bluuebunny/crossref_metadata_2025 huggingface dataset:
Supplementary Data from Microenvironmental Landscape of Human Melanoma Brain Metastases in Response to Immune Checkpoint Inhibition
Authors: Alvarez-Breckenridge Christopher , Markson Samuel C. , Stocking Jackson H. , Nayyar Naema , Lastrapes Matt , Strickland Matthew R. , Kim Albert E. , de Sauvage Magali , Dahal Ashish , Larson Juliana M. , Mora Joana L. , Navia Andrew W. , Klein Robert H. , Kuter Benjamin M. , Gill Corey M. , Bertalan Mia , Shaw Brian , Kaplan Alexander , Subramanian Megha , Jain Aarushi , Kumar Swaminathan , Danish Husain , White Michael , Shahid Osmaan , Pauken Kristen E. , Miller Brian C. , Frederick Dennie T. , Hebert Christine , Shaw McKenzie , Martinez-Lage Maria , Frosch Matthew , Wang Nancy , Gerstner Elizabeth , Nahed Brian V. , Curry William T. , Carter Bob , Cahill Daniel P. , Boland Genevieve Marie , Izar Benjamin , Davies Michael A. , Sharpe Arlene H. , Suvà Mario L. , Sullivan Ryan J. , Brastianos Priscilla K. , Carter Scott L.
Published: April 2023

Supplementary Data from Microenvironmental Landscape of Human Melanoma Brain Metastases in Response to Immune Checkpoint Inhibition
Data from Calnexin Impairs the Antitumor Immunity of CD4+ and CD8+ T Cells
Authors: Chen Yichen , Ma Da , Wang Xi , Fang Juan , Liu Xiangqi , Song Jingjing , Li Xinye , Ren Xianyue , Li Qiusheng , Li Qunxing , Wen Shuqiong , Luo Liqun , Xia Juan , Cui Jun , Zeng Gucheng , Chen Lieping , Cheng Bin , Wang Zhi
Published: April 2023

Abstract
Elucidation of the mechanisms of T-cell–mediated antitumor responses will provide information for the rational design and development of cancer immunotherapies. Here, we found that calnexin, an endoplasmic reticulum (ER) chaperone protein, is significantly upregulated in oral squamous cell carcinoma (OSCC). Upregulation of its membranous expression on OSCC cells is associated with inhibited T-cell infiltration in tumor tissues and correlates with poor survival of patients with OSCC. We found that calnexin inhibits the proliferation of CD4+ and CD8+ T cells isolated from the whole blood of healthy donors and patients with OSCC and inhibits the secretion of IFNγ, TNFα, and IL2 from these cells. Furthermore, in a melanoma model, knockdown of calnexin enhanced the infiltration and effector functions of T cells in the tumor microenvironment and conferred better control of tumor growth, whereas treatment with a recombinant calnexin protein impaired the infiltration and effector functions of T cells and promoted tumor growth. We also found that calnexin enhanced the expression of PD-1 on CD4+ and CD8+ T cells by restraining the DNA methylation status of a CpG island in the PD-1 promoter. Thus, this work uncovers a mechanism by which T-cell antitumor responses are regulated by calnexin in tumor cells and suggests that calnexin might serve as a potential target for the improvement of antitumor immunotherapy.
Data from GITR Pathway Activation Abrogates Tumor Immune Suppression through Loss of Regulatory T-cell Lineage Stability
Authors: Schaer David A. , Budhu Sadna , Liu Cailian , Bryson Campbell , Malandro Nicole , Cohen Adam , Zhong Hong , Yang Xia , Houghton Alan N. , Merghoub Taha , Wolchok Jedd D.
Published: April 2023

Abstract
Ligation of GITR (glucocorticoid-induced TNF receptor-related gene, or TNFRSF18) by agonist antibody has recently entered into early-phase clinical trials for the treatment of advanced malignancies. Although the ability of GITR modulation to induce tumor regression is well documented in preclinical studies, the underlying mechanisms of action, particularly its effects on CD4+Foxp3+ regulatory T cells (Treg), have not been fully elucidated. We have previously shown that GITR ligation in vivo by agonist antibody DTA-1 causes more than 50% reduction of intratumor Tregs with down modulation of Foxp3 expression. Here, we show that the loss of Foxp3 is tumor dependent. Adoptively transferred Foxp3+ Tregs from tumor-bearing animals lose Foxp3 expression in the host when treated with DTA-1, whereas Tregs from naïve mice maintain Foxp3 expression. GITR ligation also alters the expression of various transcription factors and cytokines important for Treg function. Complete Foxp3 loss in intratumor Tregs correlates with a dramatic decrease in Helios expression and is associated with the upregulation of transcription factors, T-Bet and Eomes. Changes in Helios correspond with a reduction in interleukin (IL)-10 and an increase in IFN-γ expression in DTA-1–treated Tregs. Together, these data show that GITR agonist antibody alters Treg lineage stability inducing an inflammatory effector T-cell phenotype. The resultant loss of lineage stability causes Tregs to lose their intratumor immune-suppressive function, making the tumor susceptible to killing by tumor-specific effector CD8+ T cells. Cancer Immunol Res; 1(5); 320–31. ©2013 AACR.
Data from A Paracrine Role for IL6 in Prostate Cancer Patients: Lack of Production by Primary or Metastatic Tumor Cells
Authors: Yu Shu-Han , Zheng Qizhi , Esopi David , Macgregor-Das Anne , Luo Jun , Antonarakis Emmanuel S. , Drake Charles G. , Vessella Robert , Morrissey Colm , De Marzo Angelo M. , Sfanos Karen S.
Published: April 2023

Abstract
Correlative human studies suggest that the pleiotropic cytokine IL6 contributes to the development and/or progression of prostate cancer. However, the source of IL6 production in the prostate microenvironment in patients has yet to be determined. The cellular origin of IL6 in primary and metastatic prostate cancer was examined in formalin-fixed, paraffin-embedded tissues using a highly sensitive and specific chromogenic in situ hybridization (CISH) assay that underwent extensive analytical validation. Quantitative RT-PCR showed that benign prostate tissues often had higher expression of IL6 mRNA than matched tumor specimens. CISH analysis further indicated that both primary and metastatic prostate adenocarcinoma cells do not express IL6 mRNA. IL6 expression was highly heterogeneous across specimens and was nearly exclusively restricted to the prostate stromal compartment—including endothelial cells and macrophages, among other cell types. The number of IL6-expressing cells correlated positively with the presence of acute inflammation. In metastatic disease, tumor cells were negative in all lesions examined, and IL6 expression was restricted to endothelial cells within the vasculature of bone metastases. Finally, IL6 was not detected in any cells in soft tissue metastases. These data suggest that, in prostate cancer patients, paracrine rather than autocrine IL6 production is likely associated with any role for the cytokine in disease progression. Cancer Immunol Res; 3(10); 1175–84. ©2015 AACR.
Data from Using Quantitative Seroproteomics to Identify Antibody Biomarkers in Pancreatic Cancer
Authors: Jhaveri Darshil T. , Kim Min-Sik , Thompson Elizabeth D. , Huang Lanqing , Sharma Rajni , Klein Alison P. , Zheng Lei , Le Dung T. , Laheru Daniel A. , Pandey Akhilesh , Jaffee Elizabeth M. , Anders Robert A.
Published: April 2023

Abstract
Pancreatic cancer is the fourth leading cause of cancer-related deaths in the United States. Less than 6% of patients survive beyond the fifth year due to inadequate early diagnostics and ineffective treatment options. Our laboratory has developed an allogeneic, granulocyte-macrophage colony-stimulating factor (GM-CSF)–secreting pancreatic cancer vaccine (GVAX) that has been tested in phase II clinical trials. Here, we employed a serum antibodies–based SILAC immunoprecipitation (SASI) approach to identify proteins that elicit an antibody response after vaccination. The SASI approach uses immunoprecipitation with patient-derived antibodies that is coupled to quantitative stable isotope–labeled amino acids in cell culture (SILAC). Using mass spectrometric analysis, we identified more than 150 different proteins that induce an antibody response after vaccination. The regulatory subunit 12A of protein phosphatase 1 (MYPT1 or PPP1R12A), regulatory subunit 8 of the 26S proteasome (PSMC5), and the transferrin receptor (TFRC) were shown to be pancreatic cancer–associated antigens recognized by postvaccination antibodies in the sera of patients with favorable disease-free survival after GVAX therapy. We further interrogated these proteins in over 80 GVAX-treated patients' pancreases and uniformly found a significant increase in the expression of MYPT1, PSMC5, and TFRC in neoplastic compared with non-neoplastic pancreatic ductal epithelium. We show that the novel SASI approach can identify antibody targets specifically expressed in patients with improved disease-free survival after cancer vaccine therapy. These targets need further validation to be considered as possible pancreatic cancer biomarkers. Cancer Immunol Res; 4(3); 225–33. ©2016 AACR.
Data from Decitabine Enhances Lymphocyte Migration and Function and Synergizes with CTLA-4 Blockade in a Murine Ovarian Cancer Model
Authors: Wang Lei , Amoozgar Zohreh , Huang Jing , Saleh Mohammad H. , Xing Deyin , Orsulic Sandra , Goldberg Michael S.
Published: April 2023

Abstract
The lack of second-line treatment for relapsed ovarian cancer necessitates the development of improved combination therapies. Targeted therapy and immunotherapy each confer clinical benefit, albeit limited as monotherapies. Ovarian cancer is not particularly responsive to immune checkpoint blockade, so combination with a complementary therapy may be beneficial. Recent studies have revealed that a DNA methyl transferase inhibitor, azacytidine, alters expression of immunoregulatory genes in ovarian cancer. In this study, the antitumor effects of a related DNA methyl transferase inhibitor, decitabine (DAC), were demonstrated in a syngeneic murine ovarian cancer model. Low-dose DAC treatment increases the expression of chemokines that recruit NK cells and CD8+ T cells, promotes their production of IFNγ and TNFα, and extends the survival of mice bearing subcutaneous or orthotopic tumors. While neither DAC nor immune checkpoint blockade confers durable responses as a monotherapy in this model, the efficacy of anti–CTLA-4 was potentiated by combination with DAC. This combination promotes differentiation of naïve T cells into effector T cells and prolongs cytotoxic lymphocyte responses as well as mouse survival. These results suggest that this combination therapy may be worthy of further consideration for improved treatment of drug-resistant ovarian cancer. Cancer Immunol Res; 3(9); 1030–41. ©2015 AACR.
Data from Altered Expression and Splicing of ESRP1 in Malignant Melanoma Correlates with Epithelial–Mesenchymal Status and Tumor-Associated Immune Cytolytic Activity
Authors: Yao Jun , Caballero Otavia L. , Huang Ying , Lin Calvin , Rimoldi Donata , Behren Andreas , Cebon Jonathan S. , Hung Mien-Chie , Weinstein John N. , Strausberg Robert L. , Zhao Qi
Published: April 2023

Abstract
Melanoma is one of the major cancer types for which new immune-based cancer treatments have achieved promising results. However, anti–PD-1 and anti–CTLA-4 therapies are effective only in some patients. Hence, predictive molecular markers for the development of clinical strategies targeting immune checkpoints are needed. Using The Cancer Genome Atlas (TCGA) RNAseq data, we found that expression of ESRP1, encoding a master splicing regulator in the epithelial–mesenchymal transition (EMT), was inversely correlated with tumor-associated immune cytolytic activity. That association holds up across multiple TCGA tumor types, suggesting a link between tumor EMT status and infiltrating lymphocyte activity. In melanoma, ESRP1 mainly exists in a melanocyte-specific truncated form transcribed from exon 13. This was validated by analyzing CCLE cell line data, public CAGE data, and RT-PCR in primary cultured melanoma cell lines. Based on ESRP1 expression, we divided TCGA melanoma cases into ESRP1-low, -truncated, and –full-length groups. ESRP1-truncated tumors comprise approximately two thirds of melanoma samples and reside in an apparent transitional state between epithelial and mesenchymal phenotypes. ESRP1 full-length tumors express epithelial markers and constitute about 5% of melanoma samples. In contrast, ESRP1-low tumors express mesenchymal markers and are high in immune cytolytic activity as well as PD-L2 and CTLA-4 expression. Those tumors are associated with better patient survival. Results from our study suggest a path toward the use of ESRP1 and other EMT markers as informative biomarkers for immunotherapy. Cancer Immunol Res; 4(6); 552–61. ©2016 AACR.
Data from Immunity to the Vacuolar ATPase Complex Accessory Unit ATP6S1 in Patients with Malignant Melanoma
Authors: Zhou Jun , Gupta Meghna , Wu Xinqi , Yoon Charles , Giobbie-Hurder Anita , Hodi F. Stephen
Published: April 2023

Abstract
The augmentation of high-titer antibodies to ATP6S1 is associated with favorable clinical outcomes in patients who received vaccination with autologous, irradiated tumor cells engineered to secrete GM-CSF and allogeneic bone marrow transplantation. Cellular immune responses to ATP6S1 are unknown. To define its role as an immune target, examination of cellular responses to ATP6S1 and immunity related to current therapies such as checkpoint blockade is needed. We used an overlapping peptide library representing the full-length ATP6S1 protein to screen for cellular responses from the peripheral blood of patients with stage III and IV melanoma. Reactive peptide pools were used to determine the individual peptide activity and epitopes. Recombinant ATP6S1 protein was used in an ELISA to assess potential correlation with humoral immune responses and changes in immunity related to CTLA-4 blockade with ipilimumab in these patients. We observed a broad array of CD4+ and CD8+ cellular responses against ATP6S1, including the identification of several MHC class I and II ATP6S1 epitopes. The generation of specific CD4+ and cytotoxic T cells revealed potent functional capability elicited by ipilimumab treatment in patients with metastatic melanoma, which revealed potent functional capability, including cytokine production, proliferation responsiveness to melanoma cell lines, and tumor-cell killing. Furthermore, the augmented humoral immune responses to ATP6S1 as a function of ipilimumab treatment were associated with beneficial clinical outcomes. These results support the continued development of ATP6S1 as a biomarker and therapeutic target. Cancer Immunol Res; 3(1); 59–67. ©2014 AACR.
Data from Neoantigen Load, Antigen Presentation Machinery, and Immune Signatures Determine Prognosis in Clear Cell Renal Cell Carcinoma
Authors: Matsushita Hirokazu , Sato Yusuke , Karasaki Takahiro , Nakagawa Tohru , Kume Haruki , Ogawa Seishi , Homma Yukio , Kakimi Kazuhiro
Published: April 2023

Abstract
Tumors commonly harbor multiple genetic alterations, some of which initiate tumorigenesis. Among these, some tumor-specific somatic mutations resulting in mutated protein have the potential to induce antitumor immune responses. To examine the relevance of the latter to immune responses in the tumor and to patient outcomes, we used datasets of whole-exome and RNA sequencing from 97 clear cell renal cell carcinoma (ccRCC) patients to identify neoepitopes predicted to be presented by each patient's autologous HLA molecules. We found that the number of nonsilent or missense mutations did not correlate with patient prognosis. However, combining the number of HLA-restricted neoepitopes with the cell surface expression of HLA or β2-microglobulin(β2M) revealed that an A-neohi/HLA-Ahi or ABC-neohi/β2Mhi phenotype correlated with better clinical outcomes. Higher expression of immune-related genes from CD8 T cells and their effector molecules [CD8A, perforin (PRF1) and granzyme A (GZMA)], however, did not correlate with prognosis. This may have been due to the observed correlation of these genes with the expression of other genes that were associated with immunosuppression in the tumor microenvironment (CTLA-4, PD-1, LAG-3, PD-L1, PD-L2, IDO1, and IL10). This suggested that abundant neoepitopes associated with greater antitumor effector immune responses were counterbalanced by a strongly immunosuppressive microenvironment. Therefore, immunosuppressive molecules should be considered high-priority targets for modulating immune responses in patients with ccRCC. Blockade of these molecular pathways could be combined with immunotherapies targeting neoantigens to achieve synergistic antitumor activity. Cancer Immunol Res; 4(5); 463–71. ©2016 AACR.
Data from 4-1BB–Enhanced Expansion of CD8+ TIL from Triple-Negative Breast Cancer Unveils Mutation-Specific CD8+ T Cells
Authors: Harao Michiko , Forget Marie-Andrée , Roszik Jason , Gao Hui , Babiera Gildy V. , Krishnamurthy Savitri , Chacon Jessica A. , Li Shumin , Mittendorf Elizabeth A. , DeSnyder Sarah M. , Rockwood Korrene F. , Bernatchez Chantale , Ueno Naoto T. , Radvanyi Laszlo G. , Vence Luis , Haymaker Cara , Reuben James M.
Published: April 2023

Abstract
Triple-negative breast cancer (TNBC) highly infiltrated with CD8+ tumor-infiltrating lymphocytes (TIL) has been associated with improved prognosis. This observation led us to hypothesize that CD8+ TIL could be utilized in autologous adoptive cell therapy for TNBC, although this concept has proven to be challenging, given the difficulty in expanding CD8+ TILs in solid cancers other than in melanoma. To overcome this obstacle, we used an agonistic antibody (urelumab) to a TNFR family member, 4-1BB/CD137, which is expressed by recently activated CD8+ T cells. This approach was first utilized in melanoma and, in this study, led to advantageous growth of TILs for the majority of TNBC tumors tested. The agonistic antibody was only added in the initial setting of the culture and yet favored the propagation of CD8+ TILs from TNBC tumors. These expanded CD8+ TILs were capable of cytotoxic functions and were successfully utilized to demonstrate the presence of immunogenic mutations in autologous TNBC tumor tissue without recognition of the wild-type counterpart. Our findings open the way for a successful adoptive immunotherapy for TNBC. Cancer Immunol Res; 5(6); 439–45. ©2017 AACR.
Data from Cell-free Tumor Microparticle Vaccines Stimulate Dendritic Cells via cGAS/STING Signaling
Authors: Zhang Huafeng , Tang Ke , Zhang Yi , Ma Ruihua , Ma Jingwei , Li Yong , Luo Shunqun , Liang Xiaoyu , Ji Tiantian , Gu Zhichao , Lu Jinzhi , He Wei , Cao Xuetao , Wan Yonghong , Huang Bo
Published: April 2023

Abstract
Tumor antigens and innate signals are vital considerations in developing new therapeutic or prophylactic antitumor vaccines. The role or requirement of intact tumor cells in the development of an effective tumor vaccine remains incompletely understood. This study reveals the mechanism by which tumor cell–derived microparticles (T-MP) can act as a cell-free tumor vaccine. Vaccinations with T-MPs give rise to prophylactic effects against the challenge of various tumor cell types, while T-MP–loaded dendritic cells (DC) also exhibit therapeutic effects in various tumor models. Such antitumor effects of T-MPs are perhaps attributable to their ability to generate immune signaling and to represent tumor antigens. Mechanically, T-MPs effectively transfer DNA fragments to DCs, leading to type I IFN production through the cGAS/STING-mediated DNA-sensing pathway. In turn, type I IFN promotes DC maturation and presentation of tumor antigens to T cells for antitumor immunity. These findings highlight a novel tumor cell-free vaccine strategy with potential clinical applications. Cancer Immunol Res; 3(2); 196–205. ©2014 AACR.
Data from VEGF Neutralization Plus CTLA-4 Blockade Alters Soluble and Cellular Factors Associated with Enhancing Lymphocyte Infiltration and Humoral Recognition in Melanoma
Authors: Wu Xinqi , Giobbie-Hurder Anita , Liao Xiaoyun , Lawrence Donald , McDermott David , Zhou Jun , Rodig Scott , Hodi F. Stephen
Published: April 2023

Abstract
Immune recognition of tumor targets by specific cytotoxic lymphocytes is essential for the effective rejection of tumors. A phase I clinical trial of ipilimumab (an antibody that blocks CTLA-4 function) in combination with bevacizumab (an antibody that inhibits angiogenesis) in patients with metastatic melanoma found favorable clinical outcomes were associated with increased tumor endothelial activation and lymphocyte infiltration. To better understand the underlying mechanisms, we sought features and factors that changed as a function of treatment in patients. Ipilimumab plus bevacizumab (Ipi-Bev) increased tumor vascular expression of ICAM1 and VCAM1. Treatment also altered concentrations of many circulating cytokines and chemokines, including increases of CXCL10, IL1α, TNFα, CXCL1, IFNα2, and IL8, with decreases in VEGF-A in most patients. IL1α and TNFα induced expression of E-selectin, CXCL1, and VCAM1 on melanoma tumor-associated endothelial cells (TEC) in vitro and promoted adhesion of activated T cells onto TEC. VEGFA inhibited TNFα-induced expression of ICAM1 and VCAM1 and T-cell adhesion, which was blocked by bevacizumab. CXCL10 promoted T-cell migration across TEC in vitro, was frequently expressed by melanoma cells, and was upregulated in a subset of tumors in treated patients. Robust upregulation of CXCL10 in tumors was accompanied by increased T-cell infiltration. Ipi-Bev also augmented humoral immune responses recognizing targets in melanoma, tumor endothelial, and tumor mesenchymal stem cells. Our findings suggest that Ipi-Bev therapy augments immune recognition in the tumor microenvironment through enhancing lymphocyte infiltration and antibody responses. IL1α, TNFα, and CXCL10, together with VEGF neutralization, contribute to Ipi-Bev–induced melanoma immune recognition. Cancer Immunol Res; 4(10); 858–68. ©2016 AACR.
Data from Tumor Subtype-Specific Cancer–Testis Antigens as Potential Biomarkers and Immunotherapeutic Targets for Cancers
Authors: Yao Jun , Caballero Otavia L. , Yung W.K. Alfred , Weinstein John N. , Riggins Gregory J. , Strausberg Robert L. , Zhao Qi
Published: April 2023

Abstract
Cancer–testis (CT) antigens are potential targets for cancer immunotherapy because of their restricted expression in immune-privileged germ cells and various malignancies. Current application of CT-based immunotherapy has been focused on CT expression–rich tumors such as melanoma and lung cancers. In this study, we surveyed CT expression using The Cancer Genome Atlas (TCGA) datasets for ten common cancer types. We show that CT expression is specific and enriched within certain cancer molecular subtypes. For example, HORMAD1, CXorf61, ACTL8, and PRAME are highly enriched in the basal subtype of breast cancer; MAGE and CSAG are most frequently activated in the magnoid subtype of lung adenocarcinoma; and PRAME is highly upregulated in the ccB subtype of clear cell renal cell carcinoma. Analysis of CT gene expression and DNA methylation indicates that some CTs are regulated epigenetically, whereas others are controlled primarily by tissue- and subtype-specific transcription factors. Our results suggest that although for some CT expression is associated with patient outcome, not many are independent prognostic markers. Thus, CTs with shared expression pattern are heterogeneous molecules with distinct activation modes and functional properties in different cancers and cancer subtypes. These data suggest a cancer subtype–orientated application of CT antigen as biomarkers and immunotherapeutic targets. Cancer Immunol Res; 2(4); 371–9. ©2013 AACR.
Data from IL2 Variant Circumvents ICOS+ Regulatory T-cell Expansion and Promotes NK Cell Activation
Authors: Sim Geok Choo , Liu Chengwen , Wang Ena , Liu Hui , Creasy Caitlin , Dai Zhimin , Overwijk Willem W. , Roszik Jason , Marincola Francesco , Hwu Patrick , Grimm Elizabeth , Radvanyi Laszlo
Published: April 2023

Abstract
Clinical responses to high-dose IL2 therapy are limited due to selective expansion of CD4+CD25+Foxp3+ T-regulatory cells (Treg), especially ICOS+ Tregs, rather than natural killer (NK) cells and effector T cells. These ICOS+ Tregs are highly suppressive and constitutively express high levels of IL2Rα (CD25) and CD39. Here, we characterized the effect of a mutant form of IL2 (F42K), which preferentially binds to the lower affinity IL2Rβγ with reduced binding to CD25, on Tregs, effector NK cells, and T-cell subsets. Unlike wild-type (WT) IL2, F42K did not efficiently induce the expansion of highly suppressive ICOS+ Tregs in peripheral blood mononuclear cells (PBMC) from healthy controls and melanoma patients. Instead, it promoted the expansion of CD16+CD56+ NK cells and CD56hiCD16− NK cell subsets in both short- and long-term cultures, with enhanced Bcl-2 expression. Stimulation of PBMCs with F42K induced expression of more NK cell activation molecules, such as NKp30, NKp44, DNAM-1, NKG2D, 4-1BB/CD137, and Tim-3, than WT IL2. F42K induced greater upregulation of TRAIL, and NK-mediated cytolytic activity was increased against both autologous and HLA-mismatched melanoma cells compared with WT IL2. Gene expression analysis revealed distinct gene expression profiles stimulated by F42K, WT IL2, and IL15. F42K therapy in vivo also induced a dramatic reduction in the expansion of ICOS+ Tregs, promoted NK cell expansion, and inhibited melanoma tumor growth more efficiently than WT IL2 and more effectively than anti–CTLA-4. Our findings suggest that F42K could be a potential substitute for WT IL2 as a cytokine therapy for cancer. Cancer Immunol Res; 4(11); 983–94. ©2016 AACR.
Data from NK Cell–Mediated Antitumor Effects of a Folate-Conjugated Immunoglobulin Are Enhanced by Cytokines
Authors: Jaime-Ramirez Alena C. , McMichael Elizabeth L. , Kondadasula SriVidya , Skinner Cassandra C. , Mundy-Bosse Bethany L. , Luedke Eric , Jones Natalie B. , Mani Aruna , Roda Julie , Karpa Volodymyr , Li Hong , Li Jilong , Elavazhagan Saranya , La Perle Krista M. , Schmitt Alessandra C. , Lu Yanhui , Zhang Xiaoli , Pan Xueliang , Mao Hsaioyin , Davis Melanie , Jarjoura David , Butchar Jonathan P. , Poi Ming , Phelps Mitch , Tridandapani Susheela , Byrd John C. , Caligiuri Michael A. , Lee Robert J. , Carson William E.
Published: April 2023

Abstract
Optimally effective antitumor therapies would not only activate immune effector cells but also engage them at the tumor. Folate conjugated to immunoglobulin (F-IgG) could direct innate immune cells with Fc receptors to folate receptor–expressing cancer cells. F-IgG bound to human KB and HeLa cells, as well as murine L1210JF, a folate receptor (FR)–overexpressing cancer cell line, as determined by flow cytometry. Recognition of F-IgG by natural killer (NK) cell Fc receptors led to phosphorylation of the ERK transcription factor and increased NK cell expression of CD69. Lysis of KB tumor cells by NK cells increased by about 5-fold after treatment with F-IgG, an effect synergistically enhanced by treatment with IL2, IL12, IL15, or IL21 (P < 0.001). F-IgG also enhanced the lysis of chronic lymphocytic leukemia cells by autologous NK cells. NK cells significantly increased production of IFNγ, MIP-1α, and RANTES in response to F-IgG–coated KB target cells in the presence of the NK cell–activating cytokine IL12, and these coculture supernatants induced significant T-cell chemotaxis (P < 0.001). F-IgG-coated targets also stimulated FcR-mediated monocyte effector functions. Studies in a murine leukemia model confirmed the intratumoral localization and antitumor activity of F-IgG, as well as enhancement of its effects by IL12 (P = 0.05). The antitumor effect of this combination was dependent on NK cells and led to decreased tumor cell proliferation in vivo. Thus, F-IgG can induce an immune response against FR-positive tumor cells that is mediated by NK cells and can be augmented by cytokine therapy. Cancer Immunol Res; 4(4); 323–36. ©2016 AACR.
Data from A Four-Factor Immunoscore System That Predicts Clinical Outcome for Stage II/III Gastric Cancer
Authors: Wen Ti , Wang Zhenning , Li Yi , Li Zhi , Che Xiaofang , Fan Yibo , Wang Shuo , Qu Jinglei , Yang Xianghong , Hou Kezuo , Zhou Wenyang , Xu Ling , Li Ce , Wang Jin , Liu Jing , Chen Liqun , Zhang Jingdong , Qu Xiujuan , Liu Yunpeng
Published: April 2023

Abstract
The American Joint Committee on Cancer (AJCC) staging system is insufficiently prognostic for operable gastric cancer patients; therefore, complementary factors are under intense investigation. Although the focus is on immune markers, the prognostic impact of a single immune factor is minimal, due to complex antitumor immune responses. A more comprehensive evaluation may engender more accurate predictions. We analyzed immune factors by immunohistochemical staining in two independent cohorts. The association with patients' survival was analyzed by the Kaplan–Meier method. Our immunoscore system was constructed using Cox proportional hazard analysis. PD-L1+ immune cells (IC), PD-L1+ tumor cells (TC), PD-1hi, and CD8More were found among 33.33%, 31.37%, 33.33%, and 49%, respectively, of patients from the discovery cohort, and 41.74%, 17.4%, 38.26%, and 30.43% from the validation cohort. PD-L1+ ICs and PD-1hi ICs correlated with poorer overall survival (OS), but PD-L1+ TCs correlated with better OS and clinical outcomes and infiltration of more CD8+ T cells. These four factors were independently prognostic after tumor/lymph nodes/metastasis (TNM) stage adjustment. An immunoscore system based on hazard ratios of the four factors further separated gastric cancer patients with similar TNM staging into low-, medium-, or high-risk groups, with significantly different survival. Our prognostic model yielded an area under the receiver operating characteristic curve (AUC) of 0.856 for prediction of mortality at 5 years, superior to that of TNM staging (AUC of 0.676). Thus, this more comprehensive immunoscore system can provide more accurate prognoses and is an essential complement to the AJCC staging system for operable gastric cancer patients. Cancer Immunol Res; 5(7); 524–34. ©2017 AACR.
Data from Development of Aggressive Pancreatic Ductal Adenocarcinomas Depends on Granulocyte Colony Stimulating Factor Secretion in Carcinoma Cells
Authors: Pickup Michael W. , Owens Philip , Gorska Agnieszka E. , Chytil Anna , Ye Fei , Shi Chanjuan , Weaver Valerie M. , Kalluri Raghu , Moses Harold L. , Novitskiy Sergey V.
Published: April 2023

Abstract
The survival rate for pancreatic ductal adenocarcinoma (PDAC) remains low. More therapeutic options to treat this disease are needed, for the current standard of care is ineffective. Using an animal model of aggressive PDAC (Kras/p48TGFβRIIKO), we discovered an effect of TGFβ signaling in regulation of G-CSF secretion in pancreatic epithelium. Elevated concentrations of G-CSF in PDAC promoted differentiation of Ly6G+ cells from progenitors, stimulated IL10 secretion from myeloid cells, and decreased T-cell proliferation via upregulation of Arg, iNOS, VEGF, IL6, and IL1b from CD11b+ cells. Deletion of csf3 in PDAC cells or use of a G-CSF–blocking antibody decreased tumor growth. Anti–G-CSF treatment in combination with the DNA synthesis inhibitor gemcitabine reduced tumor size, increased the number of infiltrating T cells, and decreased the number of Ly6G+ cells more effectively than gemcitabine alone. Human analysis of human datasets from The Cancer Genome Atlas and tissue microarrays correlated with observations from our mouse model experiments, especially in patients with grade 1, stage II disease. We propose that in aggressive PDAC, elevated G-CSF contributes to tumor progression through promoting increases in infiltration of neutrophil-like cells with high immunosuppressive activity. Such a mechanism provides an avenue for a neoadjuvant therapeutic approach for this devastating disease. Cancer Immunol Res; 5(9); 718–29. ©2017 AACR.
Data from PVRIG and PVRL2 Are Induced in Cancer and Inhibit CD8+ T-cell Function
Authors: Whelan Sarah , Ophir Eran , Kotturi Maya F. , Levy Ofer , Ganguly Sudipto , Leung Ling , Vaknin Ilan , Kumar Sandeep , Dassa Liat , Hansen Kyle , Bernados David , Murter Benjamin , Soni Abha , Taube Janis M. , Fader Amanda Nickles , Wang Tian-Li , Shih Ie-Ming , White Mark , Pardoll Drew M. , Liang Spencer C.
Published: April 2023

Abstract
Although checkpoint inhibitors that block CTLA-4 and PD-1 have improved cancer immunotherapies, targeting additional checkpoint receptors may be required to broaden patient response to immunotherapy. PVRIG is a coinhibitory receptor of the DNAM/TIGIT/CD96 nectin family that binds to PVRL2. We report that antagonism of PVRIG and TIGIT, but not CD96, increased CD8+ T-cell cytokine production and cytotoxic activity. The inhibitory effect of PVRL2 was mediated by PVRIG and not TIGIT, demonstrating that the PVRIG–PVRL2 pathway is a nonredundant signaling node. A combination of PVRIG blockade with TIGIT or PD-1 blockade further increased T-cell activation. In human tumors, PVRIG expression on T cells was increased relative to normal tissue and trended with TIGIT and PD-1 expression. Tumor cells coexpressing PVR and PVRL2 were observed in multiple tumor types, with highest coexpression in endometrial cancers. Tumor cells expressing either PVR or PVRL2 were also present in numbers that varied with the cancer type, with ovarian cancers having the highest percentage of PVR−PVRL2+ tumor cells and colorectal cancers having the highest percentage of PVR+PVRL2− cells. To demonstrate a role of PVRIG and TIGIT on tumor-derived T cells, we examined the effect of PVRIG and TIGIT blockade on human tumor-infiltrating lymphocytes. For some donors, blockade of PVRIG increased T-cell function, an effect enhanced by combination with TIGIT or PD-1 blockade. In summary, we demonstrate that PVRIG and PVRL2 are expressed in human cancers and the PVRIG–PVRL2 and TIGIT–PVR pathways are nonredundant inhibitory signaling pathways.

See related article on p. 244
Data from PD-L1 Binds to B7-1 Only In Cis on the Same Cell Surface
Authors: Chaudhri Apoorvi , Xiao Yanping , Klee Alyssa N. , Wang Xiaoxu , Zhu Baogong , Freeman Gordon J.
Published: April 2023

Abstract
Programmed death ligand 1 (PD-L1)–mediated immunosuppression regulates peripheral tolerance and is often co-opted by tumors to evade immune attack. PD-L1 binds to PD-1 but also binds to B7-1 (CD80) to regulate T-cell function. The binding interaction of PD-L1 with B7-1 and its functional role need further investigation to understand differences between PD-1 and PD-L1 tumor immunotherapy. We examined the molecular orientation of PD-L1 binding to B7-1 using cell-to-cell binding assays, ELISA, and flow cytometry. As expected, PD-L1–transfected cells bound to PD-1–transfected cells, and B7-1 cells bound to CD28 or CTLA-4–transfected cells; however, PD-L1 cells did not bind to B7-1 cells. By ELISA and flow cytometry with purified proteins, we found PD-L1 and B7-1 had a strong binding interaction only when PD-L1 was flexible. Soluble PD-1 and B7-1 competed for binding to PD-L1. Binding of native PD-L1 and B7-1 in cis on the same cell surface was demonstrated with NanoBiT proximity assays. Thus, PD-L1–B7-1 interaction can occur in cis on the same cell but not in trans between two cells, which suggests a model in which PD-L1 can bend via its 11-amino acid, flexible stalk to bind to B7-1 in cis, in a manner that can competitively block the binding of PD-L1 to PD-1 or of B7-1 to CD28. This binding orientation emphasizes the functional importance of coexpression of PD-L1 and B7-1 on the same cell. We found such coexpression on tumor-infiltrating myeloid cells. Our findings may help better utilize these pathways in cancer immunotherapy. Cancer Immunol Res; 6(8); 921–9. ©2018 AACR.
Data from Peptide Blocking of PD-1/PD-L1 Interaction for Cancer Immunotherapy
Authors: Li Chunlin , Zhang Nengpan , Zhou Jundong , Ding Chen , Jin Yaqing , Cui Xueyuan , Pu Kefeng , Zhu Yimin
Published: April 2023

Abstract
Immunotherapy has become a promising alternative therapeutic approach for cancer patients. Interruption of immune checkpoints, such as CTLA-4 and PD-1, has been verified to be a successful means for cancer therapy in clinical trials. mAb targeting PD-L1 has been approved to treat urothelial carcinoma, non–small cell lung cancer, or Merkel cell carcinoma by the FDA. However, the high cost of the antibody can limit its application. In our study, targeting PD-L1 peptide (TPP-1), which specifically binds to PD-L1 with high affinity, was identified through bacterial surface display methods. Using a T-cell activation assay and mixed lymphocyte reaction, TPP-1 was verified to interfere with the interaction of PD-1/PD-L1. To examine the inhibitory effect of TPP-1 on tumor growth in vivo, a xenograft mouse model using H460 cells was established. The growth rate of tumor masses in TPP-1 or PD-L1 antibody–treated mice was 56% or 71% lower than that in control peptide–treated mice, respectively, indicating that TPP-1 inhibits, or at least retards, tumor growth. IHC of the tumors showed that IFNγ and granzyme B expression increased in the TPP-1 or PD-L1 antibody–treated groups, indicating that TPP-1 attenuates the inhibitory effect of PD-L1 on T cells and that T cells may get reactivated. On the basis of our data, TPP-1 peptide could work as an alternative to antibodies for tumor immunotherapy. Cancer Immunol Res; 6(2); 178–88. ©2017 AACR.
Data from USP22 Deubiquitinates CD274 to Suppress Anticancer Immunity
Authors: Huang Xing , Zhang Qi , Lou Yu , Wang Junli , Zhao Xinyu , Wang Lin , Zhang Xiaozhen , Li Shanshan , Zhao Yulan , Chen Qi , Liang Tingbo , Bai Xueli
Published: April 2023

Abstract
PD-1 (CD279)–PD-L1 (CD274) inhibitory signaling is critical for cancer immune evasion, and thus has become one of the major targets in anticancer immunotherapy. There are several studies that demonstrate the potent effects of posttranslational modifications of CD274 on immune inactivation and suppression, such as ubiquitination, phosphorylation, glycosylation, and palmitoylation. However, the regulatory mechanisms for CD274 deubiquitination are still largely unclear. Here, we identified ubiquitin-specific protease 22 (USP22) as a novel deubiquitinase of CD274. USP22 directly interacted with the C terminus of CD274, inducing its deubiquitination and stabilization. Across multiple cancer types, USP22 was highly expressed and frequently altered in liver cancer, closely correlating with poor prognosis of these patients. Genetic depletion of USP22 inhibited liver cancer growth in an immune system–dependent manner, increased tumor immunogenicity and tumor-infiltrating lymphocytes, and improved therapeutic efficacy of CD274-targeted immunotherapy and CDDP-based chemotherapy in mice. We demonstrate that targeting USP22 is a promising strategy to potentiate anticancer immunity for CD274-amplified cancer.
Data from Assessing the Magnitude of Immunogenic Cell Death Following Chemotherapy and Irradiation Reveals a New Strategy to Treat Pancreatic Cancer
Authors: Ye Jian , Mills Bradley N. , Zhao Tony , Han Booyeon J. , Murphy Joseph D. , Patel Ankit P. , Johnston Carl J. , Lord Edith M. , Belt Brian A. , Linehan David C. , Gerber Scott A.
Published: April 2023

Abstract
Pancreatic ductal adenocarcinoma (PDAC) continues to have a dismal prognosis, in part, due to ineffective treatment strategies. The efficacy of some chemotherapies and especially radiotherapy is mediated partially by the immune system. Therefore, we hypothesized that profiling the immune response following chemotherapy and/or irradiation can be used as a readout for treatment efficacy but also to help identify optimal therapeutic schedules for PDAC. Using murine models of PDAC, we demonstrated that concurrent administration of stereotactic body radiotherapy (SBRT) and a modified dose of FOLFIRINOX (mFX) resulted in superior tumor control when compared with single or sequential treatment groups. Importantly, this combined treatment schedule enhanced the magnitude of immunogenic cell death, which in turn amplified tumor antigen presentation by dendritic cells and intratumoral CD8+ T-cell infiltration. Concurrent therapy also resulted in systemic immunity contributing to the control of established metastases. These findings provide a rationale for pursuing concurrent treatment schedules of SBRT with mFX in PDAC.
Data from IL15 Enhances CAR-T Cell Antitumor Activity by Reducing mTORC1 Activity and Preserving Their Stem Cell Memory Phenotype
Authors: Alizadeh Darya , Wong Robyn A. , Yang Xin , Wang Dongrui , Pecoraro Joseph R. , Kuo Cheng-Fu , Aguilar Brenda , Qi Yue , Ann David K. , Starr Renate , Urak Ryan , Wang Xiuli , Forman Stephen J. , Brown Christine E.
Published: April 2023

Abstract
Improvements in the quality and fitness of chimeric antigen receptor (CAR)-engineered T cells, through CAR design or manufacturing optimizations, could enhance the therapeutic potential of CAR-T cells. One parameter influencing the effectiveness of CAR-T cell therapy is the differentiation status of the final product: CAR-T cells that are less-differentiated and less exhausted are more therapeutically effective. In the current study, we demonstrate that CAR-T cells expanded in IL15 (CAR-T/IL15) preserve a less-differentiated stem cell memory (Tscm) phenotype, defined by expression of CD62L+CD45RA+ CCR7+, as compared with cells cultured in IL2 (CAR-T/IL2). CAR-T/IL15 cells exhibited reduced expression of exhaustion markers, higher antiapoptotic properties, and increased proliferative capacity upon antigen challenge. Furthermore, CAR-T/IL15 cells exhibited decreased mTORC1 activity, reduced expression of glycolytic enzymes and improved mitochondrial fitness. CAR-T/IL2 cells cultured in rapamycin (mTORC1 inhibitor) shared phenotypic features with CAR-T/IL15 cells, suggesting that IL15-mediated reduction of mTORC1 activity is responsible for preserving the Tscm phenotype. CAR-T/IL15 cells promoted superior antitumor responses in vivo in comparison with CAR-T/IL2 cells. Inclusion of cytokines IL7 and/or IL21 in addition to IL15 reduced the beneficial effects of IL15 on CAR-T phenotype and antitumor potency. Our findings show that IL15 preserves the CAR-T cell Tscm phenotype and improves their metabolic fitness, which results in superior in vivo antitumor activity, thus opening an avenue that may improve future adoptive T-cell therapies.
Data from Hypoxia-Induced VISTA Promotes the Suppressive Function of Myeloid-Derived Suppressor Cells in the Tumor Microenvironment
Authors: Deng Jie , Li Jiannan , Sarde Aurelien , Lines J. Louise , Lee Yu-Chi , Qian David C. , Pechenick Dov A. , Manivanh Richard , Le Mercier Isabelle , Lowrey Christopher H. , Varn Frederick S. , Cheng Chao , Leib David A. , Noelle Randolph J. , Mabaera Rodwell
Published: April 2023

Abstract
Tumor hypoxia is a negative prognostic factor that is implicated in oncogenic signal activation, immune escape, and resistance to treatment. Identifying the mechanistic role of hypoxia in immune escape and resistance to immune-checkpoint inhibitors may aid the identification of therapeutic targets. We and others have shown that V-domain Ig suppressor of T-cell activation (VISTA), a negative checkpoint regulator in the B7 family, is highly expressed in the tumor microenvironment in tumor models and primary human cancers. In this study, we show that VISTA and HIF1α activity are correlated in a cohort of colorectal cancer patients. High VISTA expression was associated with worse overall survival. We used the CT26 colon cancer model to investigate the regulation of VISTA by hypoxia. Compared with less hypoxic tumor regions or draining lymph nodes, regions of profound hypoxia in the tumor microenvironment were associated with increased VISTA expression on tumor-infiltrating myeloid-derived suppressor cells (MDSC). Using chromatin immunoprecipitation and genetic silencing, we show that hypoxia-inducible factor (HIF)-1α binding to a conserved hypoxia response element in the VISTA promoter upregulated VISTA on myeloid cells. Further, antibody targeting or genetic ablation of VISTA under hypoxia relieved MDSC-mediated T-cell suppression, revealing VISTA as a mediator of MDSC function. Collectively, these data suggest that targeting VISTA may mitigate the deleterious effects of hypoxia on antitumor immunity.
Data from Tumor-Specific Regulatory T Cells from the Bone Marrow Orchestrate Antitumor Immunity in Breast Cancer
Authors: Ge Yingzi , Böhm Hans-Henning , Rathinasamy Anchana , Xydia Maria , Hu Xiaoying , Pincha Mudita , Umansky Ludmila , Breyer Christopher , Hillier Michael , Bonertz Andreas , Sevko Alexandra , Domschke Christoph , Schuetz Florian , Frebel Helge , Dettling Steffen , Herold-Mende Christel , Reissfelder Christoph , Weitz Jürgen , Umansky Viktor , Beckhove Philipp
Published: April 2023

Abstract
Endogenous antitumor effector T-cell responses and immune-suppressive regulatory T cells (Treg) critically influence the prognosis of patients with cancer, yet many of the mechanisms of how this occurs remain unresolved. On the basis of an analysis of the function, antigen specificity, and distribution of tumor antigen–reactive T cells and Tregs in patients with breast cancer and transgenic mouse tumor models, we showed that tumor-specific Tregs were selectively activated in the bone marrow (BM) and egressed into the peripheral blood. The BM was constantly depleted of tumor-specific Tregs and was instead a site of increased induction and activity of tumor-reactive effector/memory T cells. Treg egress from the BM was associated with activation-induced expression of peripheral homing receptors such as CCR2. Because breast cancer tissues express the CCR2 ligand CCL2, the activation and egress of tumor antigen–specific Tregs in the BM resulted in the accumulation of Tregs in breast tumor tissue. Such immune compartmentalization and redistribution of T-cell subpopulations between the BM and peripheral tissues were achieved by vaccination with adenoviral vector–encoded TRP-2 tumor antigen in a RET transgenic mouse model of spontaneous malignant melanoma. Thus, the BM simultaneously represented a source of tumor-infiltrating Tregs and a site for the induction of endogenous tumor-specific effector T-cell responses, suggesting that both antitumor immunity and local immune suppression are orchestrated in the BM.
Data from LncRNA AK036396 Inhibits Maturation and Accelerates Immunosuppression of Polymorphonuclear Myeloid–Derived Suppressor Cells by Enhancing the Stability of Ficolin B
Authors: Tian Xinyu , Zheng Yu , Yin Kai , Ma Jie , Tian Jie , Zhang Yue , Mao Lingxiang , Xu Huaxi , Wang Shengjun
Published: April 2023

Abstract
Long noncoding RNAs (lncRNA) are emerging as crucial regulators of cell biology. However, the role of lncRNAs in the development and function of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC) remains unclear. Here, we identified that the lncRNA F730016J06Rik (AK036396) was highly expressed in PMN-MDSCs and that lncRNA AK036396 knockdown promoted the maturation and decreased the suppressive function of PMN-MDSCs. Ficolin B (Fcnb), the expression of which could be assessed as a surrogate for PMN-MDSC development, was the predicted target gene of lncRNA AK036396 based on microarray results. LncRNA AK036396 knockdown attenuated Fcnb protein stability in a manner dependent on the ubiquitin-proteasome system. Moreover, Fcnb inhibition downregulated the suppressive function of PMN-MDSCs. In addition, the expression of human M-ficolin, which is an ortholog of mouse Fcnb, was increased and positively correlated with arginase1 (ARG1) expression. This suppressive molecule is released by MDSCs, and its production is commonly used to represent the suppressive activity of MDSCs in patients with lung cancer, suggesting clinical relevance for these findings. These results indicate that lncRNA AK036396 can inhibit maturation and accelerate immunosuppression of PMN-MDSCs by enhancing Fcnb protein stability.