Cell Checkpoint Inhibitors Research Articles

Abstract 4723: Targeting the IDO and TDO pathway through inhibition of the aryl hydrocarbon receptor

Abstract Though checkpoint inhibitors have become the cornerstone of immune-based oncology therapy, the majority of cancer patients do not benefit from these agents. Resistance to checkpoint inhibitors is due in part to the presence of immunosuppressive molecules which prevent immune activation despite T cell checkpoint inhibition. One such immunosuppressive molecule is kynurenine. Kynurenine, metabolized from tryptophan by indoleamine-pyrrole 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase 2 (TDO2), binds to the Aryl hydrocarbon receptor (AHR) in multiple immune cell types which leads to a net immunosuppressive environment. IDO1 inhibitors combined with checkpoint inhibitors have led to substantial clinical benefit in early clinical trials, validating disruption of the kynurenine pathway as a therapeutic strategy and substantiating that metabolites within human cancers induce immune cell tolerance. Given that kynurenine can be synthesized by both IDO1 and TDO2 and that AHR is activated by endogenous ligands other than kynurenine, AHR inhibition provides a novel approach to reverse immunosuppression in a broad range of tumor types. We will present data demonstrating that AHR antagonism can affect multiple immune cell types within the tumor microenvironment, reverse immune suppression and lead to tumor growth inhibition in tumor models. In addition, AHR inhibition combined with checkpoint inhibitors leads to enhanced anti-tumor activity and improved survival in these models relative to either agent alone. Kynurenine can lead to increased levels of many immunosuppressive cytokines including IL22 in human T-cells which is reversed with AHR inhibition. In addition, we have previously shown that kynurenine addition during T-cell maturation leads to a more quiescent T-effector cells while AHR antagonism during maturation maintains T-cells in a more naïve-like, T-central memory state. In vivo studies in the B16 and the B16-IDO overexpressing models demonstrate that kynurenine-mediated activation of the AHR pathway leads to faster tumor growth and a more immunosuppressive tumor microenvironment, with FOXP3 T-cells playing a major role. There are also more suppressive myeloid cells in the IDO overexpressing model. AHR antagonism inhibits growth in the B16-IDO model and reverses the immunosuppressive microenvironment as indicated by changes in T cell and myeloid cell populations as well as AHR-dependent gene expression. Similar activity is also seen in the CT26 model. In addition, enhanced tumor growth inhibition and survival benefit is seen in combination with checkpoint inhibitors. Our data indicate that AHR is a viable target for reversing immune suppression in the tumor microenvironment and could have activity in many tumor types due to activation via kynurenine and multiple other potential ligands. Citation Format: Jeremy Tchaicha, Karen McGovern, Luis Felipe Campesato, Silvia Coma, Xiaoyan Michelle Zhang, Meghan Walsh, Jill Cavanaugh, Taha Merghoub, Jedd Wolchok, Mark Manfredi. Targeting the IDO and TDO pathway through inhibition of the aryl hydrocarbon receptor [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 4723.

Abstract 615: Increased somatic hypermutation in the immunoglobulin sequences of melanoma patients who have durable response to checkpoint inhibitor therapy

Abstract Patient immune response to tumor can drive profound and durable clinical benefit, but the contribution of antibodies to this benefit is poorly understood. To further our understanding of the role of the humoral response, we compared antibody sequence repertoires of durable responder and non-responder melanoma patients before and during checkpoint inhibitor treatment. We collected pre- and on-treatment peripheral blood from 26 melanoma patients treated with pembrolizumab (9), ipilimumab (8) or nivolumab+ipilimumab (9). Of the 26 patients, 14 were durable responders (RECIST stable disease, partial response, or complete response for at least 6 months) and 12 were non-responders. We generated natively paired heavy and light chain antibody sequences from individual IgG plasmablasts (CD19+CD20-CD38+CD3-CD14-IgA-IgM-IgD-). Sequences were obtained from a total of 26,725 plasmablasts. These sequences were used to reconstruct lineages (sets of plasmablasts likely derived from a common progenitor B cell) where each lineage shares heavy and light V genes, CDR3 length and 80% sequence similarity in CDR3s. Durable responders exhibited an increase in somatic hypermutation (SHM) after initiation of treatment. In contrast, no significant change in SHM was observed in non-responders. Higher SHM was also observed in on-treatment repertoires of responders compared with those of non-responders suggesting that T cell checkpoint inhibitors promote activation of humoral immunity in clinical responders. Furthermore, persistent antibody lineages (observed both before and during treatment) showed higher SHM than lineages observed at only a single time point. Comparison of lineages between patients identified antibodies with high sequence similarity suggesting these antibodies may have arisen from convergent selection, i.e., different patients raising antibodies against shared or similar epitopes. These putative convergent antibodies were enriched in IgG2. IgG2 usage was also higher overall in durable responders than non-responders, even in samples taken prior to treatment. The higher levels of IgG2 observed in responders and in the potentially convergent sets of antibodies suggest that IgG2 antibodies may play a role in effective anti-tumor responses. In conclusion, our analysis of plasmablast repertoires from melanoma patients suggests that treatment with checkpoint inhibitors promotes SHM in durable responders, and that responder plasmablasts are enriched in IgG2. Moreover, patients make potentially convergent antibodies that are also enriched for the IgG2 subclass. Our observations demonstrate that the humoral immune response is remodeled in patients with anti-tumor responses and support a role for the humoral arm of the immune response in driving clinical benefit in patients treated with checkpoint inhibitors. Citation Format: Ngan Nguyen, Alusha Mamchak, Mariano Severgnini, Kevin S. Williamson, Xinqi Wu, Elliott F. Drabek, Michael Manos, Xiaomu Chen, Xiaobin Tang, Zoe Amiri, Chantia Carroll, Yvonne Leung, Dongkyoon Kim, Wayne Volkmuth, Norman Greenberg, Daniel Emerling, William H. Robinson, Guy Cavet, F. Stephen Hodi. Increased somatic hypermutation in the immunoglobulin sequences of melanoma patients who have durable response to checkpoint inhibitor therapy [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 615.

Immunotherapies: Exploiting the Immune System for Cancer Treatment.

Cancer is a condition that has plagued humanity for thousands of years, with the first depictions dating back to ancient Egyptian times. However, not until recent decades have biological therapeutics been developed and refined enough to safely and effectively combat cancer. Three unique immunotherapies have gained traction in recent decades: adoptive T cell transfer, checkpoint inhibitors, and bivalent antibodies. Each has led to clinically approved therapies, as well as to therapies in preclinical and ongoing clinical trials. In this review, we outline the method by which these 3 immunotherapies function as well as any major immunotherapeutic drugs developed for treating a variety of cancers.

Targeting Immune Cell Checkpoints during Sepsis.

Immunosuppression is increasingly being recognized as one of the causes of increased morbidity and mortality during sepsis. Both innate and adaptive immune system dysfunction have been shown to cause an impaired ability to eradicate the primary infection and also lead to frequent occurrence of secondary opportunistic infections. Pre-clinical and clinical studies have shown that inhibitory immune checkpoint molecules, including programmed death-1 (PD-1), programmed death ligand-1 (PD-L1), cytotoxic T lymphocyte antigen-4 (CTLA-4), T cell membrane protein-3 (TIM-3), Lymphocyte activation-gene-3 (LAG-3) and 2B4, are upregulated during the course of sepsis. Engagement of these inhibitory molecules on various immune cells has been consistently shown to inhibit innate immune cell functions (e.g., phagocytosis, cytokine production and pathogen clearance) and also lead to impaired T cell competence. In numerous pre-clinical models of sepsis, therapeutic agents aimed at blocking engagement of inhibitory immune checkpoints on immune cells have been shown to improve innate and adaptive immune cell functions, increase host resistance to infection and significantly improve survival. Therefore, immunotherapy with immune cell checkpoint inhibitors holds significant potential for the future of sepsis therapy and merits further investigation.

Engineered cells for costimulatory enhancement combined with IL-21 enhance the generation of PD-1-disrupted CTLs for adoptive immunotherapy

Blockade of the immune cell checkpoint inhibitors programmed death 1 (PD-1) and programmed death-ligand 1 (PD-L1) has become a powerful tool in cancer treatment, which is effective across various solid cancer types and hematologic malignancies. Our previous studies showed that by reducing immune tolerance, clustered regularly interspaced short palindromic repeats-associated protein 9 (CRISPR-Cas9) modified cytotoxic T lymphocytes (CTLs) rank highly in terms of immune responses and cytotoxicity. In this study, a genetically modified K562 cell line with surface expression of 4-1BBL was developed to expand PD-1-disrupted CTLs in vitro for further adoptive immunotherapy against cancer. Our findings demonstrate that after a long-term, up to 28days, engineered cells for costimulatory enhancement (ECCE) combined with IL-21 promote the expansion of PD-1-disrupted CTLs with a less differentiated “young” phenotype, enhanced immune response and superior cytotoxic effector characteristics. These new in vitro conditions represent a nimble and cost-effective approach to developing PD-1-disrupted CTLs with improved therapeutic potential.

Abstract 2661: A broad approach for the selection of therapeutic mAbs to TNFR-superfamily members for use in combination with an anti-PD-1 mAb

Abstract The rise of therapeutic antibodies directed towards T cell checkpoint inhibitors has paved the way to a novel and exciting frontier of cancer treatments. Spearheaded by anti-CTLA-4 and PD-1/PD-L1 inhibitors, many investigators have strived to uncover combinatory therapies that could synergize with CTLA-4 and PD-1 inhibitors to improve the overall survival of cancer patients. The TNFR-superfamily has been seen as an ideal combinatory target in providing co-stimulatory T cell signals in sync with the unleashing of T cells from their CTLA-4 and PD-1 brakes. The ideal outcome of these combinations is the induction of T cell memory and effector function towards tumour cells. Current clinical-stage agents against TNFR-superfamily targets include conventional or Fc-engineered therapeutic antibodies and multimeric ligands that aim to cluster and activate targets such as OX40, 4-1BB, GITR, and CD27. For OX40 and GITR, wildtype IgG1 variants offer target cell depletion of OX40/GITR-high expressing Treg cells, and also provide agonism through FcgR-mediated crosslinking. We have taken a broad approach of screening IgG1, IgG2, Fc-engineered and tetravalent antibodies to select for potent therapeutic antibody candidates to TNFR-superfamily targets, and conducted humanized mouse studies as a screen for candidate potency. Citation Format: Cheng Chen, Xiaoniu Miao, Yao Yan, Liang Tang, Bingliang Chen, Junjian Liu, Xiaolin Liu, Michael Yu, Andy Tsun. A broad approach for the selection of therapeutic mAbs to TNFR-superfamily members for use in combination with an anti-PD-1 mAb [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 2661. doi:10.1158/1538-7445.AM2017-2661

Combination therapy of cancer with cancer vaccine and immune checkpoint inhibitors: A mathematical model.

In this paper we consider a combination therapy of cancer. One drug is a vaccine which activates dendritic cells so that they induce more T cells to infiltrate the tumor. The other drug is a checkpoint inhibitor, which enables the T cells to remain active against the cancer cells. The two drugs are positively correlated in the sense that an increase in the amount of each drug results in a reduction in the tumor volume. We consider the question whether a treatment with combination of the two drugs at certain levels is preferable to a treatment by one of the drugs alone at ‘roughly’ twice the dosage level; if that is the case, then we say that there is a positive ‘synergy’ for this combination of dosages. To address this question, we develop a mathematical model using a system of partial differential equations. The variables include dendritic and cancer cells, CD4+ and CD8+ T cells, IL-12 and IL-2, GM-CSF produced by the vaccine, and a T cell checkpoint inhibitor associated with PD-1. We use the model to explore the efficacy of the two drugs, separately and in combination, and compare the simulations with data from mouse experiments. We next introduce the concept of synergy between the drugs and develop a synergy map which suggests in what proportion to administer the drugs in order to achieve the maximum reduction of tumor volume under the constraint of maximum tolerated dose.

Neoadjuvant ipilimumab + nivolumab (IPI+NIVO) in palpable stage III melanoma: Updated data from the OpACIN trial and first immunological analyses.

9586 Background: The combination of IPI+NIVO induces high response rates and improved overall survival in late stage melanoma. T cell checkpoint inhibition is of greatest value at the moment of TCR triggering and therefore dependent on the amount of antigen present, indicating that adjuvant immunotherapy may work most efficiently, when initiated prior to surgery. Methods: Two-arm Phase 1b feasibility trial of 20 high risk AJCC stage IIIB and IIIC melanoma patients with palpable nodal disease receiving the combination of IPI 3mg/kg and NIVO 1mg/kg, either adjuvant four courses after surgery, or split two courses neo-adjuvant and two courses adjuvant. Results: In this update all 20 patients are evaluable. Neo-adjuvant application of IPI+NIVO was feasible and no surgery-associated adverse events were attributed to (neo-)adjuvant therapy. 18/20 patients had to stop early due to grade 3/4 toxicities. Neo-adjuvant IPI+NIVO reduced tumor load in 8/10 patients (3 pCR, 4 near-pCRs [minimal remaining micrometastases], 1 pPR [75% reduction], 1 SD and 1 PD). To date, after a median follow-up of 45 weeks (range 13-74), none of the responders in the neoadjuvant arm has relapsed. Relapse was observed for both non-responders within the neo-adjuvant arm and for 3 patients within the adjuvant arm. TCR sequencing and MHC tetramer-based analysis to compare the induction and expansion of tumor (neo-)antigen-specific T cell responses between both treatment arms are underway and will be presented. Conclusions: The combination of IPI+NIVO in the (neo-)adjuvant treatment setting for high risk stage III melanoma patients is feasible and induces very frequent responses. At the same time, severe grade 3/4 toxicity is more frequent than expected from stage IV melanoma patient study data. These results indicate that IPI+NIVO is a promising combination for neo-adjuvant treatment in stage III melanoma. Adjusted combination schemes are currently tested in the phase 2 OpACIN-neo trial, with the aim of reducing toxicity while preserving efficacy. Clinical trial information: NCT02437279.

Next generation approaches for tumor vaccination.

Tumor vaccines have been an attractive concept in the immunotherapy of cancer based on the central role of tumor-associated antigens in allowing the immune system to recognize cancer cells and the large variety of platforms in which to present such antigens to the immune system. Early clinical studies of vaccines, however, were largely disappointing. Recent evidence that cancer-mediated T cell suppression may prevent T cell activation is leading to renewed interest in vaccine development. The use of T cell checkpoint inhibitors alone has revolutionized the contemporary treatment of human cancer, and has suggested that the emergence of neoantigens may be an important biomarker of therapeutic response. Thus, the possibility of using more personalized vaccines targeting relevant neoantigens alone and in combination with T cell checkpoint blockade is a new area of active clinical investigation. In this review we will discuss the central role of antigens in tumor immunotherapy, describe how vaccines may be developed in the context of modern genomic profiling of tumor cells and provide a forward looking perspective on how tumor vaccines may be incorporated into the current landscape of cancer therapy.

CRISPR-Cas9-mediated disruption of PD-1 on human T cells for adoptive cellular therapies of EBV positive gastric cancer

ABSTRACTThe successful use of immune cell checkpoint inhibitors PD-1 and PD-L1, over the past 5 y has raised the concern of using immunotherapy to treat various cancers. Epstein-Barr virus-associated gastric cancer (EBVaGC) exhibits high infiltration of lymphocytes and high amplification of immune-related genes including PD-L1 as distinguished from Epstein-Barr virus-non-associated gastric cancer (EBVnGC). Here, we presume that this PD-1/PD-L1 pathway may hinder the efficacy of adoptive T cell therapy toward EBVaGC. These studies reveal possibility of generating PD-1-disrupted CTL by CRISPR-Cas9 system and demonstrate enhanced immune response of these PD-1-disrupted CTLs to the EBV-LMP2A antigen and superior cytotoxicity to the EBV-positive gastric cancer cell. In addition, when combined with low-dose radiotherapy, these PD-1-disrupted CTLs mediated an impressive antitumor effect in a xenograft mouse model of EBVaGC. Taken together, these studies illustrate PD-1/PD-L1-mediated immune tolerance of EBVaGC and provide a new strategy for targeting immune checkpoints to break the tolerance for the T cell-based adoptive therapy.

Cancer immunotherapy: from the lab to clinical applications-Potential impact on cancer centres' organisation.

This report covers the Immunotherapy sessions of the 2016 Organisation of European Cancer Institutes (OECI) Oncology Days meeting, which was held on 15th–17th June 2016 in Brussels, Belgium. Immunotherapy is a potential cancer treatment that uses an individual’s immune system to fight the tumour. In recent years significant advances have been made in this field in the treatment of several advanced cancers. Cancer immunotherapies include monoclonal antibodies that are designed to attack a very specific part of the cancer cell and immune checkpoint inhibitors which are molecules that stimulate or block the inhibition of the immune system. Other cancer immunotherapies include vaccines and T cell infusions. This report will summarise some of the research that is going on in this field and will give us an update on where we are at present.

PI3Kγ is a molecular switch that controls immune suppression.

Macrophages play critical, but opposite, roles in acute and chronic inflammation and cancer. In response to pathogens or injury, inflammatory macrophages express cytokines that stimulate cytotoxic T cells, whereas macrophages in neoplastic and parasitic diseases express anti-inflammatory cytokines that induce immune suppression and may promote resistance to T cell checkpoint inhibitors. Here we show that macrophage PI 3-kinase γ controls a critical switch between immune stimulation and suppression during inflammation and cancer. PI3Kγ signalling through Akt and mTor inhibits NFκB activation while stimulating C/EBPβ activation, thereby inducing a transcriptional program that promotes immune suppression during inflammation and tumour growth. By contrast, selective inactivation of macrophage PI3Kγ stimulates and prolongs NFκB activation and inhibits C/EBPβ activation, thus promoting an immunostimulatory transcriptional program that restores CD8+ T cell activation and cytotoxicity. PI3Kγ synergizes with checkpoint inhibitor therapy to promote tumour regression and increased survival in mouse models of cancer. In addition, PI3Kγ-directed, anti-inflammatory gene expression can predict survival probability in cancer patients. Our work thus demonstrates that therapeutic targeting of intracellular signalling pathways that regulate the switch between macrophage polarization states can control immune suppression in cancer and other disorders.

Abstract 4020: Inhibition of KIT in vivo modifies immune cell populations to improve the efficacy of checkpoint inhibitors in syngeneic mouse tumor models

Abstract The KIT receptor tyrosine kinase plays critical roles in GIST and a range of other tumor types, and when activated by somatic mutation serves as a potent oncogenic driver in these neoplasms. Whereas small molecule inhibitors of the tyrosine kinase domain of KIT have demonstrated considerable success in treatment of GIST patients with some KIT mutations, there remains a need to develop additional KIT-directed therapies for treatment of GIST patients that fail to respond to these agents, or that develop resistance during therapy. Furthermore, KIT is prominently expressed on mast cells, which promote an immune evasive environment within the tumor microenvironment by influencing the function of other immune cells, such as myeloid derived suppressor cells (MDSC’s). However, the potential impact of KIT inhibition on anti-tumor immunity has not been studied extensively to date. KTN0158 is a humanized anti-KIT IgG1 monoclonal antibody that specifically binds to the extracellular domain of KIT, and is being developed as a potential therapy for cancer and other mast cell-related diseases such as neurofibromatosis type 1 (NF1). KTN0158 inhibits KIT signaling and function in vitro and in vivo and has demonstrated anti-tumor activity in dogs with mast cell tumors expressing either wild-type or mutant KIT. In order to explore the potentially broad anti-tumor benefit of KIT inhibition by an antibody, a series of syngeneic tumor models were evaluated for sensitivity and immune cell profile changes in response to a surrogate antibody that recognizes mouse KIT (ACK2), dosed as either a single agent or in combination with T cell checkpoint inhibitors. Within this panel of tumor models, ACK2 strongly enhanced the anti-tumor activity provided by an anti-CTLA4 antibody in the colon26 model, while exhibiting no anti-tumor activity when dosed as a single agent. The colon26 cell line does not express detectable levels of KIT, and does not exhibit sensitivity to ACK2 or SCF treatment in proliferation assays performed in vitro, suggesting that this effect of ACK2 is not due to a direct effect on the tumor cells. Strikingly, ACK2 treatment was associated with pronounced effects on immune cell populations, including a profound decrease in both intra-tumoral and splenic monocytic MDSC counts. These observations suggest that KIT inhibition in vivo has the potential to modulate immune cell populations including MDSC's and mast cells, thereby targeting immune suppressive innate immune cells in the tumor and sensitizing the tumor to immune checkpoint inhibitors. Furthermore, the data provide a rationale to investigate KTN0158 in combination with immune checkpoint inhibitors across a range of human tumor types. Citation Format: Andrew J. Garton, Lori Lopresti-Morrow, Scott Seibel, Theresa LaVallee, Richard Gedrich. Inhibition of KIT in vivo modifies immune cell populations to improve the efficacy of checkpoint inhibitors in syngeneic mouse tumor models. [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 4020.

Abstract 559: Small molecule T-reg inhibitors for cancer immunotherapy

Abstract Immune evasion is a hallmark feature of tumors as they employ various strategies to suppress the immune system's ability to recognize and destroy cancer cells. T cell checkpoint inhibitors such as anti-PD1 and anti-CTLA-4 antibodies spearhead the immune response against a variety of tumors. Numerous studies suggest that the complex immunosuppressive milieux require the development of additional therapeutic agents to potentiate active drugs and thereby broaden the utility and increase the therapeutic indices of revolutionary immune-oncology treatment modalities. The presence of immunosuppressive regulatory T cells (Tregs) in tumors, keeping tumoricidal Teffector cells in check, signals poor prognosis. Thus, depletion of Tregs or impairment of Treg function is an attractive therapeutic approach for cancer. USP7, a deubiquitylase enzyme implicated as a critical node in several cancer signaling pathways, has recently emerged as an essential factor in maintaining Treg functions. Progenra identified small molecule USP7 inhibitors and employed them to show that Treg specific inhibition of USP7 results in impairment of Treg function leading to immune activation, commensurate with the ablation of Foxp3, a transcription factor that is a target of USP7 and is essential to Treg activation. This USP7 inhibitor class was subsequently lead optimized, and selected USP7 inhibitors were evaluated in ADME/PK studies and shown to impair Treg functions and to exhibit powerful anti-tumor activity against syngeneic lung tumor models in immunocompetent mice. In addition, Progenra's USP7 inhibitors significantly augmented the antitumor activity of anti-PD1 antibody, CAR T cell therapy, and cancer vaccines. These studies provide a strong rationale for the use of USP7 inhibitors in combination therapy protocols to improve the efficacy of currently approved cancer immunotherapy agents. Citation Format: Suresh Kumar, Jian Wu, Liquing Wang, Feng Wang, Matthew P. Kodrasov, Saket Agarwal, Ivan Sokirniy, Thomas Yeckley, Joseph Weinstock, Michael R. Mattern, Wayne W. Hancock. Small molecule T-reg inhibitors for cancer immunotherapy. [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 559.

Abstract 564: Combination therapy of chemokine receptor inhibition plus PD-L1 blockade potentiates antitumor effects in a murine model of breast cancer

Abstract Trafficking and expansion of myeloid derived suppressor cells (MDSCs) plays a major role in the immune suppression of tumors. MDSCs express chemokine receptors which likely mediate their recruitment to the tumor microenvironment. Suppression of T cells is also mediated by the interaction of programmed death-1 (PD-1) and its ligands which are abundantly expressed in cancer cells and immune infiltrates, including MDSCs. Here, we show that targeting both pathways through administration of a small molecule chemokine receptor antagonist (CCX9588, which blocks CCR1) and a PD-L1 monoclonal antibody (mAb) significantly reduced tumor burden in mice who received orthotopic transplants of 4T1 cells, a cell line used to model triple negative breast cancer. Primary tumor growth was modestly reduced by either agent (PD-L1 mAb or CCR1 inhibitor) alone, but the combination of CCR1 inhibitor plus PD-L1 mAb resulted in significantly reduced tumor progression as compared to either of the single agents. Furthermore, lung metastasis was also significantly reduced by CCR1 antagonist and combination treatment. Orthotropic 4T1 cell engraftment induced robust expansion of CD11b+Ly6Ghi Ly6Chi MDSCs, a subpopulation of which express CCR1. Analysis of the tumor infiltrating cells revealed that CCX9588 significantly reduced the number of MDSCs and increased CD8 T cells infiltration in primary tumors, suggesting that CCR1 blockade of MDSC trafficking in combination with PD-L1 mAb translates into reduced tumor burden, possibly through increased CD8 T cell response against the tumor. Analysis of human breast cancer patient samples from The Cancer Genome Atlas (TCGA) database revealed that the CCR1 ligands, MCP-7 (CCL7) and RANTES (CCL5) are present at significantly higher levels in breast cancers as compared to normal breast tissue. Interestingly, ligands for CCR1 and PD-1 are significantly higher in triple negative breast cancer samples than in the other breast cancer subtypes. These data are the first to show that CCR1 chemokine receptor antagonists can act synergistically with PDL-1 inhibitors, and suggest a novel approach for potentiating the activity of immune cell checkpoint inhibitors in one of the most aggressive forms of breast cancer. Citation Format: Heiyoun Jung, Linda Ertl, Karen Ebsworth, Christine Janson, Penglie Zhang, Punna Sreenivas, Thomas Schall, Israel Charo. Combination therapy of chemokine receptor inhibition plus PD-L1 blockade potentiates antitumor effects in a murine model of breast cancer. [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 564.

Revving Up Dendritic Cells while Braking PD-L1 to Jump-Start the Cancer-Immunity Cycle Motor

Although it is successful for some, most melanoma patients are refractory to T cell checkpoint inhibition. In this issue of Immunity, Merad and colleagues (2016) describe a dendritic-cell-based strategy to heighten the efficacy of therapeutic anti-PD-L1 and BRAF inhibitors in mouse melanoma models.

Abstract B017: A soluble form of CD80 inhibits PD-L1 immune suppression and stimulates T cells through CD28-specific pathways indicating potential for increased therapeutic activity over checkpoint inhibition alone

Abstract Antibodies that block the PD-1/PD-L1 pathway have demonstrated therapeutic success against several cancer types, but do not induce a response in the majority of patients. The development of treatments that provide activation signals to T cells by targeting co-stimulatory receptors, in addition to checkpoint inhibition, may increase anti-tumor immunity and the percent of responding patients. CD80/B7-1 is a co-stimulatory molecule that also interacts with PD-L1. CD80 is expressed by antigen presenting cells and ligates to CD28 providing co-stimulation during T cell activation. Furthermore, PD-L1 interacts with T cell CD80 in a reverse signaling pathway that promotes T cell anergy. The ability of CD80 to block inhibitory and promote stimulatory signaling pathways in T cell activation suggests potential therapeutic value. We have shown that soluble CD80 fused to the Fc region of IgG (CD80-Fc) blocks PD-1/PD-L1 interactions thus restoring activation of CD4+ and CD8+ T cells. Moreover, CD80-Fc was more effective in vitro than PD-1/PD-L1 monoclonal antibodies at restoring T cell activation, which suggests that CD80-Fc may be stimulating CD28 in addition to blocking PD-1/PD-L1 signaling. Stimulation of T cell CD28 in the absence of other signals induces downstream transcription, and signaling pathway activation. In fact, agonist CD28 antibody can induce functions in T cells in both naïve and activated cells, even in the absence of TCR activation. We now demonstrate that CD80-Fc does indeed stimulate T cells via CD28 activation by demonstrating comparable downstream results of CD80-Fc and agonist CD28 antibody treatment of both human and mouse T cells. Therefore, CD80-Fc may provide a novel therapeutic approach to target T cell checkpoint inhibition in addition to providing stimulatory signaling to improve T cell activation and anti-tumor immunity. Citation Format: Tiha M. Long, Suzanne Ostrand-Rosenberg. A soluble form of CD80 inhibits PD-L1 immune suppression and stimulates T cells through CD28-specific pathways indicating potential for increased therapeutic activity over checkpoint inhibition alone. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr B017.

Abstract A90: Combination therapy of chemokine receptor inhibition plus PDL-1 blockade potentiates anti-tumor effects in a murine model of breast cancer

Abstract Trafficking and expansion of myeloid derived suppressor cells (MDSCs) plays a major role in immune suppression of tumor. MDSCs express chemokine receptors which likely mediate their recruitment to the tumor microenvironment. Suppression of T cells is also mediated by the interaction of programmed death-1 (PD-1) and its ligands which are abundantly expressed in cancer cells and immune infiltrates, including MDSCs. Here, we show that targeting both pathways through administration of a small molecule chemokine receptor antagonist (CCX9588 which blocks CCR1) and an anti-PDL1 antibody significantly reduced tumor burden in an orthotopic breast cancer mouse model. Primary tumor growth and lung metastasis were modestly reduced by either agent (anti-PDL-1 or CCR1 inhibitor) alone, but the combination of CCR1 inhibitor plus anti-PDL1 antibody resulted in significantly reduced tumor burden as compared to either of the single agents. Orthotropic breast cancer cell engraftment induces robust expansion of CD11b+Ly6Ghi Ly6Chi MDSCs, a subpopulation of which express CCR1. Analysis of the tumor infiltrating cells revealed that CCX9588 significantly reduced the number of MDSCs in primary tumors, showing that CCR1 blockade of MDSC trafficking translates into reduced tumor burden. Analysis of human breast cancer patient samples from The Cancer Genome Atlas (TCGA) database revealed that the CCR1 ligands, MCP-7 (CCL7) and RANTES (CCL5) are present at significantly higher levels in breast cancers as compared to normal breast tissue. Furthermore, CCR1 and its ligands, as well as PDL-1 are significantly higher in triple negative breast cancer samples than in the other breast cancer subtypes. These data are the first to show that CCR1 chemokine receptor antagonists can act synergistically with PDL-1 inhibitors, and suggest a novel approach for potentiating the activity of immune cell checkpoint inhibitors in the most aggressive forms of breast cancer. Citation Format: Heiyoun Jung. Combination therapy of chemokine receptor inhibition plus PDL-1 blockade potentiates anti-tumor effects in a murine model of breast cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A90.

The checkpoint inhibitor TIGIT limits antitumor and antiviral CD8+ T cell responses

Success in eliciting tumor rejection by therapeutic blockade of T cell checkpoint inhibitors, such as PD-1/PD-L1 and CTLA-4, has spurred re-evaluation of additional receptors that regulate antitumor responses. Here we summarize our recent studies characterizing the role of co-inhibitory receptor TIGIT in antitumor and other chronic CD8+ T cell responses.

Abstract ES6-3: Tumor infiltrating lymphocytes in breast cancer

Abstract Interest in evaluating the presence of tumor infiltrating lymphocytes (TILs) in breast cancer is growing due to robust data supporting significant prognostic associations as well as the success of the new T cell checkpoint inhibitors in other solid organ cancer types. The positive prognostic associations between TILs present at diagnosis in triple negative (TN) and HER2-overexpressing breast cancer and better outcomes after standard adjuvant therapy strongly suggests that these breast cancer subtypes will be amendable to immunotherapeutic approaches. Current research suggests that TILs represent pre-existing host anti-tumor immunity, with the data reporting that the more TILs that are present in the tumor at diagnosis, the stronger the pre-existing immune response and better clinical outomes. However, only a minority of patients present with high levels of TILs at diagnosis. Current data also reports that biopsies from metastatic disease still can contain significant amounts of TILs. This suggests that these patients can generate an immune response even in advanced disease, albeit weak, as the median values of TILs present seem to be less. This presentation will discuss the data on the prognostic associations of TILs, our current understanding about the content of TILs in breast cancer and how they could be potentially triggered and/or suppressed. I will also discuss our recently published guidelines on how to evaluate TILs in breast cancer (Salgado R et al, 2014). Furthermore I will speculate on how we could potentially augment or re-kick start a more effective anti-tumor immune response in patients with breast cancer. I will also talk about the potential future clinical utility of TILs in breast cancer- it is currently not recommended that TILs are used in treatment decision-making concerning chemotherapy or no chemotherapy in TN breast cancer or trastuzuamb vs no trastuzumab in HER2-positive disease. Phase II efficacy data evaluating T cell checkpoint inhibitors is not yet available in advanced TN and HER2-positive breast cancer, however, phase I data using monotherapy T cell checkpoint inhibition in heavily pre-treated disease seems promising with objective responses meeting RECIST criteria seen. It is unclear what the best biomarker will be in breast cancer patients for immunotherapeutic approaches, but targeting patients with some degree of already pre-existing anti-tumor immunity seems logical. Citation Format: Sherene Loi. Tumor infiltrating lymphocytes in breast cancer [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr ES6-3.