Tackling the Tumor Microenvironment: Beyond T-cells最新文献

{"title":"Abstract A091: IL-33 activates antitumoral toxicity in eosinophils through stimulation of contact-dependent degranulation","authors":"F. Mattei, Carla Buccione, S. Andreone, F. Spadaro, A. Ninno, Jacopo Mancini, C. Zanetti, I. Parolini, F. Iosi, A. Tinari, V. Lucarini, A. Gerardino, Giovanna Ziccheddu, L. Businaro, C. Afferni, G. Schiavoni","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A091","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A091","url":null,"abstract":"The alarmin IL-33 plays pleiotropic roles in allergy, autoimmunity and inflammation through binding to its specific receptor ST2 expressed by most hematopoietic cells. Emerging evidences suggest an involvement of this cytokine also in cancer immunity, although its function remains ill-defined. Eosinophils (EOS) are a rare blood population playing critical roles in allergic inflammation and parasitic responses. We recently showed that EOS play an essential role in anti-tumor responses against melanoma growth and pulmonary metastasis mediated by IL-33 in vivo.In the present study we analyzed the mechanisms by which IL-33 mediates tumor infiltration and antitumoral activities of EOS. We show that IL-33 indirectly stimulates the recruitment of EOS inducing tumor-derived chemokines CCL24 and CCL5. Furthermore, IL-33 directly activates EOS inducing the expression of adhesion molecules, such as the integrin CD11b, resulting in efficient contact-dependent tumor cell killing. In co-culture experiments, IL-33 activated EOS tightly bond to tumor cells, forming increased numbers of conjugates, with respect to resting eosinophils. Confocal laser-scanning microscopy (CLSM) of eosinophil-tumor cell conjugates revealed polarization of the pore-forming eosinophilic cationic protein (ECP) and of CD11b on the cell synapses exclusively in IL-33-activated, but not resting, EOS. Furthermore, we show that IL-33 activated EOS release larger amounts of extracellular vesicles (EV) with respect to resting EOS. Transmission electron microscopy (TEM) revealed increased degranulation and EV release of IL-33-activated EOS following cell contact with target tumor cells. Our results advocate for an eosinophil-mediated tumoricidal function promoted by IL-33, thus opening perspectives for novel cancer immunotherapy strategies. Citation Format: Fabrizio Mattei, Carla Buccione, Sara Andreone, Francesca Spadaro, Adele De Ninno, Jacopo Mancini, Cristiana Zanetti, Isabella Parolini, Francesca Iosi, Antonella Tinari, Valeria Lucarini, Annamaria Gerardino, Giovanna Ziccheddu, Luca Businaro, Claudia Afferni, Giovanna Schiavoni. IL-33 activates antitumoral toxicity in eosinophils through stimulation of contact-dependent degranulation [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 A091.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"59 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"79766160","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 A074: Spatially resolved deep antigen profiling of single cells in FFPE tissue samples through CODEXTM","authors":"M. Gallina, Atri Choksi, N. Nikulina, Jaskirat Singh, G. Dakshinamoorthy, Joseph Kim, S. Mistry, J. Kennedy‐Darling","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A074","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A074","url":null,"abstract":"Spatially resolved, deep antigen profiling of tissue specimens is crucial for investigating the architecture and cell diversity in complex matrices, such as the tumor microenvironment (TME). The correlation of these two parameters with the progression of a variety of pathologies, ranging from cancer to autoimmune diseases, is still vastly unknown due to the lack of technologies that enable the detection of both high content and spatial resolution. Formalin-fixed, paraffin-embedded (FFPE) tissues constitute the ideal sample for these investigations as they retain tissue morphology at room temperature for long periods of time, which is convenient and cost-effective, and are available in vast specimen archives. Akoya Biosciences, Inc., is commercializing CODEXTM (CO-Detection by indEXing), a multiparameteric imaging platform that performs the concurrent detection and quantification of dozens of antigens with single cell resolution within a tissue specimen. The CODEX platform uses a DNA-based barcode library to label antibodies and iterative cycles of adding and removing cognate dye-labeled oligonucleotides to reveal the staining pattern for a subset of the target markers per cycle. The entire data acquisition process is fully automated using the CODEX instrument, which integrates with existing microscopes. To support high-multiplexing capabilities, more than 45 different antibody clones have been screened and validated for the detection of target epitopes in human FFPE secondary lymphoid tissues for both healthy and cancerous specimens using the CODEX platform. Proof-of-concept data have also been obtained for different cancer types, including melanoma and head and neck cancer in both normal and TMA formats. Analysis software has been developed to extract meaning from CODEX datasets and is included as part of the CODEX platform. The analysis pipeline includes image drift compensation, deconvolution and segmentation to measure integrated fluorescence intensity for each cell across tens of parameters. In this manner, infiltrating lymphocytes and other immune cells have been identified within the TME. With the high multiplexing capabilities, a variety of T-cells can be identified to detect the ratios of both immunosuppressive and immune-activating subtypes. Analysis of FFPE tissues using the CODEX platform demonstrates the unparalleled opportunities for simultaneously detecting tens of antigens with single-cell resolution within a tissue specimen. Citation Format: Maria Elena Gallina, Atri Choksi, Nadya Nikulina, Jaskirat Singh, Gajalakshmi Dakshinamoorthy, Joseph Kim, Sejal Mistry, Julia Kennedy-Darling. Spatially resolved deep antigen profiling of single cells in FFPE tissue samples through CODEXTM [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 A074.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"16 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77236536","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 A059: Tackling the tumor microenvironment with CD38 blockade to enhance cancer immunotherapy","authors":"Limo Chen, L. Diao, X. Yi, B. L. Rodriguez, Yanli Li, P. Villalobos, T. Cascone, Xi Liu, L. Tan, P. Lorenzi, Jared J. Fradette, D. Peng, F. Skoulidis, Youhong Fan, J. Rodriguez-Canales, V. Papadimitrakopoulou, E. Dmitrovsky, L. Byers, Jing Wang, I. Wistuba, Jim Heymach, D. Gibbons","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A059","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A059","url":null,"abstract":"The single agent or the combination of anti-PD-1, anti-PD-L1, and anti-CTLA-4 is an effective strategy that is being clinically explored to treat a variety of cancer types. Some patients display primary resistance to the treatment, while others relapse after treatment. Resistance is a major issue that needs to be addressed. Using multiple immunocompetent syngeneic and K-rasLA1/+p53R172HΔg/+ spontaneous animal models of lung cancer, we have explored the mechanisms of resistance to treatments by evaluating the molecular and cellular immune profiles of the tumor microenvironment. We observed that tumor-bearing mice treated with PD-1/PD-L1 blocking antibodies developed resistance through the up-regulation of CD38. We also observed this in the combination therapy of anti-PD-1 and anti-CTLA-4, suggesting that CD38 is a major mechanism of resistance to immune checkpoint inhibitors. In vitro and in vivo studies demonstrated that CD38 impacted CD8+ T-cell function via adenosine receptor signaling and dendritic cell-mediated B7 signaling. Antibody-mediated cell depletion assays were conducted to validate the mechanisms. To determine the applicability to patients, we analyzed 793 lung cancer patients’ specimens with immunohistochemistry staining and assessed biomarker relationships in multiple large independent patient databases (~1,900 tumors). Pathologic analysis revealed positive immunohistochemical staining for CD38 on tumor cells in 15-23% of cases, and bioinformatic analyses revealed a strong correlation between CD38 expression and the immune signature. Lastly, targeting CD38 abolished the treatment resistance by modulating the adenosine levels and thereby enhancing the effector immune cell infiltrates into the tumor microenvironment. Based on our study, CD38 blockade improves the efficacy of single-agent anti-PD-1/PD-L1, or with anti-CTLA-4 combination in lung cancer. CD38 could potentially serve as a novel biomarker of resistance for immune checkpoint inhibition. The data from this study provide a unique target, biomarker, and therapeutic strategy that can be translated into the clinical practice. Citation Format: Limo Chen, Lixia Diao, Xiaohui Yi, Bertha Leticia Rodriguez, Yanli Li, Pamela Villalobos, Tina Cascone, Xi Liu, Lin Tan, Philip Lorenzi, Jared Fradette, David Peng, Ferdinandos Skoulidis, Youhong Fan, Jaime Rodriguez-Canales, Vassiliki Papadimitrakopoulou, Ethan Dmitrovsky, Lauren A Byers, Jing Wang, Ignasio Wistuba, Jim Heymach, Don Gibbons. Tackling the tumor microenvironment with CD38 blockade to enhance cancer immunotherapy [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 A059.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"5 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"86493192","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 A061: STING, an immune biomarker for colorectal cancer","authors":"H. Chon, Chan Kim","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A061","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A061","url":null,"abstract":"Background: STING is an innate immune sensor for cytosolic DNA. The activation of STING signaling is indispensable in Type I interferon response and the evocation of anticancer immune response by CD8+ T-cells. The aim of this study is to characterize intratumoral STING expression pattern and its clinical implication in colorectal cancer (CRC). Methods: We analyzed STING and CD8 expression in 225 CRC patients who underwent surgical resection. Clinicopathologic variables and survival outcomes were analyzed according to STING expression level. Results: Distinct STING expression was observed in tumor specimens of CRCs. Patients with higher STING expression seemed to have early stage cancer (P = 0.001) with increased intratumoral CD8+ T-cell infiltration (P Citation Format: Hongjae Chon, Chan Kim. STING, an immune biomarker for colorectal 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 A061.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"87424577","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 A051: Melanoma displays an evolutionarily conserved resistance to upregulation of prophagocytic signals and to CD47 blockade","authors":"K. L. Anderson, K. Snyder, D. Ito, Debra C. Lins, L. Mills, K. Weiskopf, N. G. Ring, A. Ring, Y. Shimizu, M. Mescher, I. Weissman, J. Modiano","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A051","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A051","url":null,"abstract":"Therapeutic activation of macrophage phagocytosis has the ability to restrain tumor growth through phagocytic clearance of tumor cells and activation of the adaptive immune response. Our objective for this study was to evaluate the effects of modulating pro- and anti-phagocytic pathways in malignant melanoma. In order to identify evolutionarily conserved mechanisms of resistance that may be important for melanoma cell survival, we utilized a multispecies approach and examined the phagocytosis of human, mouse, and dog melanoma cells in vitro. We blocked the interaction between CD47 on tumor cells and SIRPα on macrophages using anti-CD47 monoclonal antibodies, SIRPα mimotopes, and a soluble fusion protein of the SIRPα extracellular domain. In all cases, we confirmed that these reagents blocked >85% of the maximum binding of CD47 to recombinant SIRPα. We observed that melanoma cells from all three species displayed unexpected resistance to phagocytosis that could not be fully mitigated by blockade of the “don’t eat me” signal CD47 or by chemotherapeutic enhancement of known “eat me” signals. In vitro, CD47 blockade minimally enhanced phagocytosis of melanoma cells, and loss of CD47 expression did not increase sensitivity to phagocytosis. In vivo, CD47 blockade enhanced the proliferation of tumor-specific CD8+ T-cells, but this response failed to inhibit tumor growth. Resistance to phagocytosis was not mediated by soluble factors, as phagocytosis of melanoma cells was not enhanced by inhibition of secretory pathways, and phagocytosis of sensitive lymphoma tumor cells was not impaired in the presence of melanoma cells or melanoma culture supernatants. siRNA-mediated knockdown of 47 prospective “don’t eat me” signals similarly did not enhance melanoma cell phagocytosis. Interestingly, in lymphoma cells, CD47 knockout alone did not enhance phagocytosis, suggesting that at least part of the pro-phagocytic effect of CD47 blockade is due to Fc receptor engagement. Restoration of CD47 expression in lymphoma cells re-established sensitivity to CD47 blockade. We conclude that melanoma cells possess an evolutionarily conserved resistance to macrophage phagocytosis. Further investigation will be needed to define and overcome the mechanisms that mediate melanoma cell resistance to innate immunity. Citation Format: Katie L. Anderson, Kristin M. Snyder, Daisuke Ito, Debra C. Lins, Lauren J. Mills, Kipp Weiskopf, Nan G. Ring, Aaron M Ring, Yoji Shimizu, Matthew F. Mescher, Irving L. Weissman, Jaime F. Modiano. Melanoma displays an evolutionarily conserved resistance to upregulation of prophagocytic signals and to CD47 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 A051.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"85450414","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 A067: Cognate interaction with CD4+ T-cells instructs M2-like macrophages to acquire M1-like phenotype","authors":"D. Eisel, W. Osen, K. Das, Franziska Hoerhold, R. König, S. Eichmüller","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A067","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A067","url":null,"abstract":"Current tumor immunotherapy approaches are based on application of checkpoint inhibitors, such as monoclonal antibodies against PD-1/PD-L1 and CTLA-4 or adoptive T-cell therapy using ex vivo expanded TILs or genetically modified autologous T-cells expressing recombinant T-cell receptors (TCRs) or chimeric antigen receptors (CARs). However, the success of these therapeutic strategies is often limited by various inhibitory immune cell types accumulating in the tumor. In particular, tumor-associated macrophages (TAMs) contribute to the immune suppressive tumor micro-milieu, since so-called M2-like macrophages suppress the function of T effector cells and promote the differentiation of regulatory T-cells (Treg) through secretion of inhibitory cytokines such as TGF-β and IL10. Tumor antigen-specific CD4+ T effector cells, on the other hand, can essentially sustain antitumoral immune responses as shown for various tumor entities. In fact, using peritoneal exudate cells (PECs) as source for macrophages we demonstrate that MHC II restricted interaction between ovalbumin (OVA) specific CD4+ T-cells and M2-like macrophages drives M1 polarization. This was confirmed by detailed gene and protein expression analyses as well as functional assays testing phagocytic and pinocytic activities of repolarized macrophages. Moreover, in a set of preclinical experiments, adoptive transfer of CD4+, OVA-specific OT-II cells into C57BL/6 mice bearing OVA expressing IAb-/- tumors resulted in increased intratumoral number of M1-like TAMs as determined by gene expression analysis and flow cytometry. Furthermore, we observed a significant survival benefit of mice treated with a combination of OVA-specific CD4+ (OT-II) and CD8+ (OT-I) cells after transplantation of B16F10-Ova tumors and a complete response in some mice that rejected the tumor cells also upon a later re-challenge. While the antitumoral effect of this adoptive transfer experiment has been already described, we now offer a possible explanation for the supportive effect of specific CD4+ cells on co-transferred CD8+ cells. Taken together, the instructive impact of CD4+ T-cells on M2-like macrophages described in this presentation points towards a so far underestimated function of the CD4+ T-cell / TAM axis in reconditioning the immunosuppressive tumor micro-milieu. Citation Format: David Eisel, Wolfram Osen, Krishna Das, Franziska Marie-Claire Hoerhold, Rainer Konig, Stefan B. Eichmuller. Cognate interaction with CD4+ T-cells instructs M2-like macrophages to acquire M1-like phenotype [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 A067.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"3 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"89703261","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 A085: High infiltration of NK cells expressing elevated LAG-3 in a subgroup of renal cell carcinoma patients","authors":"M. Lee, P. Järvinen, H. Nísen, O. Brück, Mette Ilander, S. Mustjoki, K. Anna","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A085","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A085","url":null,"abstract":"Background: Renal cell carcinoma (RCC) is considered one of the most immunogenic cancers with the highest number of indel mutations, frequent infiltration of T-cells, presence of many antigen-specific T-cell clones, and high immuno-oncologic (IO) sensitivity. Since less is known about natural killer (NK) cells in RCC, our primary aim was to investigate the intratumoral phenotype of NK cells as well as further assess the overall immune landscape of the tumors, peripheral blood (PB) and adjacent healthy kidney tissue, which may be critical for patient prognosis and predictions to targeted immunotherapies. Methods: We used multi-parameter flow cytometry together with a comprehensive immunostaining panel containing a total of 56 fundamental markers to cancer immunology to immunophenotype the tumor, adjacent healthy kidney tissue and presurgical peripheral blood (PB) samples from 31 RCC patients who underwent partial or radical nephrectomies. To study the intratumoral T-cell clonalities, T-cell receptor beta (TCRβ) deep sequencing was carried out with eight tumors. Results: Using hierarchical clustering (Spearman correlation distance and Ward linkage) and correlation analyses (Spearman correlation), we discovered that our patient cohort clustered into two distinct subgroups defined by a high (NKhigh, n=11; mean 29.7%) and low (NKlow, n=20; mean 9.4%) percentage of NK cells among the intratumoral lymphocyte population. Accordingly, the NKhigh subgroup had a lower percentage of T-cells (mean 36.9%) than the NKlow group (mean 65.7%), and overall, a significant negative correlation between T and NK cells was discovered. Our TCRβ sequencing results revealed a positive correlation between T-cell clonality and the intratumoral T-cell percentage, whereas the higher proportion of tumor NK cells associated with low T-cell clonality, possibly due to a polyclonal T-cell population. When we compared the expressions of the most clinically relevant IO markers (LAG-3 and PD-1) on the NK cells, LAG-3 was more expressed in the NKhigh group than in the NKlow group (21.3% vs 9.8%; p=0.08). In contrast, no differences were observed with PD-1. Clinical parameters such as tumor grade (Fuhrman), weight, size (diameter), the presence of necrosis, gender, or age of the patients did not differ between the two subgroups. To examine the overall immune landscape of RCC, we compared the cells from the tumor, PB, and healthy kidney tissue of seven patients. Our results showed that tumors have more NK cells compared to their corresponding T-cell-rich PB and healthy tissue counterparts, supporting our findings that some tumors accumulate NK cells. Compared to the adjacent healthy tissue, PD-1 and LAG-3 expressions were higher in the intratumoral CD8 cells. The expressions of PD-1 and LAG-3 on PB CD8+ T-cells or NK cells did not correlate with their intratumoral counterparts, whereas a positive correlation was found between the PB and tumor CD4+ T-cells for both LAG-3 and PD-1. Conclus","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","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":"89846966","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 A073: CODEXTM: A novel platform for spatially resolved deep antigen profiling of single cells in tissue samples","authors":"M. Gallina, Atri Choksi, N. Nikulina, Jaskirat Singh, G. Dakshinamoorthy, Joseph Kim, S. Mistry, J. Kennedy‐Darling","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A073","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A073","url":null,"abstract":"The tumor microenvironment (TME) comprises a multitude of cell types that collectively create an immunosuppressive environment, enabling tumor growth. Understanding the spatial organization of these tissues requires a technology that can identify the presence of multiple markers and correlate it to their specific spatial location within the same tissue. Characteristics of the TME including infiltrating immune cells, angiogenesis and the presence of other nonmalignant cells influence important phenomena like drug delivery, treatment effectiveness and, ultimately, clinical outcomes. Current methodologies for analyzing the spatial dimension of tissues, e.g., traditional immunofluorescence (IF) and immunohistochemistry (IHC), are limited to measuring a few parameters simultaneously, thereby restricting the number of phenotypes that can be identified. Conversely, single-cell technologies like mass cytometry and next-generation sequencing-based tools provide multiplexing capabilities, but at the expense of the associated spatial information. Akoya Biosciences, Inc., is commercializing CODEXTM (CO-Detection by indEXing), a multiparameteric imaging platform that allows the simultaneous detection and quantification of dozens of target epitopes in single cells within a single tissue section. This innovative platform is an end-to-end solution that comprises three components: 1) the CODEX fluidics instrument, 2) a suite of specialized CODEX reagents, and 3) an analysis pipeline. The CODEX workflow involves a barcoding system such that each antibody moiety is conjugated to a proprietary tag. Panels of CODEX antibodies are used to stain tissue specimens en masse in a single step. Staining data for sets of antibodies are revealed across iterative cycles using corresponding dye-labeled barcodes. The CODEX fluidics instrument integrates with existing microscope units for a fully automated data collection process. CODEX data are processed to achieve noise reduction and analyzed at the single-cell level through a segmentation algorithm based on nuclear staining.More than 80 antibodies have been validated on the CODEX platform for analysis of human and mouse fresh-frozen and FFPE tissues. Preliminary studies on fresh-frozen tissues with developed antibodies demonstrate an unprecedented capability for revealing the spatial correlation between a variety of target epitopes. These results demonstrate the enormous potential of the CODEX technology to identify spatial correlations in the TME through highly multiplexed detection with single-cell resolution. Citation Format: Maria Elena Gallina, Atri Choksi, Nadya Nikulina, Jaskirat Singh, Gajalakshmi Dakshinamoorthy, Joseph Kim, Sejal Mistry, Julia Kennedy-Darling. CODEXTM: A novel platform for spatially resolved deep antigen profiling of single cells in tissue samples [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","volume":"5 3","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91482794","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 A077: Microenvironmental factors shape resistance patterns to immune checkpoint blockade","authors":"Sheng Gu, Xihao Hu, Xiaoqing Wang, Ziyi Li, Nicole Traugh, Xia Bu, Xiaofang Xing, G. Freeman, Myles A. Brown, Xiaole Shirley Liu","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A077","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A077","url":null,"abstract":"Immune checkpoint blockade (ICB) therapy has dramatically improved the prognosis of several types of cancer. However, only a small proportion of patients respond. Although multiple biomarkers/mechanisms of resistance to ICB have been identified, it remains elusive to what extent the cellular/molecular mechanisms contribute to the heterogeneity of ICB response. To this end, we applied clonal barcoding for lineage tracing of cancer cells following control IgG or ICB treatment in a transplantation mouse model. We identified significant clonal heterogeneity within cancer cells in response to ICB, suggestive of a minority of pre-existing ICB-resistant clones prior to treatment. Furthermore, counter-intuitively, ICB-responding tumors harbored a higher proportion of ICB-resistant clones than nonresponding tumors post-treatment, indicating that the tumor microenvironment might dictate the initial response to ICB. Integrated gene expression and immune repertoire analyses of the tumor microenvironment identified more T-cell and B cell infiltration in the ICB responders, and found that BCR class switch is associated with ICB response. Our study established a system to assess the contributions of cancer cell-intrinsic and microenvironmental factors in response to ICB treatment, and identified B cell infiltration and repertoire constitution as a novel biomarker for ICB response. Citation Format: Shengquing Stan Gu, Xihao Sherlock Hu, Xiaoqing Shawn Wang, Ziyi Li, Nicole Traugh, Xia Bu, Xiaofang Xing, Gordon Freeman, Myles Brown, Xiaole Shirley Liu. Microenvironmental factors shape resistance patterns to immune 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 A077.","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","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":"90420433","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 A095: Discovering the immune profiles of a novel antifolate receptor alpha IgE antibody associated with monocyte-mediated antitumor functions","authors":"Mano Nakamura, H. J. Bax, J. Spicer, K. Lacy, S. Tsoka, D. Josephs, S. Karagiannis","doi":"10.1158/2326-6074.CRICIMTEATIAACR18-A095","DOIUrl":"https://doi.org/10.1158/2326-6074.CRICIMTEATIAACR18-A095","url":null,"abstract":"Monoclonal antibodies are an established modality for the clinical management of many cancers. To date, all antibodies approved for cancer treatment are of the IgG class, and many are targeted towards non-solid tumors. We hypothesized that antibodies of the IgE class may be able to recruit and activate immune cells against tissue resident tumors based on very high affinity of IgE for cognate Fcϵ Receptors on potent effector cells such as masT-cells, monocytes and macrophages. A novel IgE-based antibody against folate receptor alpha (FRα) for ovarian carcinoma was developed and demonstrated to engender superior anti-tumor effector functions compared with those triggered by its IgG counterpart in different in vivo models of cancer. We have now translated this concept to a first-in-man clinical trial. We have previously reported that monocytes and macrophages are important IgE effector cells against tumors. However, the underlying mechanisms of how IgE exerts antitumor immunity by engaging these effector cells are insufficiently characterized. Here, we sought to investigate the immune profile signatures triggered by the engagement of tumor antigen-specific IgE with Fc-receptors on the surface of monocytes, a key immune effector cell recruited by IgE antibodies. In order to investigate the above aim, the following methods were utilized: Antibody dependenT-cell-mediated cytotoxicity (ADCC) and phagocytosis (ADCP) were measured by flow cytometry. Cell stimulation studies with cytokines or by cross-linking of IgE monoclonal antibodies on the surface of monocytes were conducted. Expression and secretion of immune mediators were assessed using quantitative-PCR and ELISA. Toxicity of cytokines towards immune and tumor cells were investigated by cell viability (MTS) assays. As a result, tumor antigen-specific IgE-mediated tumor cell cytotoxicity mediated by human monocytes correlated with increased levels of TNFα, MCP-1 and IL-10. Cross-linking of IgE, but not IgG, on the monocyte surface, triggered up-regulation of the proinflammatory mediator TNFα. TNFα stimulation of monocytes and of tumor cells triggered stimulation of MCP-1 by both cells. Neither cross-linking of IgE or IgG antibodies on tumor cells or monocytes, nor stimulation of these cells with TNFα or MCP-1, was sufficient to upregulate IL-10. However, IL-10 expression was induced by monocytic cells when stimulated by TNFα and MCP-1 in combination, and by stimulation with IL-10 in an autocrine manner. None of these stimulation conditions was sufficient to upregulate IL-10 in cancer cell lines of different origins. None of the three cytokines had direct cytotoxic effects towards immune or tumor cells. However, blockade of TNFα receptor on monocytes reduced ADCC and increased levels of the chemoattractant MCP-1 recruited monocytes in chemotaxis assays in vitro and macrophages in tumor lesions in a rat model of cancer treated with anti-tumor IgE. These findings suggest that IgE-dependent monocyte-med","PeriodicalId":22141,"journal":{"name":"Tackling the Tumor Microenvironment: Beyond T-cells","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":"87195424","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}