Maintenance of Immune Balance: Effects of Targeted and Immune Therapies最新文献

{"title":"Abstract A134: Single-cell roadmap of the evolution of T-cell response during anti-LAG3 and anti-PD1 combination treatment in metastatic melanoma patients","authors":"J. Huuhtanen, Henna Hakanen, T. Lönnberg, O. Dufva, K. Peltola, S. Mäkelä, M. Hernberg, P. Bono, K. Anna, S. Mustjoki","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A134","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A134","url":null,"abstract":"Despite the impressive impact of CTLA4 and PD1-targeted immuno-oncologic (IO) therapies, a large proportion of patients fail to respond. The observed variance in treatment efficacy has been linked to heterogeneity in the immune cell distribution of individual patients. Lymphocyte activation gene 3 (LAG3, CD223) is one of the newest IO targets entering the clinic. Triggering of LAG3 on T-cells by HLA-DR -ligands has a well-established role in the negative regulation of T-cell function. Although for example on activated regulatory T-cells (Treg) LAG3 is widely expressed, the detailed effects of LAG3 on various cell types are still unknown. We profiled over 30,000 CD45+ lymphocytes cells using a novel paired single-cell RNA and T-cell receptor (TCR) αβ chain (10x Genomics) sequencing method for peripheral blood samples from two patients receiving anti-LAG3 and anti-PD1 combination treatment in multicentre phase I trial. The sequenced cells were from IO-treatment naive metastatic melanoma patients from before, after 4 weeks and after 12 weeks of the start of therapy. For validation, we performed TCRβ-sequencing and flow cytometry analysis of a larger cohort of melanoma patients (n = 12) enrolled in the same study. To gain in-depth understanding of the immune cell subsets, we used a recently described method to seek matching mutual neighbors between patients to normalize the interpatient variation to enable a systematic comparison across patients. To identify phenotypic clusters, we used a graph theory-based clustering method SNN-Cliq and built predictive machine learning classifiers to assess the reproducibility of learnt clusters. After optimizing our choice of input genes and parameters, we identified 16 distinct clusters that define the T-cell roadmap of anti-LAG3 and anti-PD1 treatment that includes 6 CD8+, 8 CD4+ (including Treg cluster), and 2 other clusters. We identified 4 CD8+ T-cell clusters of increasing cytotoxicity profile using a cytotoxicity score and the pseudotime algorithm Monocle3. The most highly cytotoxic clusters increased and a cluster of lower cytotoxicity score decreased during the treatment. In addition, we defined 2 CD8+ exhaustion clusters. During treatment, we observed a decrease in the exhausted T-cell cluster defined by LAG3 and PDCD1 expression, but an increase in TIGIT+ exhaustion cluster. Furthermore, ordering the cluster of FOXP3+ Treg cells along pseudotime trajectory revealed two different fates for Treg cells, where the other fate showed significantly decreased expression of LAG3 and PDCD1, adding evidence of the effect of the treatment on Treg cells. TCRαβ analysis revealed 19 individual expanded clonotypes of size of 100 sequenced individual cells. The true transcriptomic heterogeneity of identical clonotypes was revealed as most clonotypes spanned several of the 16 different clusters, challenging our view of clonal expansion. Furthermore, we were able to assess the temporal phenotypic changes in individual clo","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"21 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75176864","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A146: Systematic discovery of immune regulatory mechanisms in tumor cells","authors":"D. Pan, A. Kobayashi, Peng Jiang, Guocheng Yuan, X. Liu, John G Doench, Xintao Qiu, P. Rao, Henry W. Long, Myles A. Brown, K. Wucherpfennig, L. F. Andrade, Rong En Tay, Adrienne M. Luoma, Daphne M Tsoucas, Klothilda Lim","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A146","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A146","url":null,"abstract":"Many human cancers are resistant to immunotherapy for reasons that are poorly understood. We used a genome-scale CRISPR/Cas9 screen to identify mechanisms of tumor cell resistance to killing by cytotoxic T-cells, the central effectors of antitumor immunity. Inactivation of >100 genes sensitized mouse B16F10 melanoma cells to killing by T-cells, including Pbrm1, Arid2 and Brd7, which encode components of the PBAF form of the SWI/SNF chromatin remodeling complex. Loss of PBAF function increased tumor cell sensitivity to interferon-γ, resulting in enhanced secretion of chemokines that recruit effector T-cells. Treatment-resistant tumors became responsive to immunotherapy when Pbrm1 was inactivated. In many human cancers, expression of PBRM1 and ARID2 inversely correlated with expression of T-cell cytotoxicity genes, and Pbrm1-deficient murine melanomas were more strongly infiltrated by cytotoxic T-cells. Citation Format: Deng Pan, Aya Kobayashi, Peng Jiang, Guo-Cheng Yuan, X. Shirley Liu, John Doench, Xintao Qiu, Prakash Rao, Henry Long, Myles A. Brown, Kai W. Wucherpfennig, Lucas Ferrari de Andrade, Rong En Tay, Adrienne M. Luoma, Daphne Tsoucas, Klothilda Lim. Systematic discovery of immune regulatory mechanisms in tumor cells [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A146.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"93 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"80480896","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A125: Pulsatile MEK inhibition improves antitumor immunity and T-cell function in Kras mutant lung cancer","authors":"Hyejin Choi, Jiehui Deng, Shuai Li, Tarik Silk, E. Brea, Jonathan A Boiarsky, E. Akbay, Paul D. Smith, T. Merghoub, Kwok-kin Wong, J. Wolchok","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A125","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A125","url":null,"abstract":"KRAS is one of the most commonly identified driver oncogene in non-small cell lung cancer (NSCLC) and is commonly associated to NSCLC that is refractory to current available modalities of treatment. Targeted therapy to inhibit MEK has shown promising tumor growth control, but is followed by quick rebound of tumor growth. Recently, cancer immunotherapy has shown great promise by activating T-cells that are suppressed in the tumor microenvironment, especially through targeting co-inhibitory molecules and their counterparts. We sought to identify the most effective therapy for the treatment of KRAS mutant NSCLC patients by targeting cancer cells and activating immune infiltrates at the same time, by studying the impact of MEK inhibition on the immune microenvironment. We utilized Kras mutant lung cancer cell lines and Kras mutant transgenic lung cancer animals. We found that pulsatile treatment of MEK inhibitors activates T-cells and releases their proliferation suppression ex vivo and in vivo. Both selumetinib and trametinib showed highly increased CTLA-4 expression and mild increase of PD-1 in T-cells in pulsatile treatment, compared to continuous treatment in vivo. In addition, the pulsatile schedule alone showed superior antitumor effect and delayed drug resistance, in contrast with continuous dosing schedule. Based on the above observations, we combined pulsatile MEK inhibitor treatment and anti-CTLA-4 and found that it showed most prolonged survival of Kras tumor-bearing mice. All together, our findings will set the foundation for a combinatorial therapeutic strategy using pulsatile targeted therapy together with immunotherapy in patients, to optimally enhance tumor apoptosis and promote long-term immune response simultaneously. Citation Format: Hyejin Choi, Jiehui Deng, Shuai Li, Tarik Silk, Elliott J. Brea, Jonathan Boiarsky, Esra A. Akbay, Paul D. Smith, Taha D. Merghoub, Kwok-Kin Wong, Jedd D. Wolchok. Pulsatile MEK inhibition improves antitumor immunity and T-cell function in Kras mutant lung cancer [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A125.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"21 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"85675233","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A140: A T-cell utilizing bispecific anti-CD3/GD2 construct mediates superior in vitro efficacy compared to CH14.18 mAb in neuroblastoma patients after allogeneic SCT","authors":"Anne-Marie Lang, G. Jung, U. Seidel, F. Heubach, A. Rabsteyn, P. Schlegel, C. Seitz, E. M. Ribeiro, R. Handgretinger, P. Lang","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A140","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A140","url":null,"abstract":"Introduction: Whereas immunotherapy in high-risk/relapsed neuroblastoma with standard antibody formats targeting the tumor associated antigen GD2 is currently used after autologous/allogeneic stem cell transplantation (SCT), T-cells have rarely been addressed as effectors. Bispecific antibodies can bring T-cells into close proximity to tumor cells and activate them through the CD3 region, which ultimately leads to targeT-cell elimination. They lack the Fc-domain and cannot trigger ADCC or CDC as NK-cells. An example for bispecific antibodies is NG-CU (developed at the Department of Immunology, University of Tuebingen), which recognizes CD3 and GD2 as target antigens. The tumor antigen specific antibody domain, derived from a fab fragment of GD2 14.18 antibody and the T-cell recruiting antibody domain, derived from single chain variable fragment (scFv fragment) of UCHT-1 (CD3) are linked by modified CH2 domain. We investigated whether the bispecific antibody NG-CU might be an alternative to the therapeutic monoclonal antibody CH14.18, which mediates CDC and ADCC through NK-cells. Method: Different antibody concentrations and effector to target ratios (E:T) were evaluated using the xCELLigence RTCA system, PBMCs (healthy donors and patients after allogeneic SCT) and the neuroblastoma cell lines LS and LAN-1. CDC was evaluated using autologous serum.Results: NG-CU showed enhanced cytotoxicity compared to CH14.18. Median specific lysis (n=3) after 12/24/48 hrs (effectors: healthy PBMCs, E:T=5:1) with LS cells was: 55/73/77% (CH14.18, 1 µg/ml) vs. 70/100/100% (NG CU, 100ng/ml). P values with Mann-Whitney test: p=0,0244; p 48 hrs targets were completely eradicated by NG-CU, while targets treated with CH14.18 still proliferated.Using patient PBMCs, significant lysis with both constructs was detected dependent on percentages and total numbers of T- and NK-cells. In patients early after SCT, NK-cells represented the majority of effectors. Here, CH14.18 produced higher lysis, whereas later on, associated with increasing T-cell counts, NG-CU was more effective. With both antibody constructs complete eradication of neuroblastoma cells (LS+LAN-1) was detectable. Conclusion: NG-CU showed enhanced cytotoxicity compared to CH14.18 even in lower concentrations and E:T ratios. The bispecific design represents an alternative to classical CH14.18 based therapies. Dependent on the differential recovery of effector cells post-transplant, either NK-cell based or T-cell based antibody constructs might result in optimal antitumor activity. Citation Format: Anne-Marie Lang, Gundram Jung, Ursula Seidel, Florian Heubach, Armin Rabsteyn, Patrick Schlegel, Christian Seitz, Emmanuelle Moraes Ribeiro, Rupert Handgretinger, Peter Lang. A T-cell utilizing bispecific anti-CD3/GD2 construct mediates superior in vitro efficacy compared to CH14.18 mAb in neuroblastoma patients after allogeneic SCT [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immun","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"07 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86021307","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A161: Targeting gastrointestinal tumors with constant region engineered anti-glycan antibodies","authors":"M. Vankemmelbeke, Thomas Kirk, J. Chua, R. Mcintosh, L. Durrant","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A161","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A161","url":null,"abstract":"Post-translational modifications, for instance protein and lipid glycosylation, are attractive targets for therapeutic antibody (mAb) development. The altered tumor glyco-code drives oncogenic features such as the ability to proliferate, metastasize as well evade immune detection. Through immunizations with colorectal cancer cell membrane extracts we have generated a panel of anti-glycan mAbs with potential for application as cancer therapeutics. These mAbs selectively target Lewis or sialylated Lewis glycans on glycoproteins and/or glycolipids, binding to a large percentage of colorectal, pancreatic and gastric tumor tissues on tumor microarrays and induce a significant tumor volume reduction combined with a survival benefit in metastatic colorectal cancer xenograft models. Underlying these potent antitumor responses is the mAbs’ ability to induce direct tumor cell killing in the absence of complement or immune effector cells. In vitro, this direct cytotoxicity is characterized by mAb-induced cellular aggregation, pore formation, release of alarmins (ATP and high mobility group box 1 protein (HMGB1)), and maturation of immature dendritic cells, thereby constituting a form of inflammatory cell death (ICD). This mode of cell killing is linked to mAb cooperative binding on repeating antigen, a characteristic mostly associated with the murine mIgG3 isotype, thus human hIgG1 formats do not exhibit this form of direcT-cell killing. We have identified the residues required for the cooperative binding behaviour, through mAb constant region (CH2/CH3) screening, and have engineered improved (‘i’) hIgG1 variants, containing these selected residues. In silico immunogenicity prediction (IEDB) suggests limited immunogenicity, which requires further validation. Functional characterization of three improved anti-glycan hIgG1 mAbs in in vitro cell-based and Biacore assays demonstrates significant direct cancer cell killing, pore formation as well as increased functional glycan affinity. Classical immune effector functions (ADCC and CDC) were maintained or improved. Importantly, these improved hIgG1 variants now show significant tumor volume reduction in vivo in a mouse colorectal xenograft model.The multifaceted killing activity of our hIgG1 anti-glycan mAbs, has the potential to synergize with checkpoint blockade, thus holding tremendous therapeutic promise for the treatment of gastrointestinal tumors. Additionally, the activity of other therapeutic mAbs could be further enhanced with our Fc-engineering strategy for introducing cooperative binding. Citation Format: Mireille Vankemmelbeke, Thomas Kirk, Jia X. Chua, Richard McIntosh, Lindy G. Durrant. Targeting gastrointestinal tumors with constant region engineered anti-glycan antibodies [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"15 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"78667998","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A148: A hydrogel platform for the delivery of specialized pro-resolving mediators to treat chronic inflammatory disease","authors":"Padmini S. Pillai, Jamie Webster, R. Langer","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A148","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A148","url":null,"abstract":"The resolution of acute inflammation is an active process orchestrated by mediators that prevent further recruitment of inflammatory cells and promote clearance of microbes and dying cells. Chronic inflammation can arise when resolution is diminished due to a scarcity of these pro-resolving mediators. Current immunosuppressive drugs for the treatment of chronic inflammatory diseases such as IBD, arthritis, or psoriasis temporarily block inflammation without initiating resolution or tissue repair, leading to the recurrence of chronic inflammation. Additionally, systemic administration of these drugs leaves patients susceptible to tuberculosis, lymphoma, and opportunistic infections. Resolvins are a class of evolutionarily conserved lipid mediators that return tissue to homeostasis without immunosuppression by promoting phagocytosis of debris and microbes, and preventing proinflammatory cytokine production and cellular infiltration. To enhance dose sparing and stability, we have developed a tunable hydrogel platform to deliver resolvins to various sites of chronic inflammation. For oral delivery, this hydrogel is further encapsulated into a pH-sensitive microparticle formulated using anionic polymers, to target the large intestine. Using a mouse model of colitis, we demonstrate the ability of the hydrogel to resolve intestinal inflammation, initiate repair of the intestinal barrier, and regulate the gut microbiota. Our tunable hydrogel platform could have profound effects on the prevention of gastrointestinal cancers and chronic inflammatory disease. Citation Format: Padmini Sushila Pillai, Jamie Webster, Robert Langer. A hydrogel platform for the delivery of specialized pro-resolving mediators to treat chronic inflammatory disease [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A148.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"2 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"74433618","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A155: Antibody-dependent cancer cell phagocytosis in macrophages induces immune escape by upregulating PD-L1 and IDO","authors":"Shicheng Su, A. Su","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A155","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A155","url":null,"abstract":"Therapeutic antibodies can exert anticancer effects via antibody-dependenT-cellular cytotoxicity (ADCC). Also, they may trigger antibody-dependenT-cellular phagocytosis (ADCP) in tumor-associated macrophages (TAMs), but how ADCP influences TAM functions and antitumor immunity remains unclear. Here, we demonstrated that TAMs undergone ADCP of breast cancer cells and lymphoma cells mediated by trastuzumab and rituximab, respectively, suppressed the proliferation and cytotoxicity of NK cells and tumor-specific CD8+ T-cells against tumor cells by increasing PD-L1 and IDO. In vivo, inhibition of PD-L1 and IDO dramatically increases the infiltration of NK cells and CD8+ T-cells in Her2+ breast cancers and enhances the therapeutic effects of anti-Her2 antibody in both human Her2 knockin mice and humanized mice. Clinically, trastuzumab, but not chemotherapy alone, significantly increased the expression of PD-L1 and IDO in the TAMs of Her2+ breast cancer patients receiving neoadjuvant therapy. Furthermore, PD-L1+ IDO+ TAM infiltration was associated with poor trastuzumab response and reduced NK and CD8+ T-cells in tumors. Collectively, our findings unveil an unexpected role of ADCP mediated by therapeutic monoclonal antibodies in cancer immunosuppression, and suggest that antibody plus immune checkpoint blockade provide synergic therapeutic effects in cancer patients. Citation Format: Shicheng Su, An Su. Antibody-dependent cancer cell phagocytosis in macrophages induces immune escape by upregulating PD-L1 and IDO [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A155.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"42 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"82595921","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A164: Specifically targeting PD-L1 in the tumor-draining lymph node unmasks its spatiotemporal role in perturbing antitumor immunity and survival","authors":"Floris Dammeijer, Mandy van Gulijk, Melanie Lukkes, M. Nimwegen, R. Hendriks, T. V. Hall, H. Vroman, J. Aerts","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A164","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A164","url":null,"abstract":"Antagonistic antibodies to programmed cell death protein 1 (PD1) and its ligand PD-L1 have revolutionized the treatment of multiple metastasized cancers including melanoma and non-small cell lung cancer. Despite these advances, a majority of patients do not achieve durable responses to these therapies. Further insights into the mode of action and potential biomarkers predicting clinical response are therefore warranted. Although the focus has been on biomarker identification in the tumor or peripheral blood, the role of PD-L1 in the tumor draining lymph node (TDLN) has not yet been investigated. As the TDLN is crucial for orchestrating antitumor immune responses, we assessed the role of PD-L1 in the TDLN on survival and anti-tumor immunity. To assess the extent of PD-L1 expression in different tissues during tumor growth and inflammation, we measured PD-L1 levels on multiple cell subsets in the tumor, TDLN and non-TDLN on baseline and following injection of activated dendritic cells (DCs) by multicolor flow cytometry. We exploited the intraperitoneal (i.p.) localization of mesothelioma tumors by injecting a range of anti-PD-L1 antibody concentrations intrapleurally (i.pl). This allowed us to investigate the role of PD-L1 in the TDLN while leaving tumoral PD-L1 intact. When sole targeting of PD-L1 expressed in the TDLN was achieved, we injected advanced tumor-bearing mice with low-dose anti-PD-L1 i.pl and compared the effects on survival and antitumor immune responses to systemic administration of the antibody alone or in combination with DC-induced immune activation. Besides the well-documented expression of PD-L1 by cells in the TME, we detected significant levels of PD-L1 in the TDLN, mainly on macrophages and dendritic cells. Furthermore, surface PD-L1 expression doubled on these cells following adoptive transfer of inflammatory bone-marrow derived DCs. Injecting a near hundredfold lower dose of 2.5µg of anti-PD-L1 antibody i.pl. blocked PD-L1 in the TDLN and prevented translocation of the antibody to other sites. In the advanced disease setting, anti-PD-L1 monotherapy or adoptive DC-transfer only marginally improved survival (median survival of 24 days in untreated mice compared to 25 days for both monotherapies). A single i.pl. injection of low-dose antibody prior to DC-administration was as effective in prolonging survival as compared to repeated high-dose systemic injection of the antibody combined with adoptive DC-transfer (median survival of 35 and 35.5 days, respectively). When investigating the effects on antitumor immune responses, we found the increase in T-cell proliferation to be dependent on systemic anti-PD-L1 administration, whereas activation of T-cells indicated by CD69-positivity, was largely dependent on TDLN-localized PD-L1. Until now, dissecting the spatial roles of PD-L1 in immune regulation has proven difficult. By using a model allowing for separate dosing of PD-L1 blocking antibodies to different anatomic compartments,","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"40 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83960898","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract A162: AB928, a dual antagonist of the A2aR and A2bR adenosine receptors, relieves adenosine-mediated immune suppression","authors":"D. Direnzo, D. Piovesan, J. Tan, D. Miles, M. Leleti, Tim Park, F. Soriano, B. Handlos, J. Jeffrey, Ehesan U. Sharif, Brandon R. Rosen, U. Schindler, J. Powers, M. Walters","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A162","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A162","url":null,"abstract":"Introduction: Adenosine, generated through the hydrolysis of extracellular adenosine monophosphate (AMP) by the ecto-nucleotidase CD73, is an important mechanism for immunosuppression in cancer development. Adenosine’s suppressive effects on immune cells are driven primarily through 2 of the 4 adenosine receptors, A2aR and A2bR. We have previously shown that adenosine-mediated suppression of T-cells can be blocked by the dual A2aR/A2bR antagonist, AB928. Herein, we show that AB928 is capable of relieving adenosine-mediated immune suppression using human in vitro cell cultures, advanced gene expression studies, and mouse syngeneic tumor models. Methods: The ability of AB928 to inhibit adenosine-mediated suppression of dendritic cell function in vitro was assessed using human monocyte-derived dendritic cells (moDC). Briefly, moDC were generated from freshly isolated CD14+ monocytes and differentiated with IL-4/GM-CSF for 7 days +/- adenosine/EHNA +/- AB928. Cells were then taken for NanoString analysis or placed in a mixed lymphocyte reaction (MLR) with CD4+ T-cells. Mouse syngeneic tumor studies were conducted using C57BL/6 mice inoculated with mouse mammary tumor AT3-OVA or melanoma B16-F10 cells. Tumors were subsequently treated with doxorubicin, oxaliplatin, or α-PD-1 +/- AB928. Results: Quantitative immunohistochemistry and analysis of public gene expression databases identified individual human tumor types that express high levels of adenosine processing enzymes. Most notably, non-small cell lung, renal, triple-negative breast, ovarian, colorectal, and gastroesophageal cancers were found to have the most favorable expression profiles for interventions targeting the adenosine pathway. Additionally, a high degree of correlation was found between transcript and protein measurements for CD73 (r2 = 0.87), illustrating the robust and reproducible nature of these techniques. In human in vitro cell cultures, moDC differentiated in the presence of adenosine showed a decreased ability to stimulate IFN-γ secretion from allogenic CD4+ T-cells in a MLR. This suppression was significantly reversed by addition of AB928. Next, multiplexed gene expression profiling using NanoString identified a cassette of 39 genes (>2.0 fold change, p Citation Format: Daniel DiRenzo, Dana Piovesan, Joanne Tan, Dillon H. Miles, Manmohan R. Leleti, Timothy Park, Ferdie Soriano, Bryan Handlos, Jenna L. Jeffrey, Ehesan U. Sharif, Brandon R. Rosen, Ulrike Schindler, Jay P. Powers, Matthew J. Walters. AB928, a dual antagonist of the A2aR and A2bR adenosine receptors, relieves adenosine-mediated immune suppression [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A162.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"51 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"81004024","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstract PR08: Mechanisms of primary resistance to PD-1 checkpoint blockade","authors":"M. Krogsgaard, Duane Moogk, Kaitao Li, Zhou Yuan, I. Osman, J. Weber, Cheng Zhu","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-PR08","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-PR08","url":null,"abstract":"Although much clinical progress has been made in harnessing the immune system to recognize and target cancer, there is still a significant lack of an understanding of how tumors evade immune recognition and the mechanisms that drive tumor resistance to both T-cell and checkpoint blockade immunotherapy. Our objective is to understand how tumor-mediated signaling through inhibitory receptors, including PD-1, combines to affect the process of T-cell recognition of tumor antigen and activation signaling. This has the goal of understanding the basis of resistance to PD-1 blockade and potentially identifying new molecular targets to enable T-cells to overcome dysfunction mediated by multiple inhibitory receptors. Biomembrane Force Probe (BFP) measurements show that that the activities of TCR-proximal signaling components affect T-cell mechanosensing and sensitivity at the earliest stages of antigen recognition and are influenced by PD-1 and other inhibitory receptors via Shp-1/2 by targeting CD28 and Lck to directly suppress TCR-pMHC-CD8 binding. Phospho-proteomics and flow cytometry-based analysis of patient-derived T-cells from PD-1 responders and nonresponders identified additional mediators, signaling components and pathways associated with PD-1 checkpoint blockade resistance. Targeting these interactions and understanding the basis of resistance to PD-1 blockade would potentially allow identification of novel biomarkers of resistance or new molecular targets to enable T-cells to overcome dysfunction during PD-1 checkpoint blockade. Citation Format: Michelle Krogsgaard, Duane Moogk, Kaitao Li, Zhou Yuan, Iman Osman, Jeffrey S Weber, Cheng Zhu. Mechanisms of primary resistance to PD-1 checkpoint blockade [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr PR08.","PeriodicalId":18169,"journal":{"name":"Maintenance of Immune Balance: Effects of Targeted and Immune Therapies","volume":"10 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90774665","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}