bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from T-cell Expression of IL10 Is Essential for Tumor Immune Surveillance in the Small Intestine
Authors: Dennis Kristen L. , Saadalla Abdulrahman , Blatner Nichole R. , Wang Shuya , Venkateswaran Vysak , Gounari Fotini , Cheroutre Hilde , Weaver Casey T. , Roers Axel , Egilmez Nejat K. , Khazaie Khashayarsha
Published: April 2023

Abstract
IL10 is attributed with immune-suppressive and anti-inflammatory properties, which could promote or suppress cancer in the gastrointestinal tract. Loss of IL10 exacerbates colonic inflammation, leading to colitis and cancer. Consistent with this, transfer of IL10-competent regulatory T cells (Treg) into mice with colitis or hereditary polyposis protects against disease, while IL10-deficient mice are predisposed to polyposis with increased colon polyp load. Little is known about the protective or pathogenic function of IL10 in cancers of the small intestine. We found CD4+ T cells and CD4+ Foxp3+ Tregs to be the major sources of IL10 in the small intestine and responsible for the increase in IL10 during polyposis in the APCΔ468 mouse model of hereditary polyposis. Targeted ablation of IL10 in T cells caused severe IL10 deficiency and delayed polyp growth. However, these polyps progressively lost cytotoxic activity and eventually progressed to cancer. Several observations suggested that the effect was due to the loss of IFNγ-dependent immune surveillance. IL10-incompetent CD4+ T cells failed to secrete IFNγ when stimulated with polyp antigens and were inefficient in T-helper-1 (TH1) commitment. By contrast, the TH17 commitment was unaffected. These findings were validated using mice whose T cells overexpress IL10. In these mice, we observed high intra-polyp cytotoxic activity and attenuation of polyposis. Thus, expression of IL10 by T cells is protective and required for immune surveillance in the small intestine. Cancer Immunol Res; 3(7); 806–14. ©2015 AACR.
Data from B7-H4 Expression by Nonhematopoietic Cells in the Tumor Microenvironment Promotes Antitumor Immunity
Authors: Rahbar Ramtin , Lin Albert , Ghazarian Magar , Yau Helen-Loo , Paramathas Sangeetha , Lang Philipp A. , Schildknecht Anita , Elford Alisha R. , Garcia-Batres Carlos , Martin Bernard , Berman Hal K. , Leong Wey L. , McCready David R. , Reedijk Michael , Done Susan J. , Miller Naomi , Youngson Bruce , Suh Woong-Kyung , Mak Tak W. , Ohashi Pamela S.
Published: April 2023

Abstract
The B7 family plays a critical role in both positive and negative regulation of immune responses by engaging a variety of receptors on lymphocytes. Importantly, blocking coinhibitory molecules using antibodies specific for CTLA-4 and PD-1 enhances tumor immunity in a subset of patients. Therefore, it is critical to understand the role of different B7 family members since they may be suitable therapeutic targets. B7-H4 is another member that inhibits T-cell function, and it is also upregulated on a variety of tumors and has been proposed to promote tumor growth. Here, we investigate the role of B7-H4 in tumor development and show that B7-H4 expression inhibits tumor growth in two mouse models. Furthermore, we show that B7-H4 expression is required for antitumor immune responses in a mouse model of mammary tumorigenesis. We found that the expression levels of B7-H4 correlate with MHC class I expression in both mouse and human samples. We show that IFNγ upregulates B7-H4 expression on mouse embryo fibroblasts and that the upregulation of B7-H4 on tumors is dependent on T cells. Notably, patients with breast cancer with increased B7-H4 expression show a prolonged time to recurrence. These studies demonstrate a positive role for B7-H4 in promoting antitumor immunity. Cancer Immunol Res; 3(2); 184–95. ©2014 AACR.
Data from Induction of NKG2D Ligands on Solid Tumors Requires Tumor-Specific CD8+ T Cells and Histone Acetyltransferases
Authors: Hu Jiemiao , Bernatchez Chantale , Zhang Liangfang , Xia Xueqing , Kleinerman Eugenie S. , Hung Mien-Chie , Hwu Patrick , Li Shulin
Published: April 2023

Abstract
NKG2D-mediated immune surveillance is crucial for inhibiting tumor growth and metastases. Malignant tumor cells often downregulate NKG2D ligands to escape from immune surveillance. High-profile studies have shown that restoring NKG2D ligand expression via genetic engineering inhibits tumor formation and progression. However, no effective in vivo approaches are available to restore these ligands across different types of solid tumors because the classic stress signal–dependent induction of this ligand in vitro is transient and has rarely been duplicated in solid tumors in vivo. We found that coadministration of an immune stimulatory signal (IL12) and chemotherapy (doxorubicin) restored the NKG2D ligand Rae-1 in multiple tumor types, including a human tumor model. The restored expression of NKG2D ligands was associated with tumor cell death and delay of tumor progression in vivo. Induction of tumor-specific NKG2D ligands required the engagement of CD8+ T cells and was regulated by the histone acetyltransferases GCN5 and PCAF. The tumor-specific restoration of NKG2D ligands in a variety of tumor models, including a human tumor model, resulted in NKG2D-dependent tumor regression and extended survival time. The elucidation of a CD8+ T cell–dependent mechanism suggests that activated NKG2D+CD8+ T-cell therapy alone may be able to restore the NKG2D ligand in tumors. Cancer Immunol Res; 5(4); 300–11. ©2017 AACR.
Data from Peptide/MHC Tetramer–Based Sorting of CD8+ T Cells to a Leukemia Antigen Yields Clonotypes Drawn Nonspecifically from an Underlying Restricted Repertoire
Authors: Hunsucker Sally A. , McGary Colleen S. , Vincent Benjamin G. , Enyenihi Atim A. , Waugh Jennifer P. , McKinnon Karen P. , Bixby Lisa M. , Ropp Patricia A. , Coghill James M. , Wood William A. , Gabriel Don A. , Sarantopoulos Stefanie , Shea Thomas C. , Serody Jonathan S. , Alatrash Gheath , Rodriguez-Cruz Tania , Lizée Gregory , Buntzman Adam S. , Frelinger Jeffrey A. , Glish Gary L. , Armistead Paul M.
Published: April 2023

Abstract
Testing of T cell–based cancer therapeutics often involves measuring cancer antigen–specific T-cell populations with the assumption that they arise from in vivo clonal expansion. This analysis, using peptide/MHC tetramers, is often ambiguous. From a leukemia cell line, we identified a CDK4-derived peptide epitope, UNC-CDK4-1 (ALTPVVVTL), that bound HLA-A*02:01 with high affinity and could induce CD8+ T-cell responses in vitro. We identified UNC-CDK4-1/HLA-A*02:01 tetramer+ populations in 3 of 6 patients with acute myeloid leukemia who had undergone allogeneic stem cell transplantation. Using tetramer-based, single-cell sorting and T-cell receptor β (TCRβ) sequencing, we identified recurrent UNC-CDK4-1 tetramer–associated TCRβ clonotypes in a patient with a UNC-CDK4-1 tetramer+ population, suggesting in vivo T-cell expansion to UNC-CDK4-1. In parallel, we measured the patient's TCRβ repertoire and found it to be highly restricted/oligoclonal. The UNC-CDK4-1 tetramer–associated TCRβ clonotypes represented >17% of the entire TCRβ repertoire—far in excess of the UNC-CDK4-1 tetramer+ frequency—indicating that the recurrent TCRβ clonotypes identified from UNC-CDK-4-1 tetramer+ cells were likely a consequence of the extremely constrained T-cell repertoire in the patient and not in vivo UNC-CDK4-1–driven clonal T-cell expansion. Mapping recurrent TCRβ clonotype sequences onto TCRβ repertoires can help confirm or refute antigen-specific T-cell expansion in vivo. Cancer Immunol Res; 3(3); 228–35. ©2015 AACR.
Data from Tumor-Associated Macrophages Promote Epigenetic Silencing of Gelsolin through DNA Methyltransferase 1 in Gastric Cancer Cells
Authors: Wang Hao-Chen , Chen Chin-Wang , Yang Chia-Lung , Tsai I-Min , Hou Ya-Chin , Chen Chang-Jung , Shan Yan-Shen
Published: April 2023

Abstract
Epigenetic repression of the tumor suppressor gelsolin (GSN) is frequently observed in cancers. Chronic inflammation can promote tumor progression via aberrant DNA methylation. In this study, we investigated the role of tumor-associated macrophages (TAMs) in DNA methylation of the GSN gene during gastric cancer progression. Immunofluorescence staining of 121 gastric cancer tissues showed aberrant localization of GSN and DNA methyltransferase 1 (DNMT1) and juxtaposition of DNMT1 and M2 TAMs. Decreased GSN protein and mRNA expression and increased DNA methylation in the GSN promoter were observed in gastric cancer cell lines and clinical specimens. To examine the effect of TAMs on DNA methylation in gastric cancer cells, we performed in vitro coculture assays and found increased DNMT1 expression but decreased GSN expression in gastric cancer cells after coculture with U937 cells. Knockdown of DNMT1 expression in gastric cancer cells could abort U937 coculture-mediated GSN downregulation. Meanwhile, CCL5 was the main chemokine upregulated in coculture medium. Treatment with CCL5 could induce DNMT1 expression in gastric cancer cells via STAT3 signaling. Inhibiting DNMT1 activity with procainamide, inhibiting DNA methylation with 5-AZA, or inhibiting CCL5/CCR5 signaling with maraviroc reduced tumor growth in vivo. In conclusion, upregulation of DNMT1 by CCL5/CCR5/STAT3 signaling is critical for TAM-mediated GSN silencing in gastric cancer. This study identified potential targets for gastric cancer therapy. Cancer Immunol Res; 5(10); 885–97. ©2017 AACR.
Data from YAP-Induced PD-L1 Expression Drives Immune Evasion in BRAFi-Resistant Melanoma
Authors: Kim Min Hwan , Kim Chang Gon , Kim Sang-Kyum , Shin Sang Joon , Choe Eun Ah , Park Su-Hyung , Shin Eui-Cheol , Kim Joon
Published: April 2023

Abstract
Activation of YAP, a Hippo pathway effector, is an important resistance mechanism to BRAF inhibitor (BRAFi) in melanoma. Emerging evidence also suggests that YAP is involved in suppression of the antitumor immune response. However, the potential direct impact of YAP activity on cytotoxic T-cell immune responses has not been explored yet. Here, we show that BRAFi-resistant melanoma cells evade CD8+ T-cell immune responses in a PD-L1–dependent manner by activating YAP, which synchronously supports melanoma cell survival upon BRAF inhibition. PD-L1 expression is elevated in BRAFi-resistant melanoma cells, in which YAP is robustly activated, and YAP knockdown decreases PD-L1 expression. In addition, constitutively active YAP (YAP-5SA) increases PD-L1 expression by binding to an upstream enhancer of the PD-L1 gene and potentiating its transcription. Both BRAFi-resistant and YAP-5SA–expressing melanoma cells suppress the cytotoxic function and cytokine production of Melan-A–specific CD8+ T cells, whereas anti–PD-1 antibody reverses the YAP-mediated T-cell suppression. Moreover, nuclear enrichment of YAP in clinical melanoma samples correlates with increased PD-L1 expression. Our findings show that YAP directly mediates evasion of cytotoxic T-cell immune responses in BRAFi-resistant melanoma cells by upregulating PD-L1, and targeting of YAP-mediated immune evasion may improve prognosis of melanoma patients. Cancer Immunol Res; 6(3); 255–66. ©2018 AACR.
Data from Endogenous CD4+ T Cells Recognize Neoantigens in Lung Cancer Patients, Including Recurrent Oncogenic KRAS and ERBB2 (Her2) Driver Mutations
Authors: Veatch Joshua R. , Jesernig Brenda L. , Kargl Julia , Fitzgibbon Matthew , Lee Sylvia M. , Baik Christina , Martins Renato , Houghton A. McGarry , Riddell Stanley R.
Published: April 2023

Abstract
T cells specific for neoantigens encoded by mutated genes in cancers are increasingly recognized as mediators of tumor destruction after immune-checkpoint inhibitor therapy or adoptive cell transfer. Much of the focus has been on identifying epitopes presented to CD8+ T cells by class I MHC. However, CD4+ class II MHC-restricted T cells have been shown to have an important role in antitumor immunity. Unfortunately, the vast majority of neoantigens recognized by CD8+ or CD4+ T cells in cancer patients result from random mutations and are patient-specific. Here, we screened the blood of 5 non–small cell lung cancer (NSCLC) patients for T-cell responses to candidate mutation-encoded neoepitopes. T-cell responses were detected to 8.8% of screened antigens, with 1 to 7 antigens identified per patient. A majority of responses were to random, patient-specific mutations. However, CD4+ T cells that recognized the recurrent KRASG12V and the ERBB2 (Her2) internal tandem duplication (ITD) oncogenic driver mutations, but not the corresponding wild-type sequences, were identified in two patients. Two different T-cell receptors (TCR) specific for KRASG12V and one T-cell receptor specific for Her2-ITD were isolated and conferred antigen specificity when transfected into T cells. Deep sequencing identified the Her2-ITD–specific TCR in the tumor but not nonadjacent lung. Our results showed that CD4+ T-cell responses to neoantigens, including recurrent driver mutations, can be derived from the blood of NSCLC patients. These data support the use of adoptive transfer or vaccination to augment CD4+ neoantigen-specific T cells and elucidate their role in human antitumor immunity.
Data from Semaphorin4D Inhibition Improves Response to Immune-Checkpoint Blockade via Attenuation of MDSC Recruitment and Function
Authors: Clavijo Paul E. , Friedman Jay , Robbins Yvette , Moore Ellen C. , Smith Ernest , Zauderer Maurice , Evans Elizabeth E. , Allen Clint T.
Published: April 2023

Abstract
Tumor infiltration by immunosuppressive myeloid cells, such as myeloid-derived suppressor cells (MDSCs), causes resistance to immunotherapy. Semaphorin4D, originally characterized for its axonal guidance properties, also contributes to endothelial cell migration and survival and modulates global immune cytokine profiles and myeloid cell polarization within the tumor microenvironment. Here, we show how a therapeutic murine Sema4D mAb improves responses to immune-checkpoint blockade (ICB) in two murine carcinoma models. Treatment of tumor-bearing mice with Sema4D mAb abrogated Ly6Ghi PMN-MDSC recruitment through reducing MAPK-dependent chemokine production by tumor cells in Murine oral cancer-1 (MOC1) tumors. PMN-MDSC suppressive capacity was reduced through inhibition of Sema4D-driven arginase expression. These changes led to enhanced tumor infiltration by CD8+ TIL and activation of tumor-draining lymph node T lymphocytes in response to tumor antigen. Sema4D mAb in combination with either CTLA-4 or PD-1 blockade enhanced rejection of tumors or tumor growth delay, resulting in prolonged survival with either treatment. This function of Sema4D mAb provides a rationale for its evaluation in combination with ICB to treat tumors with immunosuppressive myeloid infiltration.
Data from CD4+ T Cell and NK Cell Interplay Key to Regression of MHC Class Ilow Tumors upon TLR7/8 Agonist Therapy
Authors: Doorduijn Elien M. , Sluijter Marjolein , Salvatori Daniela C. , Silvestri Serenella , Maas Saskia , Arens Ramon , Ossendorp Ferry , van der Burg Sjoerd H. , van Hall Thorbald
Published: April 2023

Abstract
One of the next challenges in cancer immunotherapy is the resistance of tumors to T-cell–based treatments through loss of MHC class I. Here, we show that under these circumstances, the Toll-like receptor (TLR)-7/8 ligand imiquimod, but not the TLR3 ligand poly I:C or TLR9 ligand CpG, mediated an effective antitumor response. The rejection of these immune-escaped cancers was mediated by NK cells and CD4+ T cells, whereas activated CD8+ T cells were dispensable. Application of the innate immune stimulator at a distant site activated NK cells and thereby elicited tumor-specific T-cell responses in tumor-bearing mice. Mechanistically, imiquimod activated NK cells to kill tumor cells, resulting in release of tumor antigens and induction of tumor-specific CD4+ T cells. These T helper cells provoked a strong induction of CXCL9 and CXCL10 in the tumor environment. Simultaneously, imiquimod induced the expression of the cognate chemokine receptor CXCR3 on peripheral lymphocytes. This ignited intratumoral CD4+ T-cell infiltration and accumulation, which was critical for tumor rejection; CXCR3 blocking antibodies mitigated the clinical response. In the effector phase, NK cell recruitment to tumors and their activation depended on CD4+ T cells. Together, we have uncovered a potent immune axis of tumor-specific CD4+ T cells and NK cells that eliminates escaped MHC-Ilow tumors. Cancer Immunol Res; 5(8); 642–53. ©2017 AACR.
Data from TLR Stimulation during T-cell Activation Lowers PD-1 Expression on CD8+ T Cells
Authors: Zahm Christopher D. , Colluru Viswa T. , McIlwain Sean J. , Ong Irene M. , McNeel Douglas G.
Published: April 2023

Abstract
Expression of T-cell checkpoint receptors can compromise antitumor immunity. Blockade of these receptors, notably PD-1 and LAG-3, which become expressed during T-cell activation with vaccination, can improve antitumor immunity. We evaluated whether T-cell checkpoint expression could be separated from T-cell activation in the context of innate immune stimulation with TLR agonists. We found that ligands for TLR1/2, TLR7, and TLR9 led to a decrease in expression of PD-1 on antigen-activated CD8+ T cells. These effects were mediated by IL12 released by professional antigen-presenting cells. In two separate tumor models, treatment with antitumor vaccines combined with TLR1/2 or TLR7 ligands induced antigen-specific CD8+ T cells with lower PD-1 expression and improved antitumor immunity. These findings highlight the role of innate immune activation during effector T-cell development and suggest that at least one mechanism by which specific TLR agonists can be strategically used as vaccine adjuvants is by modulating the expression of PD-1 during CD8+ T-cell activation. Cancer Immunol Res; 6(11); 1364–74. ©2018 AACR.
Data from A STING Agonist Given with OX40 Receptor and PD-L1 Modulators Primes Immunity and Reduces Tumor Growth in Tolerized Mice
Authors: Foote Jeremy B. , Kok Marleen , Leatherman James M. , Armstrong Todd D. , Marcinkowski Bridget C. , Ojalvo Laureen S. , Kanne David B. , Jaffee Elizabeth M. , Dubensky Thomas W. , Emens Leisha A.
Published: April 2023

Abstract
Stimulator of interferon genes (STING) signaling induces IFNβ production by intratumoral dendritic cells (DC), driving T-cell priming and recruitment into the tumor microenvironment (TME). We examined to what extent preexisting antigen-specific tolerance influenced the efficacy of in situ delivery of a potent STING-activating cyclic dinucleotide (CDN), ADU S-100, against established HER-2+ breast tumors. ADU S-100 induced HER-2–specific CD8+ T-cell priming and durable tumor clearance in 100% of nontolerant parental FVB/N mice. In contrast, ADU S-100 did not sufficiently prime HER-2–specific CD8+ T cells in tolerant neu/N mice, resulting in only delayed tumor growth and tumor clearance in 10% of the mice. No differences in IFNβ production, DC priming, or HER-2–specific CD8+ T-cell trafficking were detected between FVB/N and neu/N mice. However, activation and expansion of HER-2–specific CD8+ T cells were defective in neu/N mice. Immune cell infiltrates of untreated tumor-bearing neu/N mice expressed high numbers of PD1 and OX40 receptors on their CD8+ T cells, and PD-L1 was highly expressed on both myeloid and tumor cells. Modulating PD-L1 and OX40 receptor signaling combined with intratumoral ADU S-100 administration enhanced HER-2–specific CD8+ T-cell activity, clearing tumors in 40% of neu/N mice. Thus, intratumoral STING agonists could potently prime tumor antigen–specific CD8+ T-cell responses, and adding PD-L1 blockade and OX40 receptor activation can overcome antigen-enforced immune tolerance to induce tumor regression. Cancer Immunol Res; 5(6); 468–79. ©2017 AACR.
Data from Regulatory T Cells in an Endogenous Mouse Lymphoma Recognize Specific Antigen Peptides and Contribute to Immune Escape
Authors: Ahmetlić Fatima , Riedel Tanja , Hömberg Nadine , Bauer Vera , Trautwein Nico , Geishauser Albert , Sparwasser Tim , Stevanović Stefan , Röcken Martin , Mocikat Ralph
Published: April 2023

Abstract
Foxp3+ regulatory T cells (Tregs) sustain immune homeostasis and may contribute to immune escape in malignant disease. As a prerequisite for developing immunologic approaches in cancer therapy, it is necessary to understand the ontogeny and the antigenic specificities of tumor-infiltrating Tregs. We addressed this question by using a λ-MYC transgenic mouse model of endogenously arising B-cell lymphoma, which mirrors key features of human Burkitt lymphoma. We show that Foxp3+ Tregs suppress antitumor responses in endogenous lymphoma. Ablation of Foxp3+ Tregs significantly delayed tumor development. The ratio of Treg to effector T cells was elevated in growing tumors, which could be ascribed to differential proliferation. The Tregs detected were mainly natural Tregs that apparently recognized self-antigens. We identified MHC class II–restricted nonmutated self-epitopes, which were more prevalent in lymphoma than in normal B cells and could be recognized by Tregs. These epitopes were derived from proteins that are associated with cellular processes related to malignancy and may be overexpressed in the tumor.
Data from Early-Life Microbiota Exposure Restricts Myeloid-Derived Suppressor Cell–Driven Colonic Tumorigenesis
Authors: Harusato Akihito , Viennois Emilie , Etienne-Mesmin Lucie , Matsuyama Shingo , Abo Hirohito , Osuka Satoru , Lukacs Nicholas W. , Naito Yuji , Itoh Yoshito , Li Jian-Dong , Merlin Didier , Gewirtz Andrew T. , Denning Timothy L.
Published: April 2023

Abstract
Gut microbiota and their metabolites are instrumental in regulating homeostasis at intestinal and extraintestinal sites. However, the complex effects of prenatal and early postnatal microbial exposure on adult health and disease outcomes remain incompletely understood. Here, we showed that mice raised under germ-free conditions until weaning and then transferred to specific pathogen-free (SPF) conditions harbored altered microbiota composition, augmented inflammatory cytokine and chemokine expression, and were hyper-susceptible to colitis-associated tumorigenesis later in adulthood. Increased number and size of colon tumors and intestinal epithelial cell proliferation in recolonized germ-free mice were associated with augmented intratumoral CXCL1, CXCL2, and CXCL5 expression and granulocytic myeloid-derived suppressor cell (G-MDSC) accumulation. Consistent with these findings, CXCR2 neutralization in recolonized germ-free mice completely reversed the exacerbated susceptibility to colitis-associated tumorigenesis. Collectively, our findings highlight a crucial role for early-life microbial exposure in establishing intestinal homeostasis that restrains colon cancer in adulthood.
Data from Soluble CD80 Protein Delays Tumor Growth and Promotes Tumor-Infiltrating Lymphocytes
Authors: Horn Lucas A. , Long Tiha M. , Atkinson Ryan , Clements Virginia , Ostrand-Rosenberg Suzanne
Published: April 2023

Abstract
Tumor cells use various immune-suppressive strategies to overcome antitumor immunity. One such method is tumor expression of programmed death ligand-1 (PD-L1), which triggers apoptotic death or anergy upon binding programmed death-1 (PD-1) on T cells. Our previous in vitro cellular studies with human and mouse PD-L1+ tumor cells demonstrated that a soluble form of the costimulatory molecule CD80 prevented PD-L1–mediated immune suppression and restored T-cell activation by binding PD-L1 and blocking interaction with PD-1. We now report that in vivo treatment of established syngeneic PD-L1+ CT26 colon carcinoma and B16F10 melanoma tumors with CD80-Fc delays tumor growth and promotes tumor-infiltrating T cells. Studies with PD-1−/− and CD28−/− mice demonstrate that soluble CD80 acts in vivo by simultaneously neutralizing PD-1 suppression and activating through CD28. We also report that soluble CD80 mediates its effects by activating transcription factors EGR1-4, NF-κB, and MAPK, downstream signaling components of the CD28 and T-cell receptor pathways. Soluble CD80 binds to CTLA-4 on activated human peripheral blood mononuclear cells. However, increasing quantities of CTLA-4 antagonist antibodies do not increase T-cell activation. These results indicate that soluble CD80 does not suppress T-cell function through CTLA-4 and suggest that CTLA-4 acts as a decoy receptor for CD80, rather than functioning as a suppressive signaling receptor. Collectively, these studies demonstrate that soluble CD80 has therapeutic efficacy in vivo in mouse tumor systems and that its effects are due to its ability to inhibit PD-1–mediated suppression while concurrently activating T cells through CD28. Cancer Immunol Res; 6(1); 59–68. ©2017 AACR.
Data from MICA-Expressing Monocytes Enhance Natural Killer Cell Fc Receptor-Mediated Antitumor Functions
Authors: Campbell Amanda R. , Duggan Megan C. , Suarez-Kelly Lorena P. , Bhave Neela , Opheim Kallan S. , McMichael Elizabeth L. , Trikha Prashant , Parihar Robin , Luedke Eric , Lewis Adrian , Yung Bryant , Lee Robert , Raulet David , Tridandapani Susheela , Groh Veronika , Yu Lianbo , Yildiz Vedat , Byrd John C. , Caligiuri Michael A. , Carson William E.
Published: April 2023

Abstract
Natural killer (NK) cells are large granular lymphocytes that promote the antitumor response via communication with other cell types in the tumor microenvironment. Previously, we have shown that NK cells secrete a profile of immune stimulatory factors (e.g., IFNγ, MIP-1α, and TNFα) in response to dual stimulation with the combination of antibody (Ab)-coated tumor cells and cytokines, such as IL12. We now demonstrate that this response is enhanced in the presence of autologous monocytes. Monocyte enhancement of NK cell activity was dependent on cell-to-cell contact as determined by a Transwell assay. It was hypothesized that NK cell effector functions against Ab-coated tumor cells were enhanced via binding of MICA on monocytes to NK cell NKG2D receptors. Strategies to block MICA–NKG2D interactions resulted in reductions in IFNγ production. Depletion of monocytes in vivo resulted in decreased IFNγ production by murine NK cells upon exposure to Ab-coated tumor cells. In mice receiving trastuzumab and IL12 therapy, monocyte depletion resulted in significantly greater tumor growth in comparison to mock-depleted controls (P < 0.05). These data suggest that NK cell–monocyte interactions enhance NK cell antitumor activity in the setting of monoclonal Ab therapy for cancer. Cancer Immunol Res; 5(9); 778–89. ©2017 AACR.
Data from Antagonism of IAPs Enhances CAR T-cell Efficacy
Authors: Michie Jessica , Beavis Paul A. , Freeman Andrew J. , Vervoort Stephin J. , Ramsbottom Kelly M. , Narasimhan Vignesh , Lelliott Emily J. , Lalaoui Najoua , Ramsay Robert G. , Johnstone Ricky W. , Silke John , Darcy Phillip K. , Voskoboinik Ilia , Kearney Conor J. , Oliaro Jane
Published: April 2023

Abstract
Chimeric antigen receptor (CAR) T-cell therapy has proven successful in the treatment of hematological malignancies, notably acute lymphoblastic leukemia and B-cell lymphoma. However, the efficacy of CAR T cells against solid tumors is poor, likely due to tumor-associated immunosuppression. Here, we demonstrated that antagonizing the “inhibitor of apoptosis proteins” with the clinical smac-mimetic, birinapant, significantly enhanced the antitumor activity of CAR T cells in a tumor necrosis factor (TNF)-dependent manner. Enhanced tumor cell death occurred independently of the perforin-mediated granule exocytosis pathway, underscoring the cytotoxic potential of CAR T-cell–derived TNF. Combining CAR T-cell therapy with birinapant significantly reduced established tumor growth in vivo, where either therapy alone was relatively ineffective. Using patient biopsy-derived tumoroids, we demonstrated the synergistic potential of combining CAR T-cell therapy with smac-mimetics. Taken together, we identified CAR T-cell–derived TNF as a potent antitumor effector, which can be further harnessed by smac-mimetics.
Data from Wnt3a Neutralization Enhances T-cell Responses through Indirect Mechanisms and Restrains Tumor Growth
Authors: Pacella Ilenia , Cammarata Ilenia , Focaccetti Chiara , Miacci Stefano , Gulino Alessandro , Tripodo Claudio , Ravà Micol , Barnaba Vincenzo , Piconese Silvia
Published: April 2023

Abstract
The Wnt/β-catenin pathway regulates T-cell functions, including the repression of effector functions to the advantage of memory development via Tcf1. In a companion study, we demonstrate that, in human cancers, Wnt3a/β-catenin signaling maintains tumor-infiltrating T cells in a partially exhausted status. Here, we have investigated the effects of Wnt3a neutralization in vivo in a mouse tumor model. Abundant Wnt3a was released, mostly by stromal cells, in the tumor microenvironment. We tested whether Wnt3a neutralization in vivo could rescue the effector capacity of tumor-infiltrating T cells, by administering an antibody to Wnt3a to tumor-bearing mice. This therapy restrained tumor growth and favored the expansion of tumor antigen–specific CD8+ effector memory T cells with increased expression of Tbet and IFNγ and reduced expression of Tcf1. However, the effect was not attributable to the interruption of T-cell–intrinsic β-catenin signaling, because Wnt3a/β-catenin activation correlated with enhanced, not reduced, T-cell effector functions both ex vivo and in vitro. Adoptively transferred CD8+ T cells, not directly exposed to the anti-Wnt3a antibody but infiltrating previously Wnt3a-neutralized tumors, also showed improved functions. The rescue of T-cell response was thus secondary to T-cell–extrinsic changes that likely involved dendritic cells. Indeed, tumor-derived Wnt3a strongly suppressed dendritic cell maturation in vitro, and anti-Wnt3a treatment rescued dendritic cell activities in vivo. Our results clarify the function of the Wnt3a/β-catenin pathway in antitumor effector T cells and suggest that Wnt3a neutralization might be a promising immunotherapy for rescuing dendritic cell activities. Cancer Immunol Res; 6(8); 953–64. ©2018 AACR.
Data from Combination Therapy Using Ruxolitinib and Oncolytic HSV Renders Resistant MPNSTs Susceptible to Virotherapy
Authors: Ghonime Mohammed G. , Cassady Kevin A.
Published: April 2023

Abstract
Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive soft-tissue sarcomas resistant to most cancer treatments. Surgical resection remains the primary treatment, but this is often incomplete, ultimately resulting in high mortality and morbidity rates. There has been a resurgence of interest in oncolytic virotherapy because of encouraging preclinical and clinical trial results. Oncolytic herpes simplex virus (oHSV) selectively replicates in cancer cells, lysing the cell and inducing antitumor immunity. We previously showed that basal interferon (IFN) signaling increases interferon-stimulated gene (ISG) expression, restricting viral replication in almost 50% of MPNSTs. The FDA-approved drug ruxolitinib (RUX) temporarily resets this constitutively active STAT signaling and renders the tumor cells susceptible to oHSV infection in cell culture. In the studies described here, we translated our in vitro results into a syngeneic MPNST tumor model. Consistent with our previous results, murine MPNSTs exhibit a similar IFN- and ISG-mediated oHSV-resistance mechanism, and virotherapy alone provides no antitumor benefit in vivo. However, pretreatment of mice with ruxolitinib reduced ISG expression, making the tumors susceptible to oHSV infection. Ruxolitinib pretreatment improved viral replication and altered the oHSV-induced immune-mediated response. Our results showed that this combination therapy increased CD8+ T-cell activation in the tumor microenvironment and that this population was indispensable for the antitumor benefit that follows from the combination of RUX and oHSV. These data suggest that JAK inhibition prior to oncolytic virus treatment augments both oHSV replication and the immunotherapeutic efficacy of oncolytic herpes virotherapy.
Data from High-Affinity GD2-Specific CAR T Cells Induce Fatal Encephalitis in a Preclinical Neuroblastoma Model
Authors: Richman Sarah A. , Nunez-Cruz Selene , Moghimi Babak , Li Lucy Z. , Gershenson Zachary T. , Mourelatos Zissimos , Barrett David M. , Grupp Stephan A. , Milone Michael C.
Published: April 2023

Abstract
The GD2 ganglioside, which is abundant on the surface of neuroblastoma cells, is targeted by an FDA-approved therapeutic monoclonal antibody and is an attractive tumor-associated antigen for cellular immunotherapy. Chimeric antigen receptor (CAR)–modified T cells can have potent antitumor activity in B-cell malignancies, and trials to harness this cytolytic activity toward GD2 in neuroblastoma are under way. In an effort to enhance the antitumor activity of CAR T cells that target GD2, we generated variant CAR constructs predicted to improve the stability and the affinity of the GD2-binding, 14G2a-based, single-chain variable fragment (scFv) of the CAR and compared their properties in vivo. We included the E101K mutation of GD2 scFv (GD2-E101K) that has enhanced antitumor activity against a GD2+ human neuroblastoma xenograft in vivo. However, this enhanced antitumor efficacy in vivo was concomitantly associated with lethal central nervous system (CNS) toxicity comprised of extensive CAR T-cell infiltration and proliferation within the brain and neuronal destruction. The encephalitis was localized to the cerebellum and basal regions of the brain that display low amounts of GD2. Our results highlight the challenges associated with target antigens that exhibit shared expression on critical normal tissues. Despite the success of GD2-specific antibody therapies in the treatment of neuroblastoma, the fatal neurotoxicity of GD2-specific CAR T-cell therapy observed in our studies suggests that GD2 may be a difficult target antigen for CAR T-cell therapy without additional strategies that can control CAR T-cell function within the CNS. Cancer Immunol Res; 6(1); 36–46. ©2017 AACR.
Data from Interleukin 33 Signaling Restrains Sporadic Colon Cancer in an Interferon-γ–Dependent Manner
Authors: Eissmann Moritz F. , Dijkstra Christine , Wouters Merridee A. , Baloyan David , Mouradov Dmitri , Nguyen Paul M. , Davalos-Salas Mercedes , Putoczki Tracy L. , Sieber Oliver M. , Mariadason John M. , Ernst Matthias , Masson Frederick
Published: April 2023

Abstract
Interleukin 33 (IL33) is an inflammatory cytokine released during necrotic cell death. The epithelium and stroma of the intestine express large amounts of IL33 and its receptor St2. IL33 is therefore continuously released during homeostatic turnover of the intestinal mucosa. Although IL33 can prevent colon cancer associated with inflammatory colitis, the contribution of IL33 signaling to sporadic colon cancer remains unknown. Here, we utilized a mouse model of sporadic colon cancer to investigate the contribution of IL33 signaling to tumorigenesis in the absence of preexisting inflammation. We demonstrated that genetic ablation of St2 enhanced colon tumor development. Conversely, administration of recombinant IL33 reduced growth of colon cancer cell allografts. In reciprocal bone marrow chimeras, the concurrent loss of IL33 signaling within radioresistant nonhematopoietic, and the radiosensitive hematopoietic, compartments was associated with increased tumor burden. We detected St2 expression within the radioresistant mesenchymal cell compartment of the colon whose stimulation with IL33 induced expression of bona fide NF-κB target genes. Mechanistically, we discovered that St2 deficiency within the nonhematopoietic compartment coincided with increased abundance of regulatory T cells and suppression of an IFNγ gene expression signature, whereas IL33 administration triggered IFNγ expression by tumor allograft-infiltrating T cells. The decrease of this IFNγ gene expression signature was associated with more aggressive disease in human colon cancer patients, suggesting that lack of IL33 signaling impaired the generation of a potent IFNγ-mediated antitumor immune response. Collectively, our data reveal that IL33 functions as a tumor suppressor in sporadic colon cancer. Cancer Immunol Res; 6(4); 409–21. ©2018 AACR.
Data from NK Cell–Specific CDK8 Deletion Enhances Antitumor Responses
Authors: Witalisz-Siepracka Agnieszka , Gotthardt Dagmar , Prchal-Murphy Michaela , Didara Zrinka , Menzl Ingeborg , Prinz Daniela , Edlinger Leo , Putz Eva Maria , Sexl Veronika
Published: April 2023

Abstract
Cyclin-dependent kinase 8 (CDK8) is a member of the transcription-regulating CDK family. CDK8 activates or represses transcription by associating with the mediator complex or by regulating transcription factors. Oncogenic activity of CDK8 has been demonstrated in several cancer types. Targeting CDK8 represents a potential therapeutic strategy. Because knockdown of CDK8 in a natural killer (NK) cell line enhances cytotoxicity and NK cells provide the first line of immune defense against transformed cells, we asked whether inhibiting CDK8 would improve NK-cell antitumor responses. In this study, we investigated the role of CDK8 in NK-cell function in vivo using mice with conditional ablation of CDK8 in NKp46+ cells (Cdk8fl/flNcr1Cre). Regardless of CDK8 expression, NK cells develop and mature normally in bone marrow and spleen. However, CDK8 deletion increased expression of the lytic molecule perforin, which correlated with enhanced NK-cell cytotoxicity in vitro. This translates into improved NK cell–mediated tumor surveillance in vivo in three independent models: B16F10 melanoma, v-abl+ lymphoma, and a slowly developing oncogene-driven leukemia. Our results thereby define a suppressive effect of CDK8 on NK-cell activity. Therapies that target CDK8 in cancer patients may enhance NK-cell responses against tumor cells. Cancer Immunol Res; 6(4); 458–66. ©2018 AACR.
Data from Macrophages and CD8+ T Cells Mediate the Antitumor Efficacy of Combined CD40 Ligation and Imatinib Therapy in Gastrointestinal Stromal Tumors
Authors: Zhang Jennifer Q. , Zeng Shan , Vitiello Gerardo A. , Seifert Adrian M. , Medina Benjamin D. , Beckman Michael J. , Loo Jennifer K. , Santamaria-Barria Juan , Maltbaek Joanna H. , Param Nesteene J. , Moral John A. , Zhao Julia N. , Balachandran Vinod , Rossi Ferdinand , Antonescu Cristina R. , DeMatteo Ronald P.
Published: April 2023

Abstract
Tyrosine kinase inhibition of gastrointestinal stromal tumors (GIST) is effective but typically culminates in resistance and is rarely curative. Immunotherapy has potential application to GIST, as we previously showed that T-cell checkpoint blockade increases the antitumor effects of imatinib. Here, we showed that ligation of CD40 using an agonistic antibody (anti-CD40) activated tumor-associated macrophages (TAMs) in vivo in a knock-in mouse model of GIST harboring a germline mutation in Kit exon 11. Activated TAMs had greater TNFα production and NFκB signaling and directly inhibited tumor cells in vitro. Anti-CD40 required concomitant therapy with imatinib for efficacy and depended on TAMs, and to a lesser extent CD8+ T cells, but not on CD4+ T cells or B cells. In an analysis of 50 human GIST specimens by flow cytometry, we found that CD40 was expressed on human TAMs and tumor cells yet was downregulated after response to imatinib. CD40 ligation did not have a direct inhibitory effect on human GIST cells. Our findings provide the rationale for combining anti-CD40 and tyrosine kinase inhibition to treat human GIST. Cancer Immunol Res; 6(4); 434–47. ©2018 AACR.
Data from Drug-Induced Senescent Multiple Myeloma Cells Elicit NK Cell Proliferation by Direct or Exosome-Mediated IL15 Trans-Presentation
Authors: Borrelli Cristiana , Ricci Biancamaria , Vulpis Elisabetta , Fionda Cinzia , Ricciardi Maria Rosaria , Petrucci Maria Teresa , Masuelli Laura , Peri Agnese , Cippitelli Marco , Zingoni Alessandra , Santoni Angela , Soriani Alessandra
Published: April 2023

Abstract
Treatment of multiple myeloma (MM) cells with sublethal doses of genotoxic drugs leads to senescence and results in increased NK cell recognition and effector functions. Herein, we demonstrated that doxorubicin- and melphalan-treated senescent cells display increased expression of IL15, a cytokine involved in NK cell activation, proliferation, and maturation. IL15 upregulation was evident at the mRNA and protein level, both in MM cell lines and malignant plasma cells from patients’ bone marrow (BM) aspirates. However, IL15 was detectable as a soluble cytokine only in vivo, thus indicating a functional role of IL15 in the BM tumor microenvironment. The increased IL15 was accompanied by enhanced expression of the IL15/IL15RA complex on the membrane of senescent myeloma cells, allowing the functional trans-presentation of this cytokine to neighboring NK cells, which consequently underwent activation and proliferation. We demonstrated that MM cell–derived exosomes, the release of which was augmented by melphalan treatment in senescent cells, also expressed IL15RA and IL15, and their interaction with NK cells in the presence of exogenous IL15 resulted in increased proliferation. Altogether, our data demonstrated that low doses of chemotherapeutic drugs, by inducing tumor cell senescence and a senescence-associated secretory phenotype, promoted IL15 trans-presentation to NK cells and, in turn, their activation and proliferation, thus enhancing NK cell–tumor immune surveillance and providing new insights for the exploitation of senescence-based cancer therapies. Cancer Immunol Res; 6(7); 860–9. ©2018 AACR.
Data from Targeting Cytokine Therapy to the Pancreatic Tumor Microenvironment Using PD-L1–Specific VHHs
Authors: Dougan Michael , Ingram Jessica R. , Jeong Hee-Jin , Mosaheb Munir M. , Bruck Patrick T. , Ali Lestat , Pishesha Novalia , Blomberg Olga , Tyler Paul M. , Servos Mariah M. , Rashidian Mohammad , Nguyen Quang-De , von Andrian Ulrich H. , Ploegh Hidde L. , Dougan Stephanie K.
Published: April 2023

Abstract
Cytokine-based therapies for cancer have not achieved widespread clinical success because of inherent toxicities. Treatment for pancreatic cancer is limited by the dense stroma that surrounds tumors and by an immunosuppressive tumor microenvironment. To overcome these barriers, we developed constructs of single-domain antibodies (VHHs) against PD-L1 fused with IL-2 and IFNγ. Targeting cytokine delivery in this manner reduced pancreatic tumor burden by 50%, whereas cytokines fused to an irrelevant VHH, or blockade of PD-L1 alone, showed little effect. Targeted delivery of IL-2 increased the number of intratumoral CD8+ T cells, whereas IFNγ reduced the number of CD11b+ cells and skewed intratumoral macrophages toward the display of M1-like characteristics. Imaging of fluorescent VHH–IFNγ constructs, as well as transcriptional profiling, demonstrated targeting of IFNγ to the tumor microenvironment. Many tumors and tumor-infiltrating myeloid cells express PD-L1, rendering them potentially susceptible to this form of targeted immunotherapy. Cancer Immunol Res; 6(4); 389–401. ©2018 AACR.
Data from Mast Cell–Dependent CD8+ T-cell Recruitment Mediates Immune Surveillance of Intestinal Tumors in ApcMin/+ Mice
Authors: Bodduluri Sobha R. , Mathis Steven , Maturu Paramahamsa , Krishnan Elangovan , Satpathy Shuchismita R. , Chilton Paula M. , Mitchell Thomas C. , Lira Sergio , Locati Massimo , Mantovani Alberto , Jala Venkatakrishna R. , Haribabu Bodduluri
Published: April 2023

Abstract
The presence of mast cells in some human colorectal cancers is a positive prognostic factor, but the basis for this association is incompletely understood. Here, we found that mice with a heterozygous mutation in the adenomatous polyposis coli gene (ApcMin/+) displayed reduced intestinal tumor burdens and increased survival in a chemokine decoy receptor, ACKR2-null background, which led to discovery of a critical role for mast cells in tumor defense. ACKR2–/–ApcMin/+ tumors showed increased infiltration of mast cells, their survival advantage was lost in mast cell–deficient ACKR2–/–SA–/–ApcMin/+ mice as the tumors grew rapidly, and adoptive transfer of mast cells restored control of tumor growth. Mast cells from ACKR2–/– mice showed elevated CCR2 and CCR5 expression and were also efficient in antigen presentation and activation of CD8+ T cells. Mast cell–derived leukotriene B4 (LTB4) was found to be required for CD8+ T lymphocyte recruitment, as mice lacking the LTB4 receptor (ACKR2–/–BLT1–/–ApcMin/+) were highly susceptible to intestinal tumor-induced mortality. Taken together, these data demonstrate that chemokine-mediated recruitment of mast cells is essential for initiating LTB4/BLT1-regulated CD8+ T-cell homing and generation of effective antitumor immunity against intestinal tumors. We speculate that the pathway reported here underlies the positive prognostic significance of mast cells in selected human tumors. Cancer Immunol Res; 6(3); 332–47. ©2018 AACR.
Data from Inhibition of the NKp44-PCNA Immune Checkpoint Using a mAb to PCNA
Authors: Kundu Kiran , Ghosh Susmita , Sarkar Rhitajit , Edri Avishay , Brusilovsky Michael , Gershoni-Yahalom Orly , Yossef Rami , Shemesh Avishai , Soria Jean-Charles , Lazar Vladimir , Joshua Ben-Zion , Campbell Kerry S. , Elkabets Moshe , Porgador Angel
Published: April 2023

Abstract
mAb-based blocking of the immune checkpoints involving the CTLA4-B7 and PD1-PDL1 inhibitory axes enhance T-cell–based adaptive immune responses in patients with cancer. We show here that antitumor responses by natural killer (NK) cells can be enhanced by a checkpoint-blocking mAb, 14-25-9, which we developed against proliferating cell nuclear antigen (PCNA). PCNA is expressed on the surface of cancer cells and acts as an inhibitory ligand for the NK-cell receptor, NKp44-isoform1. We tested for cytoplasmic- and membrane-associated PCNA by FACS- and ImageStream-based staining of cell lines and IHC of human cancer formalin fixed, paraffin embedded tissues. The mAb, 14-25-9, inhibited binding of chimeric NKp44 receptor to PCNA and mostly stained the cytoplasm and membrane of tumor cells, whereas commercial antibody (clone PC10) stained nuclear PCNA. NK functions were measured using ELISA-based IFNγ secretion assays and FACS-based killing assays. The NK92-NKp44-1 cell line and primary human NK cells showed increased IFNγ release upon coincubation with mAb 14-25-9 and various solid tumor cell lines and leukemias. Treatment with 14-25-9 also increased NK cytotoxic activity. In vivo efficacy was evaluated on patient-derived xenografts (PDX)-bearing NSG mice. In PDX-bearing mice, intravenous administration of mAb 14-25-9 increased degranulation (CD107a expression) of intratumorally injected patient autologous or allogeneic NK cells, as well as inhibited tumor growth when treated long term. Our study describes a mAb against the NKp44-PCNA innate immune checkpoint that can enhance NK-cell antitumor activity both in vitro and in vivo.
Data from T-cell Activity against AML Improved by Dual-Targeted T Cells Stimulated through T-cell and IL7 Receptors
Authors: Krawczyk Eric , Zolov Sergey N. , Huang Kevin , Bonifant Challice L.
Published: April 2023

Abstract
The development of engineered T cells to treat acute myeloid leukemia (AML) is challenging due to difficulty in target selection and the need for robust T-cell expansion and persistence. We designed a T cell stimulated to kill AML cells based on recognition of the AML-associated surface marker CLEC12A, via secretion of a CLEC12AxCD3 bispecific “engager” molecule (CLEC12A-ENG). CLEC12A-ENG T cells are specifically activated by CLEC12A, are not toxic to hematopoietic progenitor cells, and exhibit antigen-dependent AML killing. Next, we coupled stimulation of T-cell survival to triggering of a chimeric IL7 receptor with an ectodomain that binds a second AML-associated surface antigen, CD123. The resulting T cells, identified as CLEC12A-ENG.CD123IL7Rα T cells, demonstrate improved activation upon dual target recognition, kill AML, and exhibit antitumor activity in xenograft models. Enhanced T-cell activation conferred by CD123.IL7Rα was dependent both on recognition of the CD123 target and on IL7Rα-mediated downstream signaling. Expression of a chimeric IL7R targeted to a second tumor-associated antigen (TAA) should improve T-cell activity not only against hematologic malignancies, but perhaps against all cancers.