bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B014: BGB324, a selective small molecule inhibitor of the receptor tyrosine kinase AXL, enhances immune checkpoint inhibitor efficacy
Authors: Gausdal Gro , Davidsen Kjersti , Wnuk-Lipinska Katarzyna , Wiertel Kathleen , Hellesøy Monica , Blø Magnus , Ahmed Lavina , Hodneland Linn , Kiprijanov Sergej , Brekken Rolf A , Lorens James B
Published: May 2016

Signaling via the AXL receptor tyrosine kinase is a key suppressor of anti-tumor innate immune response. AXL is expressed on several cells associated with the suppressive tumor immune microenvironment including natural killer cells, dendritic cells and tumor-associated macrophages. AXL is also an important regulator of tumor plasticity related to epithelial-to-mesenchymal transition (EMT) that contributes to anti-tumor immune evasion. Hence AXL signaling contributes uniquely to tumor intrinsic and microenvironmental immune suppression in tumors. We therefore evaluated whether blocking AXL signaling with BGB324, a selective clinical-stage small molecule Axl kinase inhibitor, enhances the effect of immune checkpoint blockade in syngeneic cancer mouse models that display limited immunogenicity.

We measured the effect of BGB324 in combination with anti-CTLA-4 and anti-PD-1 in the mammary adenocarcinoma 4T1/Balb/C syngeneic mouse model. BGB324 (50 mg/kg bid) significantly enhanced responsiveness to anti-CTLA-4/anti-PD-1 treatment (10 mg/kg of each, 4 doses) in Balb/C mice bearing 4T1 tumors. The combination of BGB324 + anti-CTLA-4/anti-PD-1 resulted in complete tumor clearance in 46.1 % of mice versus complete tumor clearance in 11.7 % of the mice treated with anti-CTLA-4/anti-PD-1 (p = 0.0087). BGB324 + anti-CTLA-4/anti-PD-1 treated tumors displayed enhanced CD8+ T cell tumor infiltration. Combination of BGB324 with immune checkpoint inhibitors is being evaluated in additional models, and detailed interrogation of AXL-dependent immune effector cell activity in tumors is in progress.

In conclusion, AXL inhibition represents a unique opportunity to target anti-tumor immune suppressive mechanisms and supports clinical translation of BGB324 in combination with cancer immunotherapy in human cancer.

Citation Format: Gro Gausdal, Kjersti Davidsen, Katarzyna Wnuk-Lipinska, Kathleen Wiertel, Monica Hellesøy, Magnus Blø, Lavina Ahmed, Linn Hodneland, Sergej Kiprijanov, Rolf A Brekken, James B Lorens. BGB324, a selective small molecule inhibitor of the receptor tyrosine kinase AXL, enhances immune checkpoint inhibitor efficacy. [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 B014.
Abstract B016: Allogenic CAR T-cells for adoptive immunotherapy
Authors: Galetto Roman , Lebuhotel Celine , Chion-Sotinel Isabelle , Francon Patricia , Gouble Agnes , Smith Julianne
Published: May 2016

Chimeric antigen receptor (CAR)-redirected T-cells have given rise to long-term durable remissions and remarkable objective response rates in patients with refractory leukemia, raising hopes that a wider application of CAR technology may lead to a new paradigm in cancer treatment. A limitation of the current autologous approach is that CAR T-cells must be manufactured on a “per patient basis”. We have developed a standardized platform for manufacturing T-cells from third-party healthy donors to generate allogeneic “off-the-shelf” engineered CAR+ T-cell–based frozen products. This platform utilizes Transcription Activator-Like Effector Nuclease (TALEN) gene editing technology to inactivate the TCRα constant (TRAC) gene, eliminating the potential for T-cells bearing alloreactive TCR's to mediate Graft versus Host Disease (GvHD). We have previously demonstrated that editing of the TRAC gene can be achieved at high frequencies, obtaining up to 80% of TCRβα negative cells. This allows us to efficiently produce TCR-deficient T-cells that have been shown to no longer mediate alloreactivity in a xeno-GvHD mouse model. We have adapted this allogeneic platform to the production of T cells targeting CD123, the transmembrane alpha chain of the interleukin-3 receptor, which is expressed in tumor cells of patients with Acute Myeloid Leukemia as well as CS1 a glycoprotein highly expressed on tumor cells of patients with multiple myeloma. We will present both in vitro and in vivo data demonstrating specific anti-tumor activity of engineered CAR T cells against cell lines expressing the targeted antigen.

Citation Format: Roman Galetto, Celine Lebuhotel, Isabelle Chion-Sotinel, Patricia Francon, Agnes Gouble, Julianne Smith. Allogenic CAR T-cells for adoptive immunotherapy. [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 B016.
Abstract B028: Transcriptional and epigenetic signatures of ex vivo propagated three distinct TCR Vδ1, TCR Vδ2 and TCR Vδ3 cell subtypes with broad specificity for malignancies
Authors: Maiti Sourindra , Deniger Drew , Dai Jianliang , Cooper Laurence
Published: May 2016

Functionally pleiotropic human γδ T cells are a unique set of immune cells that exhibit qualities of both innate and adaptive immune responses. In contrast to αβ T cells, γδ T cells recognize their ligands independent of MHC and are infrequent (1%–5% of T cells) in peripheral blood. Studies have shown that γδ T cells exhibit endogenous cytotoxicity towards tumor cells by directly recognizing a diverse array of Tumor Associated Antigen (TAA) and have the ability to present TAA to elicit an antitumor response. This broad recognition of antitumor activity is achieved because these cells express a diverse TCR γδ repertoire (combination of Vδ1, Vδ2, or Vδ3 with one of fourteen Vγ chains). Because of their characteristics γδ T cells are attractive targets for clinical manipulation and tumor immunotherapy. To obtain clinically significant numbers of cells for immunotherapy current ex vivo γδ T cells expansion protocol uses Zoledronic acid, an aminobisphosphonate that results only in selective propagation of Vγ9Vδ2. These cells however, showed clinical response against both solid and hematologic tumors. We have previously shown a novel ex vivo expansion protocol using artificial antigen presenting cells (aAPC), IL2 and IL21 assist aAPC in activating and expanding polyclonal γδ T-cells (Vδ1, Vδ2 and Vδ1neg Vδ2neg T cell subtypes) to clinically significant numbers. These expanded cell subsets secrete proinflammatory cytokines, lysed a broad range of malignancies and improved survival in ovarian cancer xenograft model. To better define the role of the subsets and their application in immunotherapy here we present high-resolution transcriptome, microRNA and secreted cytokine-chemokine signatures of ex vivo expanded subsets. Our data provide an atlas of transcriptional, epigenetic and secretome of three defined subsets, and subset-specific signatures. The data presented here will have a broader impact in understanding the immunobiology of γδ T cell subsets and facilitate improving the human application of these cells in cancer immunotherapy.

Citation Format: Sourindra Maiti, Drew Deniger, Jianliang Dai, Laurence Cooper. Transcriptional and epigenetic signatures of ex vivo propagated three distinct TCR Vδ1, TCR Vδ2 and TCR Vδ3 cell subtypes with broad specificity for malignancies. [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 B028.
Abstract B031: Heat-inactivated modified vaccinia virus ankara induces type I IFN and antitumor immunity via the cytosolic DNA-sensing pathway
Authors: Dai Peihong , Wang Weiyi , Serna-Tamayo Cristian , Zamarin Dimitriy , Shuman Stewart , Merghoub Taha , Wolchok Jedd D. , Deng Liang
Published: May 2016

Type I interferon (IFN), well known for its antiviral activity, is a critical component of cancer immune surveillance through its direct actions on cancer cells as well as on tumor microenvironment. Advanced melanoma remains a therapeutic challenge despite recent progress in targeted therapy and immunotherapy. Novel approaches are needed to alter the tumor immune suppressive microenvironment and to facilitate the recognition of tumor antigens that leads to antitumor immunity. Poxviruses are cytosolic DNA viruses that have been investigated as oncolytic and immunotherapeutic agents. We recently reported that the highly attenuated modified vaccinia virus Ankara (MVA), a safe vaccine for smallpox, triggers type I IFN production in conventional dendritic cells (cDCs) via the cytosolic DNA sensor cGAS and its adaptor STING, and that it requires transcription factors IRF3 and IRF7. Here we show that infection of cDCs with heat-inactivated MVA leads to higher levels of IFN induction than MVA. This induction is also mediated by the cytosolic DNA-sensing pathway cGAS/STING/IRF3/IRF7. In addition, we found that intratumoral injection of Heat-MVA caused tumor eradication in a murine B16 melanoma model as well as the generation of adaptive anti-tumor immunity. Furthermore, Heat-MVA-induced anti-tumor therapy is less effective in STING, IRF7, or Batf3-deficient mice than in wild-type mice, indicating that both the innate immune-sensing pathway and CD8α+DCs are essential for Heat-MVA-based immunotherapy. Lastly, the combination of intratumoral delivery of Heat-MVA with systemic delivery of anti-CTLA-4, PD-1 or PD-L1 antibodies achieved enhanced efficacy in tumor eradication and survival than Heat-MVA alone. Our results have strong implications for the development of poxvirus-based cancer immunotherapeutics as well novel strategies to overcome resistance to immune checkpoint blockade therapy.

Citation Format: Peihong Dai, Weiyi Wang, Cristian Serna-Tamayo, Dimitriy Zamarin, Stewart Shuman, Taha Merghoub, Jedd D. Wolchok, Liang Deng. Heat-inactivated modified vaccinia virus ankara induces type I IFN and antitumor immunity via the cytosolic DNA-sensing pathway. [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 B031.
Abstract B036: Monoclonal antibodies against GARP/TGF-β1 complexes inhibit the immunosuppressive activity of human regulatory T cells in vivo
Authors: Liénart Stéphanie , Cuende Julia , Saunders Michael , Woning Bas van der , Haard Hans De , Coulie Pierre G. , Lucas Sophie
Published: May 2016

Regulatory T lymphocytes (Tregs) are essential to prevent auto-immunity, but excessive Treg function contributes to cancer progression by inhibiting anti-tumor immune responses. Tregs exert contact-dependent inhibition of immune cells through the production of active TGF-β1. On the Treg cell surface, TGF-β1 is in an inactive form bound to membrane protein GARP and then activated by an unknown mechanism. Here, we demonstrate that GARP is involved in this activation mechanism. Two anti-GARP monoclonal antibodies were generated that block the production of active TGF-β1 by human Tregs. These antibodies recognize a conformational epitope that requires amino-acids GARP137-139 within GARP/TGF-β1 complexes. A variety of antibodies recognizing other GARP epitopes did not block active TGF-β1 production by Tregs. In a model of xenogeneic graft-versus-host disease in NSG mice, the blocking antibodies inhibited the immunosuppressive activity of human Tregs. These antibodies may serve as therapeutic tools to boost immune responses to infection or cancer, via a mechanism of action distinct from that of currently available immuno-modulatory antibodies. Used alone or in combination with tumor vaccines or antibodies targeting the CTLA4 or PD1/PD-L1 pathways, blocking anti-GARP antibodies may improve the efficiency of cancer immunotherapy.

Citation Format: Stéphanie Liénart, Julia Cuende, Michael Saunders, Bas van der Woning, Hans De Haard, Pierre G. Coulie, Sophie Lucas. Monoclonal antibodies against GARP/TGF-β1 complexes inhibit the immunosuppressive activity of human regulatory T cells in vivo. [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 B036.
Abstract B046: Mechanisms of T lymphocyte activation revealed by super-resolution microscopy
Authors: Traver Maria , Campanello Leonard , Paul Suman , Losert Wolfgang , Shroff Hari , Schaefer Brian
Published: May 2016

Tight regulatory control of lymphocyte activation is necessary to avoid the deleterious consequences of an uncontrolled immune response. A complicated web of up-regulatory and down-regulatory processes governs a key activation pathway in lymphocytes, the antigen-receptor-to-NF-κB pathway; however, the complex interplay between positive and negative regulation of this pathway remains poorly understood. We have utilized cutting-edge super-resolution imaging technologies to examine the spatial and temporal organization of the POLKADOTS signalosome, which is inducibly formed in response to antigen stimulation of T lymphocytes. Our preliminary results suggest that this filamentous signalosome is divided into two distinct subregions: an activation domain which recruits signal transduction elements, and a downregulation domain which promotes macroautophagic degradation of the signalosome. Fixed and live cell imaging of the activation process indicates that POLKADOTS filaments converge on the microtubule-organizing center as the core filament protein Bcl10 is progressively degraded by autophagosomes. Furthermore, following Bcl10 degradation, the POLKADOTS components Malt1 and phospho-IKK are transferred to a peri-nuclear aggresomal structure. Surprisingly, our data suggest these aggresomal accumulations continue to promote NF-κB nuclear localization and T cell activation. Together, these results provide new insights into the complex regulatory processes which govern T lymphocyte activation, and for the first time suggest a signaling function for aggresomes.

Citation Format: Maria Traver, Leonard Campanello, Suman Paul, Wolfgang Losert, Hari Shroff, Brian Schaefer. Mechanisms of T lymphocyte activation revealed by super-resolution microscopy. [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 B046.
Abstract B048: The MHC class II transactivator CIITA inhibits the persistent activation of NF-kB by Human T cell Lymphotropic Virus type-1 Tax-1 oncoprotein
Authors: Forlani Greta , Abdallah Rawan , Accolla Roberto S. , Tosi Giovanna
Published: May 2016

Human T cell Lymphotropic Virus type-1 (HTLV-1) Tax-1 protein, a key mediator of HTLV-1-induced T-cell transformation, deregulates diverse cell-signalling pathways. Among them, the NF-kB pathway is constitutively activated by Tax-1, which binds to NF-kB family members and activates the IkB-kinase (IKK). Upon phosphorylation-dependent degradation of IkB, NF-kB migrates into the nucleus and mediates Tax-1-stimulated gene expression. We show that CIITA, the transcriptional regulator of Major Histocompatibility Complex Class II genes, inhibits the activation of the canonical NF-kB pathway by Tax-1 and map the region mediating this effect. CIITA affects the subcellular localization of Tax-1, which is mostly retained in the cytoplasm, and this correlates with an impaired migration of RelA into the nucleus. Cytoplasmic and nuclear CIITA mutants reveal that CIITA exploits different strategies to suppress Tax-1-mediated NF-kB activation in both subcellular compartments. CIITA interacts with Tax-1 without preventing Tax-1 binding to both IKKg and RelA. Nevertheless, CIITA affects Tax-1-induced IKK activity causing the retention of the inactive p50/RelA/IkB complex in the cytoplasm. Nuclear CIITA associates with Tax-1/RelA in nuclear bodies blocking the activation of NF-kB-responsive genes by Tax-1. Thus, CIITA inhibits critical cytoplasmic and nuclear steps of Tax-1-mediated NF-kB activation. These results, together with our previous finding that CIITA acts as a restriction factor inhibiting Tax-1-promoted HTLV-1 gene expression and replication, indicate that CIITA is a versatile molecule that might also counteract Tax-1 transforming activity. Unveiling the molecular basis of CIITA-mediated inhibition of Tax-1 functions may be important in defining new strategies to control HTLV-1 spreading and oncogenic potential.

Citation Format: Greta Forlani, Rawan Abdallah, Roberto S. Accolla, Giovanna Tosi. The MHC class II transactivator CIITA inhibits the persistent activation of NF-kB by Human T cell Lymphotropic Virus type-1 Tax-1 oncoprotein. [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 B048.
Abstract B059: Slow maturation of early phagosomes promotes a TAP independent antigen cross-presentation pathway
Authors: Sengupta Debrup , Cresswell Peter
Published: May 2016

Early phagocytic compartments play a role in protein sorting, signaling and neutralizing pathogens. In addition, certain subsets of dendritic cells (DCs) load peptides derived from exogenous antigens on to MHC class-I molecules in early phagosomes. This process plays a key role in antigen cross-presentation, a process particularly important in the activation of naïve CD8+ T-cells when it is known as cross-priming and is essential for mounting an immune response against tumor and viral antigens. The functional properties of early phagosomes that facilitate cross-presentation remain unclear. Here we show that the expression of Rab GTPase mutants (Rab5CA, Rab7DN and Rab22CA) that impair maturation of phagosomes enhance antigen cross-presentation efficiency of DCs as well as an engineered derivative of 293T cells capable of cross-presentation. Enhanced antigen cross-presentation observed upon expression of the Rab5CA, Rab7DN and Rab22CA mutants remains dependent on proteasome activity. However, expression of the mutants renders cross-presentation independent of the activity of Transporter associated with antigenic peptide (TAP). These findings taken together reveal a novel TAP independent but proteasome dependent mechanism of MHC class I-restricted antigen processing and peptide loading that is facilitated by slow maturation of endocytic compartments.

Citation Format: Debrup Sengupta, Peter Cresswell. Slow maturation of early phagosomes promotes a TAP independent antigen cross-presentation pathway. [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 B059.
Abstract B069: Systemic alterations in T cell subpopulations of breast cancer patients
Authors: Pinho Mariana Pereira , Barbuto José Alexandre Marzagão
Published: May 2016

Breast cancer patients have immune system functional deviations, that affect immune cells that infiltrate the tumor, but also cells from the blood compartment, even in the absence of a metastatic disease. Blood monocytes of these patients, for example, have impaired ability to differentiate into dendritic cells. Little is known, however, about systemic alterations in T cells from these patients. To address this issue, we evaluated T cell populations in peripheral blood of cancer patients, comparing it to those of healthy donors. For this, peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Paque centrifugation, stained with antibodies to different T cell markers, and evaluated by flow cytometry. Patients had lower CD4:CD8 ratio then healthy donors (Healthy: 2.91±0.31; Cancer: 2.06±0.32; n=10; p=0.04). There was a positive correlation between the histological grade of the tumor and the CD4:CD8 ratio, where those with the lower grade had the lower CD4:CD8 ratio (r2=0.5679; p=0.03). Regarding CD4+ T cells, cancer patients had the same percentage of effector (CD45RA+CCR7-) and central memory (CD45RA-CCR7+) CD4+ T cells, but significantly lower naïve (CD45RA+CCR7+) CD4+ T cells (Healthy: 30.82±5.94, n=8; Cancer: 14.21±3.46, n=10; p=0.02) and a tendency to have higher effector memory (CD45RA-CCR7-) CD4+ T cells (Healthy: 35.13±5.54, n=8; Cancer: 48.64±4.52, n=10; p=0.08). When comparing CD8+ T cells, patients also had significantly lower naïve CD8+ T cells (Healthy: 42.74±6.97, n=8; Cancer: 19.16±3.42, n=10; p=0.005) and higher effector memory CD8+ T cells (Healthy: 29.69±5.35, n=8; Cancer: 45.37±5.15, n=10; p=0.05). Interestingly, cancer patients seemed to have higher percentage of effector CD8+ T cells than healthy patients (Healthy: 19.54±5.30, n=8; Cancer: 30.30±5.49, n=10; p=0.18). No difference in the expression of the IL-2 high affinity receptor (CD25) was observed between individuals with or without breast cancer. In conclusion, breast cancer patients had significant alterations in the percentage of the different peripheral blood T cell populations, suggesting that these may affect their ability to respond to the tumor, but also to other antigens.

Financial Support: FAPESP (2014/25988-1).

Citation Format: Mariana Pereira Pinho, José Alexandre Marzagão Barbuto. Systemic alterations in T cell subpopulations of breast cancer patients. [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 B069.
Abstract B081: Tissue repair function of regulatory T cells during infection and cancer progression
Authors: Hu Wei , Arpaia Nicholas , Bos Paula D. , Green Jesse , Rudensky Alexander
Published: May 2016

Regulatory T (Treg) cells dampen immune responses to avoid excessive, deleterious inflammation in the context of autoimmunity, allergic reactions, and infection. Treg cells also impede anti-tumor immunity. Therefore, modulation of Treg cell activity is of great therapeutic potential for treating many debilitating inflammatory diseases and cancer. Recent studies have implicated a specialized tissue-resident Treg subset in limiting metabolic inflammation in the adipose tissue and in promoting repair of muscle injury. These Treg cells exhibit a unique transcriptional signature represented by high expression of the EGF family member amphiregulin (Areg), suggesting an additional role for Treg cells in tissue maintenance and remodeling. Whether Treg cells regulate tissue maintenance in broader settings, such as during infection and cancer progression, however, remains unknown. In the present study, we observed an Areg+ Treg subset in lung-infiltrating Treg cells following influenza infection. Selective ablation of Areg in Treg cells led to severe impairment of lung function and tissue damage without evident change in viral burden or antiviral response, indicating that the tissue maintenance function of Treg cells is distinct from their immune regulatory roles. Remarkably, we found that tumor infiltrating Treg cells highly expressed Areg. Since Areg is known to exert pleiotropic oncogenic activities, Areg production by Treg cells may be crucial for establishing the tumor microenvironment. Future studies will focus on: (1) Further examine the role for Areg+ Treg cells in tissue remodeling, during infection as well as cancer progression; (2) Characterize the Areg+ Treg cell subset; (3) Define the differentiation pathway of the Areg+ Treg cells. These studies will improve our understanding of both the immune and non-immune functions of Treg cells, and could provide new strategies for selectively targeting particular aspects of Treg cell function therapeutically in the treatment and control of inflammatory diseases and cancer progression.

Note:This abstract was not presented at the conference.

Citation Format: Wei Hu, Nicholas Arpaia, Paula D. Bos, Jesse Green, Alexander Rudensky. Tissue repair function of regulatory T cells during infection and cancer progression. [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 B081.
Abstract B083: Human tumor-based identification of immune checkpoint targets
Authors: McQuade Thomas , Wang Yanyan , Doty Daniel , Amirina Najmia , Vadde Sri , Chamarthi Hareesh , Nguyen Cokey , Ranganath Sheila , Watkins-Yoon Jennifer
Published: May 2016

The recent advances in immune checkpoint (IC) blockade highlight the need to develop novel IC drug combinations. However, it remains challenging to determine which IC should be targeted in a particular indication. Enumeral uses an innovative approach to identify relevant ICs for which blockade could restore immune cell function. We use our proprietary single cell technology to assess the tumor microenvironment of human tissues and to elucidate the appropriate IC to antagonize on tumor infiltrating lymphocytes. Once appropriate IC targets are found in human tumors, monoclonal antibodies are generated and validated in vitro and ex-vivo. We tested combinations of monoclonal antibodies against ICs at the single cell level in tumor samples for reversal of immune cell exhaustion. Several anti-PD-1 and anti-TIM-3 lead candidates have been characterized in preparation for clinical studies using this strategy.

Citation Format: Thomas McQuade, Yanyan Wang, Daniel Doty, Najmia Amirina, Sri Vadde, Hareesh Chamarthi, Cokey Nguyen, Sheila Ranganath, Jennifer Watkins-Yoon. Human tumor-based identification of immune checkpoint targets. [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 B083.
Abstract B088: Can the pro-tumorigenic activity of Th17 cells be a therapeutic opportunity?
Authors: Gagliani Nicola , Vasely Carolina Amezcua , Xu Hao , Huber Samuel , Flavell Richard
Published: May 2016

Chronic inflammation in the intestine increases the risk of developing colorectal cancer. This becomes evident in patients with inflammatory bowel disease (IBD), such as ulcerative colitis and Crohn's patients, who indeed have a higher probability of developing colitis-associated cancers. The main aim of the project is to control colorectal cancer development by targeting Th17 cells, a subset of CD4 T cells, which are fundamental in promoting chronic intestinal inflammation when they escape the physiological controls. By the combination of different cutting edges technologies, including fate-cytokine reporter mouse models and RNA seq., we found that formerly pro-inflammatory Th17 cells can convert into regulatory T cells (Tregs) and acquire a strong IL-10-mediated anti-inflammatory function (Gagliani N. et al.; Nature; 2015). Moreover we proved that Th17 cells infiltrate the intestinal tumors and produce IL-22, a cytokine known to be pro-tumorigenic (Unpublished data). Considering that (i) Th17 cells infiltrate and support tumor development, (ii) could be converted into Tregs and (iii) that IL-10 production it has been shown to have anti-tumorigenic effect, we are about to test whether by forcing the conversion of pro-tumrogienc Th17 into anti-tumorigenic regulatory T cells, it is possible to reduce intestinal tumor development in preclinical mouse models.

Citation Format: Nicola Gagliani, Carolina Amezcua Vasely, Hao Xu, Samuel Huber, Richard Flavell. Can the pro-tumorigenic activity of Th17 cells be a therapeutic opportunity? [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 B088.
Abstract B089: Macrophage-mediated trogocytosis leads to death of antibody-opsonized tumor cells
Authors: Velmurugan Ramraj , Challa Dilip , Ram Sripad , Ober Raimund , Ward E. Sally
Published: May 2016

The interplay between innate immune cells and antibodies to control cancerous growth is of central importance for mediating direct anti-tumor effects and the induction of long-lived anti-tumor immunity. Phagocytosis of antibody-opsonized cells by macrophages through Fc gamma receptor interactions is well characterized and leads to cell death through target cell engulfment into phagosomes. By contrast, the contribution of the related process called trogocytosis, involving the ingestion of limited amounts of the target cell by phagocytic cells, to cell attrition is less certain. In the current study we have developed a high throughput, quantitative assay to distinguish whole cell phagocytosis and trogocytosis for different macrophage:breast cancer cell combinations. These analyses, combined with long term microscopic imaging, demonstrate that trogocytosis leads to death of HER2-overexpressing breast tumor cells. The implementation of multifocal plane microscopy to investigate trogocytic events at high spatiotemporal resolution reveals that these processes involve tubular extensions of opsonized tumor cells that are subsequently pinched off by the recipient macrophage. Of relevance to the design of second generation antibodies with improved efficacy, antibody engineering to enhance Fc gamma receptor interactions results in increased trogocytic activity. Collectively, these studies have significance to the rapidly expanding use of antibodies to treat cancer.

This research was supported in part by grants from the Cancer Prevention and Research Institute of Texas (CPRIT, RP 110069) and the National Institutes of Health (R01 GM85575).

Citation Format: Ramraj Velmurugan, Dilip Challa, Sripad Ram, Raimund Ober, E. Sally Ward. Macrophage-mediated trogocytosis leads to death of antibody-opsonized tumor cells. [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 B089.
Abstract B112: Identifying determinants of melanoma response to immune checkpoint inhibitors via preclinical modeling
Authors: Day Chi-Ping , Meskini Rajaa El , Graff-Cherry Cari , Michalowski Aleksandra , Ohler Zoe Weaver , Dyke Terry Van , Merlino Glenn
Published: May 2016

Immune checkpoint inhibitors (ICPi; e.g. anti-CTLA-4, anti-PD-1, anti-PD-L1) have shown great promise for melanoma treatment; however, sustained responses are achieved only in a fraction of patients, while the majority does not respond at all. Though load and patterns of mutations have been shown to be associated with IMT responses, the determinants have never been well characterized. In this study we aimed to identify predictive markers of melanoma response to ICPi by preclinical modeling. Three genetically engineered mice (GEM) were built to model melanomagenesis driven by different carcinogenic processes: (1) UV in albino BRAF(V600E); PTEN-knockout C57BL/6 mice, (2) carcinogen DMBA in pigmented Hgf-tg; CDK4(R24C) C57BL/6 mice, and (3) UV in pigmented Hgf-tg C57BL/6 mice. Melanoma cells derived from these models were subjected for exome sequencing and immunogenicity test by vaccination and challenge. Their responses to anti-CTLA-4 antibody were then tested in preclinical settings. Model (1), (2) and (3) are non-, medium-, and high-immunogenic, respectively. Consistent with published data, responses to anti-CTLA-4 are well correlated with the immunogenicity of individual melanoma models. Moreover, co-administration of BRAF or MEK inhibitors with immunotherapy did not enhance therapeutic efficacy. Based on exome sequencing, all the models exhibit coding mutations in genes of neuronal development, MAPK pathways, and G protein-coupled pathways, but UV-type mutations (C to T and G to A) were enriched only in model (1). Interestingly, deleterious mutations in DNA damage response (DDR) genes existed only in immunogenic models (2) and (3). These results suggested that carcinogenic mechanisms determine tumor immunogenicity and therefore response to IMT. Consistent with recent data from clinical studies, mutations in DDR genes may be predictive of IMT response. Currently we are evaluating the functions of specific DDR genes in melanoma, and the possibility they may serve as therapeutic targets in combination therapies with ICPi.

Citation Format: Chi-Ping Day, Rajaa El Meskini, Cari Graff-Cherry, Aleksandra Michalowski, Zoe Weaver Ohler, Terry Van Dyke, Glenn Merlino. Identifying determinants of melanoma response to immune checkpoint inhibitors via preclinical modeling. [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 B112.
Abstract B118: Preclinical model of patient-derived tumor transplant in humanized mice for cancer immunotherapy
Authors: Wulf-Goldenberg Annika , Stecklum Maria , Fichtner Iduna , Hoffmann Jens
Published: May 2016

The identification and validation of new targets for antitumor immune therapy is still a challenge for the preclinical research as the classical syngeneic tumor models are of limited translational value and patient-derived human tumor xenograft models (PDX) are growing on immunodeficient animals.

To overcome these constraints our aim is the development of PDX models on mice with a functional human immune system to improve predictability of drug efficacy and safety.

We reconstituted a human immune system by engrafting human hematopoietic stem cells isolated from cord blood in immunodeficient mice. A stable expression of lymphoid cell lineages in peripheral blood and secondary lymphoid tissues could be detected by flow cytometry and immunohistochemistry analysis.

At the time when the human immune system is developed, established patient-derived tumors were transplanted on these humanized mice. Human tumors developed on humanized mice without evidence of rejection. Tumors from different tumor entities grow on humanized mice with a similar level of engraftment compared to not-humanized mice.

We also tested the effect of ipilimumab (anti-CTLA-4 [cytotoxic T-lymphocyte antigen-4] mAB) and the humanized monoclonal antibody herceptin in these tumor-bearing humanized mice. We found ipilimumab could lead to a slight tumor growth delay and an increased percentage of T-cells in the blood and in the tumor.

Initially results revealed that our humanized mouse models could enable a more appropriate preclinical assessment of immune-based therapeutic antitumor strategies especially when combining the humanized mouse with patient-derived xenografts.

Citation Format: Annika Wulf-Goldenberg, Maria Stecklum, Iduna Fichtner, Jens Hoffmann. Preclinical model of patient-derived tumor transplant in humanized mice for cancer immunotherapy. [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 B118.
Abstract B127: Tumor eradication in a murine model of glioma with vaccination and anti-CTLA-4 but not anti-PD-1
Authors: Field Cameron S. , Hunn Martin K. , Ferguson Peter M. , Yagita Hideo , Hermans Ian F. , Ancelet Lindsay R.
Published: May 2016

Vaccine-mediated cancer treatment is generally insufficient to bestow long-term survival unless suppressive mechanisms are overcome. Given the success of antibody-mediated immune checkpoint blockade in relieving suppression of anti-tumor T cell responses that develop naturally in tumor-burdened hosts, we investigated whether checkpoint blockade can be used to improve the efficacy of vaccine-induced T cell responses. Combining an adjuvanted whole-cell vaccine with antibodies to cytotoxic T lymphocyte antigen-4 (α-CTLA-4) resulted in regression of established intracranial tumors, whereas neither treatment was effective as a monotherapy. In contrast, combining the vaccine with antibodies to programmed death-1 (α-PD-1) failed to provide any survival advantage despite α-PD-1 being effective against the same tumor implanted subcutaneously, which was attributed to poor access of α-PD-1 to T cell effectors in the brain. The effective combination of vaccine and α-CTLA-4 was associated with enhanced proliferation and accumulation of T cells in the lymphoid tissues without any obvious changes in the stimulatory function of antigen-presenting cells or the number of regulatory T cells, suggesting recently primed T cells were the targets of checkpoint inhibition. While tumors regressing under this combined treatment were highly infiltrated with a variety of leukocytes, tumor eradication was strictly dependent on CD4+ T cells. These results demonstrate that vaccination can be combined effectively with checkpoint blockade to induce effective T cell-mediated responses against glioma. However, new strategies are required to improve access of inhibitors to the brain if immune checkpoints on intratumoral effector cells are to be targeted.

Citation Format: Cameron S. Field, Martin K. Hunn, Peter M. Ferguson, Hideo Yagita, Ian F. Hermans, Lindsay R. Ancelet. Tumor eradication in a murine model of glioma with vaccination and anti-CTLA-4 but not anti-PD-1. [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 B127.
Abstract B136: Development of PD-L1/L2 over-expressing mouse liver cancer models to test the efficacy of anti-PD1 therapy for hepatocellular carcinoma
Authors: Ou Da-Liang , Chen Shu-Ching , Su Yu-Chia , Cheng Ann-Lii , Hsu Chiun
Published: May 2016

Anti-PD1 therapy has shown promising anti-tumor activities in multiple cancer types, including hepatocellular carcinoma (HCC), and PD-L1 expression in tumor cells was considered a potential predictive marker for treatment efficacy. The present study explored whether PD-L1/L2 expression may affect treatment efficacy of anti-PD1 therapy for HCC. PD-L1 (MR203953; Origene Technologies, Rockville, MD) or PD-L2 (MR222499; Origene Technologies) was transfected into the BNL-MEA murine liver cancer cell lines and stable clones were selected. Effects of PDL1/L2 over-expression were confirmed by Western blotting and immunofluorescence assay. Effects of PDL1/L2 over-expression on in vitro growth characteristics were compared by MTT, colony formation, and 3-D culture assays. Orthotopic liver cancer models were established by sub-capsular injection of parental or transfected cancer cells into the left lobe of mouse liver. The anti-tumor efficacy of anti-mouse PD1 therapy (clone 4H2, Ono Pharmaceutical Co), 0.2 mg by intraperitoneal injection on day 5, 8, 11, 14, and 17 after tumor implantation, was measured by change in tumor volume using the orthotopic liver cancer models. Distribution of T-cell sub-populations in tumor-infiltrating lymphocytes (TILs) after treatment were measured by flow cytometry.

The growth characteristics of parental and PD-L1/L2-expressing BNL-MEA cells did not differ significantly, as measured by MTT, colony formation, and 3-D culture assays. Survival of animals bearing the orthotopic liver tumor with or without PD-L1/L2 over-expression was also similar. Anti-PD1 therapy appeared more efficacious in PD-L1-over-expressing liver cancer models than the parental or PD-L2-over-expressing models. Distinctive TILs distribution patterns between parental and PD-L1/L2 over-expressing liver cancer models were found after anti-PD1 therapy.

Conclusions: The PD-L1/L2 expressing liver cancer models may help evaluate the efficacy and mechanisms of action of anti-PD1 therapy for HCC.

Citation Format: Da-Liang Ou, Shu-Ching Chen, Yu-Chia Su, Ann-Lii Cheng, Chiun Hsu. Development of PD-L1/L2 over-expressing mouse liver cancer models to test the efficacy of anti-PD1 therapy for hepatocellular carcinoma. [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 B136.
Abstract B172: Oncogene-mediated immune microenvironments in lung cancer provide a rationale for combination therapies
Authors: Havel Lauren S. , Gao Dingcheng , Altorki Nasser , Mittal Vivek
Published: May 2016

Non-small cell lung cancer (NSCLC) constitutes ~80% of all lung cancer cases and has a 5-year survival rate of only 15-20%. Over the past decade, targeted therapies have shown some efficacy in NSCLC patients who present drug sensitive mutations. However, acquired drug resistance has been a major obstacle to achieving durable responses. Novel and more effective combinatorial treatments are urgently needed in the clinic to improve the dismal prognosis of NSCLC patients. Recently, immune checkpoint inhibitors have led to durable response rates in NSCLC patients of up to 25%, but a large portion of patients continue to show minimal or no response. Notably, the mechanisms responsible for regulating the tumor's response to immune checkpoint inhibitors have not been elucidated. There is increasing evidence that oncogenes modulate expression of immune regulatory genes, leading to the notion that therapies that target specific oncogene pathways may influence diverse aspects of the immune response. To this end, we have focused on mouse models of NSCLC carrying mutations in the KRAS or EGFR oncogenes or the EML4-ALK translocation. These mutations were chosen because 1) together they account for >40% of the NSCLC patients and 2) FDA approved targeted therapies are available for EML4-ALK and EGFR mutants. We have performed comprehensive immunophenotyping in each of these models and observed mutation-specific distribution and abundance of specific immune subsets, suggesting diverse mechanisms and varying degrees of immune invasion/exclusion. These findings have predictive value in determining which tumor has the most likelihood of responding to specific immune checkpoint inhibitors (anti-PD-1, anti-PD-L1 or anti-CTLA4). We will discuss the implication of this approach in improving response rates to other standard of care treatments such as targeted therapies, radiation and/or chemotherapy in combination with the checkpoint inhibitors. Overall, this work has the potential to provide a mechanism-based rationale for combining standard of care treatments and immunotherapies for the treatment of NSCLC.

Citation Format: Lauren S. Havel, Dingcheng Gao, Nasser Altorki, Vivek Mittal. Oncogene-mediated immune microenvironments in lung cancer provide a rationale for combination therapies. [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 B172.
Abstract IA20: Cancer subtypes and their immune microenvironments
Authors: Becht Etienne , Giraldo Nicolas A. , Reynies Aurelien De , Laurent-Puig Pierre , Beuselinck Benoit , Zucman-Rossi Jessica , Dieu-Nosjean Marie Caroline , Sautes-Fridman Catherine , Fridman Wolf H.
Published: May 2016

In the vast majority of cancer types, as demonstrated and exemplified in colorectal cancer (CRC) and non-small cell lung cancer (NSCLC), a high density of CD8+ T cells associated with high densities of tertiary lymphoid structures (TLS) in the tumor microenvironment correlates with a good prognosis for the patient. However, an opposite association has been reported in primary clear cell renal cell carcinoma (RCC) and several other cancers. Our team studied the immune infiltrates of pulmonary metastases from CRC and RCC. As in the primary tumors, a high density of CD8+ T cells correlated with good prognosis for CRC metastases, while it correlated with a bad prognosis for RCC metastases. These results suggested that the identity of the tumor cells, rather than the organ where they grow, is critical for shaping the immune contexture of a given tumor.

Transcriptomic analyses of large cohorts of human CRC and RCC allow to define molecular subgroups associated with distinct risks of tumor progression. We therefore characterized the immune microenvironments of the molecular subgroups of CRC and RCC. In CRC, the MSI-enriched subgroup identified patients with high cytotoxic T cells infiltration and favorable prognosis. In CRC and RCC, we confirmed that patients with low tumoral T and B cell infiltrations presented with high risk of death; In addition, in both cancer types, we identified subgroups of poor-prognosis patients with high tumoral lymphocyte infiltration, in the context of high expression of genes related to inflammation, immunosuppression, and angiogenesis.

Integration of molecular and immune tumor phenotypes refines prognostic cancer groups. Moreover, these classifications allow to select immunotherapies appropriate to specific, potentially responding, groups of patients.

Citation Format: Etienne Becht, Nicolas A. Giraldo, Aurelien De Reynies, Pierre Laurent-Puig, Benoit Beuselinck, Jessica Zucman-Rossi, Marie Caroline Dieu-Nosjean, Catherine Sautes-Fridman, Wolf H. Fridman. Cancer subtypes and their immune microenvironments. [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 IA20.
Abstract IA23: T cell memory and exhaustion
Authors: Ahmed Rafi
Published: May 2016

Chronic antigen stimulation during persistent infections or cancer leads to functional exhaustion of CD8 T cells. The inhibitory receptor PD-1 plays a major role in this T cell dysfunction and blockade of this inhibitory pathway restores function in exhausted T cells. T cell exhaustion and the role of PD-1 in chronic infection were first documented in the mouse model of chronic LCMV infection and these findings were then extended to chronic viral infections in non-human primates and humans. In this talk I will discuss combinatorial strategies that enhance PD-1 directed immunotherapy and describe how PD-1 blockade acts not only at the level of increasing T cell numbers but plays a key role in target cell elimination. An interesting aspect of PD-1 biology is that PD-1 is also expressed during the early phase of T cell activation. In this talk I will also discuss how this early PD-1 expression is regulated and its potential role in CD8 T cell differentiation.

Citation Format: Rafi Ahmed. T cell memory and exhaustion. [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 IA23.