bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B48: PI3Kδ-specific inhibitors may act as adjuvants in the immunotherapy of glioblastoma
Authors: Azam Zulfikar , Wei Shao , To Tony
Published: March 2020

Glioblastoma is the most devastating form of primary brain tumor. Despite surgery, radiotherapy, and concomitant temozolomide chemotherapy, the survival rate remains disappointing. Multiple factors including a diverse array of signaling pathways play significant roles in glioblastoma pathogenesis. Of note, the phosphoinositide 3-kinase (PI3K) signaling pathway is often aberrant in glioblastoma, leading to uncontrolled cell proliferation, migration, and invasion into nearby normal brain tissues. The PI3Kδ isoform, predominantly found in leukocytes, is expressed in a number of tumors including glioblastoma. We had previously shown that PI3Kδ is also expressed in glioma cells such as the U87-MG cell line. Using siRNA knockdown technique, we demonstrated that it is important in glioma cell migration and invasion. Interestingly, its expression has been reported to associate with glioma cell resistance to erlotinib. To further study the mechanistic role of PI3Kδ in glioma pathogenesis, we created two knockout cell strains of U87-MG by CRISPR-Cas9 technology. We performed global transcriptome and pathway analysis comparing the parent cell line U87-MG with these knockout cell strains. Our results showed that the mRNA expression of Pleckstrin 2 (PLEK2) is dramatically reduced in PI3Kδ knockout cells. This is confirmed by quantitative RT-PCR. Western blot analysis showed that PLEK2 protein level is reduced by more than 90% and the phosphorylation of Akt at 308, but not 473, is also reduced. PLEK2 contributes to lamellipodia formation and cell spreading via actin cytoskeleton rearrangement, ultimately affecting cell migration and invasion. Our recent findings further strengthen the important role of PI3Kδ in glioma cell motility. It has been shown recently that inhibition of PI3Kα/δ directly enhances effector CD8+ T-cell activity. Thus, use of PI3Kδ-specific inhibitors may reduce expression of PLEK2 thereby reduces glioma cell migration and at the same time promotes antitumor immunity. PI3Kδ inhibitors may also be used as adjuvants together with immune checkpoint inhibitors such as pembrolizumab to prolong the survival of glioblastoma patients.

Citation Format: Zulfikar Azam, Shao Wei, Tony To. PI3Kδ-specific inhibitors may act as adjuvants in the immunotherapy of glioblastoma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B48.
Abstract B5: Reversing epigenetic gene silencing to overcome immune evasion in CNS malignancies
Authors: Ratnam Nivedita M. , Sonnemann Heather M. , Gilbert Mark R. , Giles Amber J.
Published: March 2020

Glioblastoma (GBM) is a lethal brain malignancy, and standard of care only offers a modest survival benefit. Emerging evidence of immune surveillance in the brain has prompted investigation of immunotherapy as a potential treatment strategy. However, preliminary studies have demonstrated impaired immune cell trafficking to the tumor, making GBMs immunologically “cold.” One mechanism of immune evasion by the tumor is to epigenetically silence the expression of chemoattractant cytokines. Previous studies in ovarian and prostate cancer have successfully established the ability of GSK126, a small-molecule inhibitor of histone methyl transferase EZH2, to promote tumor T-cell infiltration and subsequent antitumor response. Mechanistically, GSK126 was shown to promote expression of interferon gamma (IFN gamma)-induced chemokines CXCL9 and CXCL10. In the present study we evaluated the therapeutic efficacy of GSK126 in preclinical models of GBM. Our studies determined that treatment of murine and human glioblastoma cell lines with GSK126 increased IFN gamma-mediated gene expression of CXCL9 and CXCL10. Transwell migration assays performed using human T cells demonstrated increased T-cell migration when exposed to conditioned medium from tumor cells treated with GSK126 and IFN gamma. In vivo, efficacy of GSK126 was tested in combination with anti-PD-1 antibody in C57Bl/6 immunocompetent mice subcutaneously implanted with the syngeneic glioma cell line CT2A. Mice treated with the combination of GSK126 and anti-PD-1 antibody showed a significant decrease in tumor volume and improved overall survival. Tumors treated with combination therapy showed increased infiltration of CD8+ IFN gamma expressing T cells and increased CXCR3+ migratory T cells in the tumor-draining lymph node. Interestingly, treatment of mice with intracranial CT2A tumors with GSK126 alone resulted in small tumors with better prognosis. Treatment-related survival benefit was lost in immune-compromised mice, suggesting that efficacy depended upon drug-related increased lymphocyte trafficking and subsequent T cell-mediated antitumor response. Analysis of post-treatment tumor by LC-MS confirmed that GSK126 crosses the blood-brain barrier. Taken together, our data suggest a potential synergistic therapeutic role for GSK126 in glioblastoma by enhancing current immunotherapy regimens with reversing the epigenetic changes that enable immune cell evasion with resultant increases in immune cell trafficking to the tumor.

Citation Format: Nivedita M. Ratnam, Heather M. Sonnemann, Mark R. Gilbert, Amber J. Giles. Reversing epigenetic gene silencing to overcome immune evasion in CNS malignancies [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B5.
Abstract B51: Novel syngeneic animal model of tobacco-associated oral cancer reveals the activity of in situ anti-CTLA-4
Authors: Wang Zhiyong , Wu Victoria H. , Allevato Michael M. , Gilardi Mara , Saddawi-Konefka Robert , He Yudou , Callejas-Valera Juan Luis , Vitale-Cross Lynn , Martin Daniel , Amornphimoltham Panomwat , Mcdermott James , Goto Yusuke , Molinolo Alfredo A. , Sharabi Andrew B. , Cohen Ezra E.W. , Chen Qianming , Lyons Guy , Alexandrov Ludmil B. , Gutkind J. Silvio
Published: March 2020

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. Tobacco use is the main risk factor for HNSCC, and tobacco-associated HNSCCs have poor prognosis and response to available treatments. Recently approved anti-PD-1 immune checkpoint inhibitors showed limited activity (≤20%) in HNSCC, highlighting the need to identify new therapeutic options. For this, mouse models that accurately reflect the complexity of the HNSCC mutational landscape and tumor immune environment are urgently needed. Here, we report the first mouse HNSCC model system that recapitulates the human tobacco-related HNSCC mutanome, in which tumors grow when implanted in the tongue of immunocompetent mice. These HNSCC lesions have similar immune infiltration and response rates to anti-PD-1 (≤20%) immunotherapy as human HNSCCs. Remarkably, we found that >70% of HNSCC lesions respond to intratumoral anti-CTLA-4. This syngeneic HNSCC mouse model provides a platform for the development of novel immunotherapeutic options for HNSCC.

Note:This abstract was not presented at the conference.

Citation Format: Zhiyong Wang, Victoria H. Wu, Michael M. Allevato, Mara Gilardi, Robert Saddawi-Konefka, Yudou He, Juan Luis Callejas-Valera, Lynn Vitale-Cross, Daniel Martin, Panomwat Amornphimoltham, James Mcdermott, Yusuke Goto, Alfredo A. Molinolo, Andrew B. Sharabi, Ezra E.W. Cohen, Qianming Chen, Guy Lyons, Ludmil B. Alexandrov, J. Silvio Gutkind. Novel syngeneic animal model of tobacco-associated oral cancer reveals the activity of in situ anti-CTLA-4 [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B51.
Abstract B52: The systematic identification of novel immune checkpoint averting the metastasis of ovarian cancer: The role of IL-20/IL24-IL20RA axis
Authors: Li Jia , Qin Xuan , Xu Mengyao , Yan Jie , Wang Longlong , Shi Yi , Xiang Rong
Published: March 2020

Ovarian cancer has high mortality among all gynecologic malignancies due to the lack of clear symptoms at early stage and the metastasis of cancer cells when diagnosed with advanced stages. The disseminated ovarian cancer cells have to overcome the immune surveillance in abdominal environment, which is rich in many types of immune cells, to survive and proliferate. However, the key genes and mechanisms preventing the metastasis of ovarian cancer cells into abdominal sites remain largely unexplored. Here, through an in vivo, genome-scale CRISPR/Cas9 knockout screening, we identified IL-20/IL24-IL20RA as a key checkpoint preventing the abdominal metastasis of ovarian cancer cells. In ovarian cancer patients, the level of IL20RA is dramatically decreased in metastatic cancer cells when compared with that in primary sites. Overexpression of IL20RA in highly metastatic ovarian cancer cells greatly suppressed the abdominal metastasis of ovarian cancer and the formation of ascites. In contrast, silencing IL20RA could increase the metastasis of ovarian cancer cells. We further examined IL02RA-mediated crosstalk between the disseminated cancer cells and the immuno-microenvironment in mouse in vivo ovarian cancer models and in vitro coculture experiments. Silencing IL20RA in ovarian cancer cells resulted in a dramatic decrease of M1 subtype of macrophages and increase of the M2 subtype of macrophages without affecting the population of T and B lymphocytes in ascites. Further mechanistic studies revealed that the ovarian cancer cells, when invaded into the abdominal sites, greatly induced the mesothelial cells in peritoneum to produce IL20RA ligands, i.e., IL-20 and IL-24, which in turn activated the IL20RA downstream signaling in ovarian cancer cells to trigger the production and maturation of IL-18. IL-18, when secreted into the microenvironment, was able to promote the polarization of macrophages into M1-subtype to clear the cancer cells. Thus, we identified a novel immune checkpoint IL20/IL24-IL20RA that was always downregulated in metastatic ovarian cancer cells to prevent the formation of an antitumor immune microenvironment, which could be utilized to develop new strategies for the immunotherapy of ovarian cancers.

Citation Format: Jia Li, Xuan Qin, Mengyao Xu, Jie Yan, Longlong Wang, Yi Shi, Rong Xiang. The systematic identification of novel immune checkpoint averting the metastasis of ovarian cancer: The role of IL-20/IL24-IL20RA axis [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B52.
Abstract B56: YAP1 inhibits host cell immunity and increases human papillomavirus (HPV) infection to drive cervical tumorigenesis
Authors: Lv Xiangmin , He Chunbo , Huang Cong , Wang Hongbo , Chen Li , Chen Peichao , DavisS John S. , Rueda Bo R. , Wang Cheng
Published: March 2020

Although human papillomavirus (HPV) is believed to be the major causative agent for cervical cancer, accumulating evidence indicates that HPV alone is insufficient to induce cervical cancer. The molecular mechanisms underlying cervical cancer development remain unclear. In the present study, we provide evidence showing that the Hippo/YAP pathway plays a critical role in cervical carcinogenesis. We found that genetic alteration of the Hippo/YAP pathway frequently occurred in human cervical cancer patients and was associated with poor prognosis of these patients. Keratin 14 promoter-driven expression of constitutively active YAP1 (YAP S127A) in mouse cervical epithelial cells induced invasive cervical squamous cell carcinoma (CVSCC), suggesting that hyperactivation of YAP1 is sufficient to induce cervical cancer. Importantly, hyperactivation of YAP1 in cervical epithelial cells not only increased the putative HPV receptor molecules and cellular susceptibility to HPV infection, but also disrupted the host cell innate immune system, resulting in failure of HPV viral recognition, suppression of type I IFN production, inhibition of the IFNRs/JAKs/STATs pathway, and blockage of production of antiviral interferon-stimulated genes. Cervical epithelial-specific YAP1 and HPV double knockin mouse models showed that YAP1 synergizes with HPV to promote the initiation and progression of CVSCC. The present study demonstrates that the Hippo/YAP pathway plays a central role in the development of CVSCC and the synergism between hyperactivated YAP1 and high-risk HPV may be the key driver of cervical cancer development. The Hippo/YAP signaling pathway may represent a promising target for developing novel strategies to prevent and treat cervical cancer.

Citation Format: Xiangmin Lv, Chunbo He, Cong Huang, Hongbo Wang, Li Chen, Peichao Chen, John S. DavisS, Bo R. Rueda, Cheng Wang. YAP1 inhibits host cell immunity and increases human papillomavirus (HPV) infection to drive cervical tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B56.
Abstract B58: Gasdermin E suppresses tumor growth by activating antitumor immunity
Authors: Zhang Zhibin , Zhang Ying , Xia Shiyu , Kong Qing , Li Shunying , Liu Xing , Junqueira Caroline , Ansara James , Sengupta Satyaki , Yao Yandan , Wu Hao , Lieberman Judy
Published: March 2020

The gasdermins (GSDM) are a family of homologous pore-forming proteins that cause pyroptosis, an inflammatory death associated with loss of cell membrane integrity and release of inflammatory mediators. Gasdermin E (GSDME, also known as DFNA5) as the most ancient member of the GSDM family has been implicated as a tumor suppressor. GSDME is epigenetically inactivated by promoter DNA methylation in a number of cancers and mutated in some other cancers. Worse 5-year survival and an increase in lymph node metastases are associated with reduced GSDME in breast cancer. During classic apoptosis, caspase-3 activates GSDME by cleaving after Asp270. GSDME activation converts noninflammatory apoptotic death to an inflammatory pyroptotic death. This switch could have a profound effect on the ability of tumor cells to survive intrinsic cellular stresses and extrinsic challenges, such as ER stress, chemotherapy drugs, or immune attack by cytotoxic lymphocytes. Here we show reduced GSDME function of 7 of 8 tested cancer-associated mutations. By manipulating GSDME expression in diverse cancer cell lines, we show that Gsdme knockout in GSDME-expressing tumors enhances, while ectopic expression in Gsdme-repressed tumors inhibits, tumor growth. Our data demonstrate that GSDME tumor-suppressive effect is cell-extrinsic, mediated by cytotoxic lymphocyte killing and abrogated in perforin-deficient mice or mice lacking lymphocytes. Tumor suppression depended on GSDME’s ability to form cell membrane pores that cause inflammatory pyroptosis, since it was completely blocked by mutating the GSDME cleavage site for pore formation. Using different cancer models, we further show that GSDME expression enhances tumor-associated macrophage phagocytosis and the number and functions of tumor-infiltrating NK and CD8+ T lymphocytes. Importantly, we identify that immune killer cells could activate GSDME-mediated pyroptosis independently of caspase activation, when killer cell cytotoxic protease granzyme B directly cleaves and activates GSDME at the same site as caspase-3 (Asp270). Therefore, immune killer cells induce pyroptosis in target GSDME-expressing cancer cells, creating an inflammatory tumor microenvironment that further facilities immune cell tumor infiltration and forms a positive feedback loop. Thus, GSDME suppresses tumor growth by activating antitumor immunity.

Citation Format: Zhibin Zhang, Ying Zhang, Shiyu Xia, Qing Kong, Shunying Li, Xing Liu, Caroline Junqueira, James Ansara, Satyaki Sengupta, Yandan Yao, Hao Wu, Judy Lieberman. Gasdermin E suppresses tumor growth by activating antitumor immunity [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B58.
Abstract B62: Development of novel chimeric antigen receptor T cells for immunotherapy of hepatocellular carcinoma
Authors: Jiang Xiaotao , Caraballo G. Leidy D. , Zhang Huajun , Chi Xiangyang , Peng Yibing , He Aiwu R. , Shao Yanxia , Cai Lun , He Yukai
Published: March 2020

Background and Aim: Several glypican 3 (hGPC3)-specific chimeric-antigen-receptor modified T cells (CARTs) are in trials for hepatocellular carcinoma (HCC), but most are constructed from one monoclonal antibody (mAb). It is unknown how targeting different hGPC3 epitopes will affect CART’s efficacy. The aim of this study is to develop novel effective CARTs by targeting different regions of hGPC3.

Methods: BalB/C mice were immunized with hGPC3 protein to generate novel mAbs that targeted different regions of hGPC3. CARTs were then built from mAbs and their antitumor effect was studied.

Results: Twenty-two mAbs were identified by ELISA. Fourteen of them bound HepG2 cells. Five mAbs were further characterized by immunohistochemical staining. Three of them (6G11, 8F8, and 12D7) were found to specifically stain HCC tumors but not adjacent normal tissues. The mAbs’ affinity was in the nanomolar range. 6G11 and 8F8 bound to hGPC3 N-(AA25-39) and C-(AA463-496) epitopes, respectively. 12D7 recognized a conformational epitope in the hGPC3 N-fragment (AA25-358). All CARTs built from the mAbs underwent expansion after GPC3+ cell stimulation. However, their effector function was significantly different. 8F8 CARTs possessed the strongest effector function. 6G11 CARTs experienced the greatest expansion, but with slightly weaker function. In contrast, 12D7 CARTs showed minimal effector function. Soluble hGPC3 did not activate CARTs or block CART activation by tumor cells. Adoptive transfer of 8F8 and 6G11, but not 12D7, CARTs generated complete regression of HCC xenografts in NSG mice.

Conclusion: The three novel CARTs that target a membrane-distal N-epitope, a membrane-proximal C-epitope, or a conformational epitope of hGPC3 possessed different effector function and antitumor effects. CARTs targeting the hGPC3 N- or C specific epitope generated potent antitumor effects, while CARTs targeting an hGPC3-N conformational epitope showed minimal effector function.

Citation Format: Xiaotao Jiang, Leidy D. Caraballo G., Huajun Zhang, Xiangyang Chi, Yibing Peng, Aiwu R. He, Yanxia Shao, Lun Cai, Yukai He. Development of novel chimeric antigen receptor T cells for immunotherapy of hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B62.
Abstract B72: Humanized NSG-Tg(Hu-IL15) mice support preclinical immune-oncology efficacy for testing of NK cell-based immunotherapy
Authors: Yao Li-Chin , Cheng Mingshan , Shultz Leonard D. , Greiner Dale L. , Brehm Michael A. , Keck James G.
Published: March 2020

The JAX® Onco-Hu® platform utilizes humanized mice engrafted with tumors to enable in vivo investigation of the interactions between the human immune system and human cancer. We have shown that humanized NOD-scid IL2Rγ null (NSG) mice bearing patient-derived xenografts (PDX) allow efficacy studies of checkpoint inhibitors. A major avenue of our investigation is to generate humanized mouse models containing a more complete human hematopoietic system and robust innate immune cell population. Nature killer (NK) cells are important players of innate defense against cancerous cells by a number of mechanisms, including antibody-dependent cell-mediated cytotoxicity (ADCC). To overcome limited NK cell development in humanized NSG mice, we have developed NSG-Tg(Hu-IL15) (JAX stock number 030890) mice constitutively expressing human IL15. Following HSC-engraftment of NSG-Tg(Hu-IL15) mice, significantly higher levels of human CD16+CD56+ NK cells are detectable as compared to NSG mice in peripheral blood during the whole course of the study (18 weeks). Levels of circulating human CD45+ cells, CD3+ T cells, CD19+ B cells, and CD33+ myeloid cells are similar between the HSC-engrafted NSG-Tg(Hu-IL15) and NSG mice. Previously we have described that the human NK cells developing in HSC-engrafted NSG-Tg(Hu-IL15) mice are functional and can lyse the NK-sensitive target cells K562 by in vitro cytotoxicity assay. We also showed that NK cells limit the growth of a PDX melanoma xenograft. Here we show that humanized NSG-Tg(Hu-IL15) mice mediate efficient ADCC against Daudi B lymphoma cells in vivo. Administration of anti-human CD20 antibody (rituximab) resulted in significant tumor growth inhibition, in both tumor volume and tumor weight. We observed consistent ADCC efficacy in humanized NSG-Tg (Hu-IL15) mice engrafted with three different HSC donors. Together these data demonstrate that HSC-engrafted NSG-Tg(Hu-IL15) mice support enhanced development of functional human NK cells and that this mouse model enables NK cell-targeted cancer immunotherapy for preclinical testing.

Citation Format: Li-Chin Yao, Mingshan Cheng, Leonard D. Shultz, Dale L. Greiner, Michael A. Brehm, James G. Keck. Humanized NSG-Tg(Hu-IL15) mice support preclinical immune-oncology efficacy for testing of NK cell-based immunotherapy [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B72.
Abstract B78: NIR light-triggered photoimmunotherapy selectively destroy tumor cells in a clinically relevant desmoplastic heterocellular model
Authors: Bano Shazia
Published: March 2020

Near-infrared light-triggered photoimmunotherapy, utilizing the simultaneous presence of light and targeted conjugates, has emerged as a promising approach to impart selective cytotoxicity, especially for more complex nanoconstructs. Targeted immunoconjugates promise unique capabilities of combining multiple therapeutic agents within a single nanoplatform and promise great advances in revolutionizing cancer treatments, promoting selective drug localization and photo-triggered release of secondary agents to tumors cells that overexpress specific receptors. This study has utilized smart, liposome-based photoactivatable multiagent nanoconjugates, doped with a photoactivatable chromophore benzoporphyrin derivative (BPD) labeled with an active targeting ligand cetuximab to target the EGFR receptor (overexpressed on various cancer cells) for quantitative molecular imaging to measure the true in vivo receptor specificity and localized photodynamic therapy following NIR activation, that further can trigger the release of encapsulated payload to pancreatic cancer spheroids. In an in vivo model, these targeted nanoconstructs demonstrate substantial pancreatic tumor necrosis in a receptor-specific manner and modulate the collagen content in desmoplastic model of pancreatic ductal adenocarcinoma (MIA PaCa-2) in the presence of patient-derived pancreatic cancer-associated fibroblasts (PCAFs). This study establishes a tunable path towards optimally effective multiagent nanoconjugates for tumor-specific accumulation and targeted destruction of cancer cells in complex cancer model to enhance the therapeutic index of the administrated drugs.

Citation Format: Shazia Bano. NIR light-triggered photoimmunotherapy selectively destroy tumor cells in a clinically relevant desmoplastic heterocellular model [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B78.
Abstract B87: NK cells lose their ability to control MHC-I-deficient tumor cells outside circulation
Authors: Requesens Marta , Bunting Mark , Vyas Maulik , Demehri Shawn
Published: March 2020

A common tumor immune-evasion mechanism is downregulation of major histocompatibility complex Class-I (MHC-I) that enables malignant cell escape from cytotoxic T-cell responses. This strategy also contributes to nonresponsiveness of tumors to T-cell targeted immune-checkpoint blockade. MHC-I is an antigen-presenting molecule that also functions as an inhibitory ligand for natural killer (NK) cells. Thus, loss of MHC-I should render cells susceptible to NK cell-mediated cytotoxicity. However, this does not always occur. Previously it was demonstrated that NK cells were able to strongly reject MHC-I-deficient bone marrow cells in the circulation, yet they failed to reject MHC-I-deficient organ grafts, suggesting restricted NK cell cytotoxicity in tissues. We hypothesized that a similar NK cell tolerance to MHC-I-deficient cells occurs in the context of solid cancers. We compared NK cell recruitment and activation using WT and MHC-I-deficient B16F10 melanoma models. We demonstrated that NK cells eradicate circulating MHC-I−/− B16F10 cells, effectively preventing tumor metastasis to the lungs. In contrast, NK cells failed to clear MHC-I−/− B16F10 tumors when implanted subcutaneously, nor was the growth of these tumors affected by NK cell depletion. Interestingly, spontaneous lymph node (LN) metastasis from MHC-I-deficient solid tumors was fully abrogated in an NK cell-dependent manner, indicating functional inhibition of NK cells in peripheral tissues, but not in circulation or draining lymphatics. In conclusion, we demonstrate NK cell tolerance to MHC-I-deficient tumor cells in tissues, but not in circulation, which suggests the presence of non-MHC-I NK cell-inhibitory signals in solid organs. Our research uncovers a fundamental aspect of NK cell biology and will open up the possibility of new NK cell-based therapeutic strategies to control solid tumors that escape cytotoxic CD8+ T-cell responses by downregulating MHC-I.

Citation Format: Marta Requesens, Mark Bunting, Maulik Vyas, Shawn Demehri. NK cells lose their ability to control MHC-I-deficient tumor cells outside circulation [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B87.
Abstract B89: Dual PD-1 and VEGFR-2 blockade induces vascular normalization and enhances antitumor immune responses in hepatocellular carcinoma
Authors: Datta Meenal , Shigeta Kohei , Hato Tai , Kitihara Shuji , Chen Ivy X. , Matsui Aya , Kikuchi Hiroto , Mamessier Emilie , Aoki Shuichi , Ramjiawan Rakesh R. , Ochiai Hiroki , Bardeesy Nabeel , Huang Peigen , Jain Rakesh K. , Cobbold Mark , Zhu Andrew X. , Duda Dan G.
Published: March 2020

Introduction: Only a fraction of hepatocellular carcinoma (HCC) patients benefit from programmed death receptor-1 (PD-1) blockade. Combining PD-1 blockade with antiangiogenic treatments has been able to substantially increase the fraction of responsive HCC patients, but the mechanism of this interaction is unknown. We recapitulated these clinical outcomes (response vs. resistance) using orthotopic murine models of HCC. VEGFR2 blockade significantly delayed primary tumor growth but failed to prolong survival, while anti-PD-1 monotherapy conferred a minor survival. However, dual anti-PD-1/VEGFR-2 therapy significantly inhibited primary tumor growth and doubled survival in multiple models. Combination therapy reprogrammed the immune microenvironment by increasing CD8+ cytotoxic T-cell infiltration and activation, improving the ratio of anti- vs. pro-tumor-associated macrophages, and reducing T regulatory cell (Treg) infiltration in HCC tissue. Moreover, we found that VEGFR-2 blockade increased the PD-1 expression in tumor-infiltrating CD4+ cells. Under anti-PD-1 therapy, CD4+ cells promote normalized vessel formation in the face of antiangiogenic therapy.

Conclusion: We show that dual anti-PD-1/VEGFR-2 therapy has a durable vessel fortification effect in HCC and can overcome treatment resistance to either treatment alone and increase survival in both anti-PD-1 therapy resistant and responsive HCC models.

Citation Format: Meenal Datta, Kohei Shigeta, Tai Hato, Shuji Kitihara, Ivy X. Chen, Aya Matsui, Hiroto Kikuchi, Emilie Mamessier, Shuichi Aoki, Rakesh R. Ramjiawan, Hiroki Ochiai, Nabeel Bardeesy, Peigen Huang, Rakesh K. Jain, Mark Cobbold, Andrew X. Zhu, Dan G. Duda. Dual PD-1 and VEGFR-2 blockade induces vascular normalization and enhances antitumor immune responses in hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B89.
Abstract B93: Tumor lymphangiogenesis improves responsiveness to radiotherapy in mouse melanoma
Authors: Mitrousis Nikolaos , Sasso Maria Stella , Weichselbaum Ralph R. , Hubbell Jeffrey A. , Swartz Melody A.
Published: March 2020

Radiotherapy (RT) constitutes a significant component of clinically used cancer therapy, with approximately 50% of all cancer patients receiving treatment over the course of their disease. RT is known to induce immunogenic cell death of tumor cells, leading to the release of tumor antigens, damage-associated molecular patterns, and other tumor-resident components such as cytokines. This phenomenon drives the initiation of antigen-specific immune responses, which have been shown to be essential for the complete response to RT. Notwithstanding its broad applicability and diverse mechanisms of action, RT does not usually lead to curative responses and combinations with other treatment modalities are needed to improve efficacy. Our laboratory and others have previously demonstrated that tumor-associated lymphatics can actively influence antitumor immune responses and potentiate immunotherapy. Here, we investigate whether lymphangiogenesis influences the antitumor efficacy and immune response elicited by RT. We employed the aggressive B16F10 melanoma model, that was either transduced to overexpress the lymphangiogenic factor VEGF-C (B16-VEGF-C) or mock-transduced (B16-ctrl). Tumor-bearing animals were treated by ablative localized RT and monitored for tumor growth and survival, while antitumor immune responses were assessed by flow cytometry and ELISA. We found that the therapeutic response to RT was stronger in B16-VEGF-C melanomas compared to B16-ctrl, as evidenced by a significant delay in tumor growth and prolonged animal survival. In agreement with this finding, B16-VEGF-C melanomas exhibited a significantly greater CD8+ T-cell infiltration and activation upon ex vivo restimulation after RT. These responses were partially inhibited by a function-blocking antibody of the main VEGF-C receptor, VEGFR3. Together, our findings demonstrate that lymhangiogenesis may be a key contributing factor underlying tumor responsiveness to RT.

Citation Format: Nikolaos Mitrousis, Maria Stella Sasso, Ralph R. Weichselbaum, Jeffrey A. Hubbell, Melody A. Swartz. Tumor lymphangiogenesis improves responsiveness to radiotherapy in mouse melanoma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B93.
Abstract B95: PD-L1-mediated nanotherapeutic targeting of glioma-infiltrating myeloid cells synergizes with radiotherapy for glioblastoma
Authors: Zhang Peng , Miska Jason , Lee-Chang Catalina , Rashidi Aida , Panek Wojciech K. , An Shejuan , Zannikou Markella , Lopez-Rosas Aurora , Han Yu , Xiao Ting , Balyasnikova Irina V. , Lesniak Maciej S.
Published: March 2020

Massive infiltration of tumor-associated myeloid cells (TAMCs), reaching 30-50% of the tumor mass, is a distinctive characteristic of glioblastoma (GBM). These cells are a critical player in generating highly immunosuppressive microenvironment in GBM. Thus, therapeutic targeting of TAMCs might dampen the immunosuppression and improve the outcome of antiglioma therapies. In this work, we developed a novel nano-immunotherapy platform to actively and specifically target glioma-infiltrating TAMCs in vivo. Based on our finding that glioma-infiltrating TAMCs exhibit the highest expression of PD-L1 among all the immune cell infiltrates, we designed a lipid nanoparticle system (LNP) with surface-engineered anti-PD-L1 therapeutic antibody. Decoration of anti-PD-L1 antibody rendered our LNP highly capable of specifically targeting TAMCs and, interestingly, routing PD-L1 to lysosomal degradation. Co-encapsulation of dinaciclib, a cyclin-dependent kinase inhibitor, into LNP caused a dramatic elimination of TAMCs and inhibition of immunosuppression in glioma. Notably, the targeting efficiency of TAMC-targeted LNP was further increased when combined with radiation therapy (RT), a standard care of GBM. The superior therapeutic efficacy in mice treated by RT+LNP combination over the RT or LNP monotherapy was demonstrated in two syngeneic murine glioma models, GL261 and CT2A. We also demonstrated that our LNP successfully targeted TAMCs harvested from human resection specimens, which further validated the translational potential and clinical relevance of our nano-immunotherapy approach. In conclusion, our work established a new nanotechnology-based immunotherapeutic approach with great potential to improve the clinical outcome in patients with GBM.

Citation Format: Peng Zhang, Jason Miska, Catalina Lee-Chang, Aida Rashidi, Wojciech K. Panek, Shejuan An, Markella Zannikou, Aurora Lopez-Rosas, Yu Han, Ting Xiao, Irina V. Balyasnikova, Maciej S. Lesniak. PD-L1-mediated nanotherapeutic targeting of glioma-infiltrating myeloid cells synergizes with radiotherapy for glioblastoma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B95.
Abstract B96: Calreticulin exposure on malignant blasts correlates with improved NK cell-mediated cytotoxicity in AML patients
Authors: Truxova Iva , Kasikova Lenka , Salek Cyril , Hensler Michal , Lysak Daniel , Holicek Peter , Bilkova Pavla , Holubova Monika , Chen Xiufen , Mikyskova Romana , Reinis Milan , Kovar Marek , Tomalova Barbora , Kline Justin , Galluzzi Lorenzo , Spisek Radek , Fucikova Jitka
Published: March 2020

In some settings, cancer cells responding to treatment undergo an immunogenic form of cell death that is associated with the abundant emission of danger signals in the form of damage-associated molecular patterns (DAMPs). Accumulating preclinical and clinical evidence indicates that danger signals play a crucial role in the (re-)activation of antitumor immune responses in vivo, thus having a major impact on patient prognosis. We have previously demonstrated that the presence of calreticulin (CRT) on the surface of malignant blasts is a positive prognostic biomarker for patients with acute myeloid leukemia (AML). CRT exposure not only correlated with enhanced T cell-dependent antitumor immunity in this setting but also affected the number of circulating NK cells upon restoration of normal hematopoiesis. Here, we report that CRT exposure on malignant blasts is associated with enhanced NK cell cytotoxic and secretory functions, both in AML patients and in vivo in mice. The ability of CRT to stimulate NK cells relies on CD11c+CD14high cells that, upon exposure to CRT, display higher levels of IL-15Rα, maturation markers (CD86 and HLA-DR), and CCR7. CRT exposure on malignant blasts also correlates with the upregulation of genes coding for type I interferons (IFNs). This suggests that CD11c+CD14high cells have increased capacity to migrate to secondary lymphoid organs, where can efficiently deliver stimulatory signals (IL-15Rα/IL-15) to NK cells. These findings delineate a multipronged, clinically relevant mechanism whereby surface-exposed CRT favors NK cell activation in AML patients.

Citation Format: Iva Truxova, Lenka Kasikova, Cyril Salek, Michal Hensler, Daniel Lysak, Peter Holicek, Pavla Bilkova, Monika Holubova, Xiufen Chen, Romana Mikyskova, Milan Reinis, Marek Kovar, Barbora Tomalova, Justin Kline, Lorenzo Galluzzi, Radek Spisek, Jitka Fucikova. Calreticulin exposure on malignant blasts correlates with improved NK cell-mediated cytotoxicity in AML patients [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B96.
Abstract B97: The immunomodulatory roles of tumor-associated lymphatics in triple-negative breast cancer
Authors: Hosseinchi Peyman , Potin Lambert , Mansurov Aslan , Maillat Léa , Hubbell Jeffrey A. , Swartz Melody A.
Published: March 2020

During cancer development, growth and remodeling of lymphatic vessels—termed lymphangiogenesis—is associated with increased metastasis rate. However, our group has recently shown that lymphangiogenesis can also increase tumor immune infiltration and subsequently enhance the responsiveness to immunotherapies in melanoma. However, the role of lymphangiogenesis in antitumor immunity in breast cancer remains unexplored. Moreover, despite the success of immunotherapies in different cancer types, their translation into the treatment of breast cancer has remained challenging. Here, we investigated the role of tumor lymphangiogenesis in immune cell infiltration and responsiveness to immunotherapy in a murine triple-negative breast tumor model. Spontaneously metastatic 4T1 tumor cells were transduced to overexpress the pro-lymphangiogenic growth factor VEGF-C to model tumor lymphangiogensis in vivo. We confirmed that VEGF-C overexpressing tumors were more lymphangiogenic than control ones, as assessed by flow cytometry and immunohistochemistry analyses. As previously reported, tumor VEGF-C signaling also increased the metastatic burden in the lungs. Crucially, lymphangiogenic tumors were infiltrated with increased levels of immune cells, including CD4 T cells and macrophages. Upon treatment with an agonist of the stimulator of interferon genes (STING) pathway, lymphangiogenic 4T1 tumors responded more favorably to the therapy compared to the control tumors, as indicated by a reduced tumor growth in the VEGF-C group. Furthermore, gene expression analysis of human triple-negative breast tumor samples from The Cancer Genome Atlas revealed positive correlations between VEGFC expression and CD4 T-cell and macrophage gene expressions. Taken together, these data suggest that in triple-negative breast cancer, similarly to melanoma, VEGF-C signaling is beneficial to the primary tumor microenvironment upon combination with proper immunotherapy despite promoting metastasis in untreated tumors.

Citation Format: Peyman Hosseinchi, Lambert Potin, Aslan Mansurov, Léa Maillat, Jeffrey A. Hubbell, Melody A. Swartz. The immunomodulatory roles of tumor-associated lymphatics in triple-negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B97.
Abstract IA07: Engineering natural killer cells for the treatment of cancer
Authors: Rezvani Katayoun
Published: March 2020

Natural killer (NK) cells play critical roles in host defense against cancer. Our group is exploring a number of avenues to enhance NK cell function against hematologic malignancies and solid tumors. These include novel strategies to expand off-the-shelf cord blood (CB)-derived NK cells based on their coculture with genetically modified feeder cells that express membrane-bound cytokines and costimulatory molecules. To redirect NK cell specificity and enhance their in vivo persistence, we have successfully transduced expanded CB NK cells with a retroviral vector incorporating the genes for CAR-CD19, IL-15, and inducible caspase-9-based suicide gene (iC9). Data from a first-in-human phase I/II clinical study to test the safety and efficacy of escalating doses of off-the-shelf iC9/CAR.19/IL-15 CB-NK cells in patients with relapsed or refractory B-lymphoid malignancies will be presented.

Citation Format: Katayoun Rezvani. Engineering natural killer cells for the treatment of cancer [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA07.
Abstract IA09: Checkpoint inhibitor-induced autoimmune diabetes and endocrine syndromes
Authors: Herold Kevan
Published: March 2020

Checkpoint inhibitor therapies have had dramatic effects on refractory malignancies but come at a price of autoimmune syndromes. The endocrine organs are frequently targeted. Autoimmune thyroid disease is common, but of particular clinical significance is the development of insulin-deficient diabetes. This talk will review the clinical and pathophysiologic data on this adverse event with consideration of ways in which it may be averted.

Citation Format: Kevan Herold. Checkpoint inhibitor-induced autoimmune diabetes and endocrine syndromes [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA09.
Abstract IA14: Molecular programs defining tumor-specific T-cell dysfunction and reprogrammability
Authors: Schietinger Andrea
Published: March 2020

Tumor-specific T cells in solid tumors are dysfunctional, allowing tumors to progress. The epigenetic regulation determining T-cell dysfunction and therapeutic reprogrammability is not well understood. We recently found that T-cell dysfunction is a dynamic antigen-driven differentiation program initiated early during tumorigenesis: initially, T cells enter a plastic dysfunctional state from which they can be functionally rescued, but with time and continued tumor antigen encounter, they ultimately enter a fixed dysfunctional state that is resistant to reprogramming. We found that plasticity and imprinting of dysfunction is determined by epigenetic programs, and we identified surface markers associated with plastic and fixed chromatin states that demarcated reprogrammable from non-reprogrammable PD1hi dysfunctional T cells within heterogeneous T-cell populations. Moreover, we recently identified the nuclear factor Tox as a critical regulator of tumor-specific T-cell differentiation. New insights into the epigenetic and molecular programs underlying tumor-specific T-cell differentiation and dysfunction will be discussed.

Citation Format: Andrea Schietinger. Molecular programs defining tumor-specific T-cell dysfunction and reprogrammability [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA14.
Abstract IA15: Noninvasive imaging of immune responses
Authors: Rashidian Mohammad
Published: March 2020

Immunotherapy has revolutionized cancer treatment. Notwithstanding the encouraging results, responses remain heterogeneous among patients and some face serious side effects. Hence, techniques to noninvasively predict and monitor each patient’s response to immunotherapy are greatly needed. Because the success of immunotherapy hinges on its ability to promote an antitumor immune response, monitoring of the immune response after administration of therapy can be used to evaluate patient response. We have recently developed robust immuno-PET imaging methods to monitor dynamics of immune cells in the tumor microenvironment in real time. These methods have been successful in evaluating the response to immunotherapy in many mouse models of cancer. We will discuss these new advances and their potential to evaluate and predict an ongoing response to cancer treatments, with a focus on checkpoint blockade treatments.

Citation Format: Mohammad Rashidian. Noninvasive imaging of immune responses [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA15.
Abstract IA19: Myeloid cell control of tumor immunity
Authors: Merad Miriam
Published: March 2020

While the success of T-cell checkpoint-targeted immunotherapies is predicated on the induction of an effector T-cell response against tumor antigens by potent antigen-presenting cells (APC), very little is known about the APC composition in human tumors during immunotherapy. APC form a diverse population of dendritic cells (DCs) and macrophages (macs) that share the capacity to capture and present cell-associated antigens to T cells and instruct the repertoire and effector versus regulatory function of T lymphocytes. Despite the key role of APC in tumor immunotherapy, targeting the APC compartment has proved difficult. This is because little is known about the APC molecular changes during tumor response or resistance to immunotherapy. In order to develop ways to modulate tumor APC and promote cancer immunity, we sought to gain a deeper understanding of the molecular and functional diversity of APC in human NSCLC. Using paired mass cytometry analysis of tumor/adjacent tissues, we recently identified major changes in APC and T-cell composition in early non-small lung cancer (NSCLC) lesions (Lavin et al., Cell 2017). More recently, we used cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) to further measure APC defects in the tumor lesions. CITE-seq analysis of the tumor lesions revealed profound alterations in tumor APC composition including a reduction of cross-presenting DC at the expense of monocyte-derived cells, which correlated with the presence of dysfunctional T cells compared to the adjacent tissue. Accordingly, we found that DC composition and molecular state controlled CD8+ T-cell accumulation at the tumor site and controlled tumor response to checkpoint blockade in preclinical models. Here, I will discuss the strategies we have developed to enhance DC functionality at the tumor site and the challenges that remain.

Citation Format: Miriam Merad. Myeloid cell control of tumor immunity [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA19.
Abstract IA21: Colorectal cancer and the microbiota
Authors: Garrett Wendy
Published: March 2020

Increasing scientific and clinical data support a role for the microbiota (the microorganisms associated with the human body) in influencing systemic and tumor microenvironment immunity, promoting tumor growth and spread, and altering the responsiveness and toxicity towards therapeutics. Herein, we discuss both human data from meta’omics analyses and data from mechanistic studies in cell culture and animal models that support that the microbiota influences colonic tumorigenesis.

Citation Format: Wendy Garrett. Colorectal cancer and the microbiota [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr IA21.