes of INCAGN01949 make it suited for clinical development, both as a single agent and in combination with other immunomodulatory antibodies or immune education strategies. Citation Format: Ana Maria Gonzalez, Mariana L. Manrique, Ekaterina Breous, David Savitsky, Jeremy Waight, Randi Gombos, Yuqi Liu, Shiwen Lin, Taha Merghoub, Daniel Hirschhorn-Cymerman, Gerd Ritter, Jedd Wolchok, Peggy Scherle, Gregory Hollis, Reid Huber, Marc Van Dijk, Robert Stein, Nicholas S. Wilson. INCAGN01949: an anti-OX40 agonist antibody with the potential to enhance tumor-specific T-cell responsiveness, while selectively depleting intratumoral regulatory T cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3204.

La presente invention concerne des anticorps qui se lient specifiquement au recepteur d'OX40 humain (OX40) et des compositions comprenant de tels anticorps. Selon un aspect specifique, les anticorps se lient specifiquement a l'OX40 humain et modulent l'activite d'OX40, par exemple, renforcent, activent ou suscitent l'activite d'OX40, ou reduisent, desactivent ou inhibent l'activite d'OX40. La presente invention concerne egalement des methodes de traitement de troubles, tels que le cancer, en administrant un anticorps qui se lie specifiquement a l'OX40 humain et module l'activite d'OX40, par exemple, renforce, active ou suscite l'activite d'OX40. L'invention concerne egalement des methodes de traitement de troubles ou maladies auto-immuns ou inflammatoires, en administrant un anticorps qui se lie specifiquement a l'OX40 humain et module l'activite d'OX40, par exemple, reduit, desactive ou inhibe l'activite d'OX40.

The present disclosure provides multispecific ( e.g ., bispecific) antibodies that specifically bind to human GITR and/or human OX40 as well as compositions comprising such antibodies. In a specific aspect, the multispecific antibodies specifically bind to human GITR and OX40 and modulate GITR and/or OX40 activity, e.g ., enhance, activate, or induce GITR and/or OX40 activity, or reduce, deactivate, or inhibit GITR and/or OX40 activity. The present disclosure also provides methods for treating disorders, such as cancer, by administering a multispecific antibody that specifically binds to human GITR and/or OX40 and modulates GITR and/or OX40 activity, e.g ., enhances, activates, or induces GITR and/or OX40 activity. Also provided are methods for treating autoimmune or inflammatory diseases or disorders, by administering a multispecific antibody that specifically binds to human GITR and/or OX40 and modulates GITR and/or OX40 activity, e.g ., reduces, deactivates, or inhibits GITR and/or OX40 activity.

Targeting tumour-infiltrating suppressive myeloid cells with a selective PI3Kγ inhibitor overcomes resistance to checkpoint blockade therapy in various mouse myeloid-rich tumour models. Therapeutic blockade of immune checkpoints with antibodies against CTLA-4 and PD-1 has proved effective against some cancer types, but clinical benefit has been limited to a subset of patients. Here Olivier De Henau et al. show that resistance to checkpoint blockade is associated with a high level of infiltration by suppressive myeloid cells in various mouse tumour models. In addition, targeting the myeloid-derived suppressor cells with a selective inhibitor of the γ isoform of phosphoinositide 3-kinase (PI3Kγ) increases sensitivity to checkpoint blockade therapy in a melanoma mouse model. Recent clinical trials using immunotherapy have demonstrated its potential to control cancer by disinhibiting the immune system. Immune checkpoint blocking (ICB) antibodies against cytotoxic-T-lymphocyte-associated protein 4 or programmed cell death protein 1/programmed death-ligand 1 have displayed durable clinical responses in various cancers1. Although these new immunotherapies have had a notable effect on cancer treatment, multiple mechanisms of immune resistance exist in tumours. Among the key mechanisms, myeloid cells have a major role in limiting effective tumour immunity2,3,4. Growing evidence suggests that high infiltration of immune-suppressive myeloid cells correlates with poor prognosis and ICB resistance5,6. These observations suggest a need for a precision medicine approach in which the design of the immunotherapeutic combination is modified on the basis of the tumour immune landscape to overcome such resistance mechanisms. Here we employ a pre-clinical mouse model system and show that resistance to ICB is directly mediated by the suppressive activity of infiltrating myeloid cells in various tumours. Furthermore, selective pharmacologic targeting of the gamma isoform of phosphoinositide 3-kinase (PI3Kγ), highly expressed in myeloid cells, restores sensitivity to ICB. We demonstrate that targeting PI3Kγ with a selective inhibitor, currently being evaluated in a phase 1 clinical trial (NCT02637531), can reshape the tumour immune microenvironment and promote cytotoxic-T-cell-mediated tumour regression without targeting cancer cells directly. Our results introduce opportunities for new combination strategies using a selective small molecule PI3Kγ inhibitor, such as IPI-549, to overcome resistance to ICB in patients with high levels of suppressive myeloid cell infiltration in tumours.

Using the immune system to control cancer has been investigated for over a century. Yet it is only over the last several years that therapeutic agents acting directly on the immune system have demonstrated improved overall survival for cancer patients in phase III clinical trials. Furthermore, it appears that some patients treated with such agents have been cured of metastatic cancer. This has led to increased interest and acceleration in the rate of progress in cancer immunotherapy. Most of the current immunotherapeutic success in cancer treatment is based on the use of immune-modulating antibodies targeting critical checkpoints (CTLA-4 and PD-1/PD-L1). Several other immune-modulating molecules targeting inhibitory or stimulatory pathways are being developed. The combined use of these medicines is the subject of intense investigation and holds important promise. Combination regimens include those that incorporate targeted therapies that act on growth signaling pathways, as well as standard chemotherapy and radiation therapy. In fact, these standard therapies have intrinsic immune-modulating properties that can support antitumor immunity. In the years ahead, adoptive T-cell therapy will also be an important part of treatment for some cancer patients. Other areas which are regaining interest are the use of oncolytic viruses that immunize patients against their own tumors and the use of vaccines against tumor antigens. Immunotherapy has demonstrated unprecedented durability in controlling multiple types of cancer and we expect its use to continue expanding rapidly.