bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Cross-Species Single-Cell Analysis of Pancreatic Ductal Adenocarcinoma Reveals Antigen-Presenting Cancer-Associated Fibroblasts
Authors: Elyada Ela , Bolisetty Mohan , Laise Pasquale , Flynn William F. , Courtois Elise T. , Burkhart Richard A. , Teinor Jonathan A. , Belleau Pascal , Biffi Giulia , Lucito Matthew S. , Sivajothi Santhosh , Armstrong Todd D. , Engle Dannielle D. , Yu Kenneth H. , Hao Yuan , Wolfgang Christopher L. , Park Youngkyu , Preall Jonathan , Jaffee Elizabeth M. , Califano Andrea , Robson Paul , Tuveson David A.
Published: March 2024

Abstract
Cancer-associated fibroblasts (CAF) are major players in the progression and drug resistance of pancreatic ductal adenocarcinoma (PDAC). CAFs constitute a diverse cell population consisting of several recently described subtypes, although the extent of CAF heterogeneity has remained undefined. Here we use single-cell RNA sequencing to thoroughly characterize the neoplastic and tumor microenvironment content of human and mouse PDAC tumors. We corroborate the presence of myofibroblastic CAFs and inflammatory CAFs and define their unique gene signatures in vivo. Moreover, we describe a new population of CAFs that express MHC class II and CD74, but do not express classic costimulatory molecules. We term this cell population “antigen-presenting CAFs” and find that they activate CD4+ T cells in an antigen-specific fashion in a model system, confirming their putative immune-modulatory capacity. Our cross-species analysis paves the way for investigating distinct functions of CAF subtypes in PDAC immunity and progression.
Significance:
Appreciating the full spectrum of fibroblast heterogeneity in pancreatic ductal adenocarcinoma is crucial to developing therapies that specifically target tumor-promoting CAFs. This work identifies MHC class II–expressing CAFs with a capacity to present antigens to CD4+ T cells, and potentially to modulate the immune response in pancreatic tumors.

See related commentary by Belle and DeNardo, p. 1001.

This article is highlighted in the In This Issue feature, p. 983
Data from Immune Surveillance in Clinical Regression of Preinvasive Squamous Cell Lung Cancer
Authors: Pennycuick Adam , Teixeira Vitor H. , AbdulJabbar Khalid , Raza Shan E. Ahmed , Lund Tom , Akarca Ayse U. , Rosenthal Rachel , Kalinke Lukas , Chandrasekharan Deepak P. , Pipinikas Christodoulos P. , Lee-Six Henry , Hynds Robert E. , Gowers Kate H.C. , Henry Jake Y. , Millar Fraser R. , Hagos Yeman B. , Denais Celine , Falzon Mary , Moore David A. , Antoniou Sophia , Durrenberger Pascal F. , Furness Andrew J. , Carroll Bernadette , Marceaux Claire , Asselin-Labat Marie-Liesse , Larson William , Betts Courtney , Coussens Lisa M. , Thakrar Ricky M. , George Jeremy , Swanton Charles , Thirlwell Christina , Campbell Peter J. , Marafioti Teresa , Yuan Yinyin , Quezada Sergio A. , McGranahan Nicholas , Janes Sam M.
Published: March 2024

Abstract
Before squamous cell lung cancer develops, precancerous lesions can be found in the airways. From longitudinal monitoring, we know that only half of such lesions become cancer, whereas a third spontaneously regress. Although recent studies have described the presence of an active immune response in high-grade lesions, the mechanisms underpinning clinical regression of precancerous lesions remain unknown. Here, we show that host immune surveillance is strongly implicated in lesion regression. Using bronchoscopic biopsies from human subjects, we find that regressive carcinoma in situ lesions harbor more infiltrating immune cells than those that progress to cancer. Moreover, molecular profiling of these lesions identifies potential immune escape mechanisms specifically in those that progress to cancer: antigen presentation is impaired by genomic and epigenetic changes, CCL27–CCR10 signaling is upregulated, and the immunomodulator TNFSF9 is downregulated. Changes appear intrinsic to the carcinoma in situ lesions, as the adjacent stroma of progressive and regressive lesions are transcriptomically similar.
Significance:
Immune evasion is a hallmark of cancer. For the first time, this study identifies mechanisms by which precancerous lesions evade immune detection during the earliest stages of carcinogenesis and forms a basis for new therapeutic strategies that treat or prevent early-stage lung cancer.

See related commentary by Krysan et al., p. 1442.

This article is highlighted in the In This Issue feature, p. 1426
Data from Antitumor T-cell Homeostatic Activation Is Uncoupled from Homeostatic Inhibition by Checkpoint Blockade
Authors: Marshall Netonia , Hutchinson Keino , Marron Thomas U. , Aleynick Mark , Hammerich Linda , Upadhyay Ranjan , Svensson-Arvelund Judit , Brown Brian D. , Merad Miriam , Brody Joshua D.
Published: March 2024

Abstract
T-cell transfer into lymphodepleted recipients induces homeostatic activation and potentiates antitumor efficacy. In contrast to canonical T-cell receptor–induced activation, homeostatic activation yields a distinct phenotype and memory state whose regulatory mechanisms are poorly understood. Here, we show in patients and murine models that, following transfer into lymphodepleted bone marrow transplant (BMT) recipients, CD8+ T cells undergo activation but also simultaneous homeostatic inhibition manifested by upregulation of immune-checkpoint molecules and functional suppression. T cells transferred into BMT recipients were protected from homeostatic inhibition by PD-1/CTLA4 dual checkpoint blockade (dCB). This combination of dCB and BMT—”immunotransplant”—increased T-cell homeostatic activation and antitumor T-cell responses by an order of magnitude. Like homeostatic activation, homeostatic inhibition is IL7/IL15-dependent, revealing mechanistic coupling of these two processes. Marked similarity in ex vivo modulation of post-BMT T cells in mice and patients is promising for the clinical translation of immunotransplant (NCT03305445) and for addressing homeostatic inhibition in T-cell therapies.
Significance:
For optimal anticancer effect, T-cell therapies including chimeric antigen receptor T-cell, tumor-infiltrating lymphocyte, and transgenic T-cell therapies require transfer into lymphodepleted recipients and homeostatic activation; however, concomitant homeostatic inhibition mitigates T-cell therapies' efficacy. Checkpoint blockade uncouples homeostatic inhibition from activation, amplifying T-cell responses. Conversely, tumors nonresponsive to checkpoint blockade or BMT are treatable with immunotransplant.

See related commentary by Ansell, p. 1487.

This article is highlighted in the In This Issue feature, p. 1469
Data from The RNA m6A Reader YTHDF2 Maintains Oncogene Expression and Is a Targetable Dependency in Glioblastoma Stem Cells
Authors: Dixit Deobrat , Prager Briana C. , Gimple Ryan C. , Poh Hui Xian , Wang Yang , Wu Qiulian , Qiu Zhixin , Kidwell Reilly L. , Kim Leo J.Y. , Xie Qi , Vitting-Seerup Kristoffer , Bhargava Shruti , Dong Zhen , Jiang Li , Zhu Zhe , Hamerlik Petra , Jaffrey Samie R. , Zhao Jing Crystal , Wang Xiuxing , Rich Jeremy N.
Published: March 2024

Abstract
Glioblastoma is a universally lethal cancer driven by glioblastoma stem cells (GSC). Here, we interrogated N6-methyladenosine (m6A) mRNA modifications in GSCs by methyl RNA immunoprecipitation followed by sequencing and transcriptome analysis, finding transcripts marked by m6A often upregulated compared with normal neural stem cells (NSC). Interrogating m6A regulators, GSCs displayed preferential expression, as well as in vitro and in vivo dependency, of the m6A reader YTHDF2, in contrast to NSCs. Although YTHDF2 has been reported to destabilize mRNAs, YTHDF2 stabilized MYC and VEGFA transcripts in GSCs in an m6A-dependent manner. We identified IGFBP3 as a downstream effector of the YTHDF2–MYC axis in GSCs. The IGF1/IGF1R inhibitor linsitinib preferentially targeted YTHDF2-expressing cells, inhibiting GSC viability without affecting NSCs and impairing in vivo glioblastoma growth. Thus, YTHDF2 links RNA epitranscriptomic modifications and GSC growth, laying the foundation for the YTHDF2–MYC–IGFBP3 axis as a specific and novel therapeutic target in glioblastoma.
Significance:
Epitranscriptomics promotes cellular heterogeneity in cancer. RNA m6A landscapes of cancer and NSCs identified cell type–specific dependencies and therapeutic vulnerabilities. The m6A reader YTHDF2 stabilized MYC mRNA specifically in cancer stem cells. Given the challenge of targeting MYC, YTHDF2 presents a therapeutic target to perturb MYC signaling in glioblastoma.

This article is highlighted in the In This Issue feature, p. 211
Data from Phenotypic Mapping of Pathologic Cross-Talk between Glioblastoma and Innate Immune Cells by Synthetic Genetic Tracing
Authors: Schmitt Matthias Jürgen , Company Carlos , Dramaretska Yuliia , Barozzi Iros , Göhrig Andreas , Kertalli Sonia , Großmann Melanie , Naumann Heike , Sanchez-Bailon Maria Pilar , Hulsman Danielle , Glass Rainer , Squatrito Massimo , Serresi Michela , Gargiulo Gaetano
Published: March 2024

Abstract
Glioblastoma is a lethal brain tumor that exhibits heterogeneity and resistance to therapy. Our understanding of tumor homeostasis is limited by a lack of genetic tools to selectively identify tumor states and fate transitions. Here, we use glioblastoma subtype signatures to construct synthetic genetic tracing cassettes and investigate tumor heterogeneity at cellular and molecular levels, in vitro and in vivo. Through synthetic locus control regions, we demonstrate that proneural glioblastoma is a hardwired identity, whereas mesenchymal glioblastoma is an adaptive and metastable cell state driven by proinflammatory and differentiation cues and DNA damage, but not hypoxia. Importantly, we discovered that innate immune cells divert glioblastoma cells to a proneural-to-mesenchymal transition that confers therapeutic resistance. Our synthetic genetic tracing methodology is simple, scalable, and widely applicable to study homeostasis in development and diseases. In glioblastoma, the method causally links distinct (micro)environmental, genetic, and pharmacologic perturbations and mesenchymal commitment.
Significance:
Glioblastoma is heterogeneous and incurable. Here, we designed synthetic reporters to reflect the transcriptional output of tumor cell states and signaling pathways' activity. This method is generally applicable to study homeostasis in normal tissues and diseases. In glioblastoma, synthetic genetic tracing causally connects cellular and molecular heterogeneity to therapeutic responses.

This article is highlighted in the In This Issue feature, p. 521
Data from Surface Proteomics Reveals CD72 as a Target for In Vitro–Evolved Nanobody-Based CAR-T Cells in KMT2A/MLL1-Rearranged B-ALL
Authors: Nix Matthew A. , Mandal Kamal , Geng Huimin , Paranjape Neha , Lin Yu-Hsiu T. , Rivera Jose M. , Marcoulis Makeba , White Kristie L. , Whitman Jeffrey D. , Bapat Sagar P. , Parker Kevin R. , Ramirez Jonathan , Deucher Anne , Phojanokong Paul , Steri Veronica , Fattahi Faranak , Hann Byron C. , Satpathy Ansuman T. , Manglik Aashish , Stieglitz Elliot , Wiita Arun P.
Published: March 2024

Abstract
Alternative strategies are needed for patients with B-cell malignancy relapsing after CD19-targeted immunotherapy. Here, cell surface proteomics revealed CD72 as an optimal target for poor-prognosis KMT2A/MLL1-rearranged (MLLr) B-cell acute lymphoblastic leukemia (B-ALL), which we further found to be expressed in other B-cell malignancies. Using a recently described, fully in vitro system, we selected synthetic CD72-specific nanobodies, incorporated them into chimeric antigen receptors (CAR), and demonstrated robust activity against B-cell malignancy models, including CD19 loss. Taking advantage of the role of CD72 in inhibiting B-cell receptor signaling, we found that SHIP1 inhibition increased CD72 surface density. We establish that CD72-nanobody CAR-T cells are a promising therapy for MLLr B-ALL.
Significance:
Patients with MLLr B-ALL have poor prognoses despite recent immunotherapy advances. Here, surface proteomics identifies CD72 as being enriched on MLLr B-ALL but also widely expressed across B-cell cancers. We show that a recently described, fully in vitro nanobody platform generates binders highly active in CAR-T cells and demonstrate its broad applicability for immunotherapy development.

This article is highlighted in the In This Issue feature, p. 1861
Data from Parallel Genomic Alterations of Antigen and Payload Targets Mediate Polyclonal Acquired Clinical Resistance to Sacituzumab Govitecan in Triple-Negative Breast Cancer
Authors: Coates James T. , Sun Sheng , Leshchiner Ignaty , Thimmiah Nayana , Martin Elizabeth E. , McLoughlin Daniel , Danysh Brian P. , Slowik Kara , Jacobs Raquel A. , Rhrissorrakrai Kahn , Utro Filippo , Levovitz Chaya , Denault Elyssa , Walmsley Charlotte S. , Kambadakone Avinash , Stone James R. , Isakoff Steven J. , Parida Laxmi , Juric Dejan , Getz Gad , Bardia Aditya , Ellisen Leif W.
Published: March 2024

Abstract
Sacituzumab govitecan (SG), the first antibody–drug conjugate (ADC) approved for triple-negative breast cancer, incorporates the anti-TROP2 antibody hRS7 conjugated to a topoisomerase-1 (TOP1) inhibitor payload. We sought to identify mechanisms of SG resistance through RNA and whole-exome sequencing of pretreatment and postprogression specimens. One patient exhibiting de novo progression lacked TROP2 expression, in contrast to robust TROP2 expression and focal genomic amplification of TACSTD2/TROP2 observed in a patient with a deep, prolonged response to SG. Analysis of acquired genomic resistance in this case revealed one phylogenetic branch harboring a canonical TOP1E418K resistance mutation and subsequent frameshift TOP1 mutation, whereas a distinct branch exhibited a novel TACSTD2/TROP2T256R missense mutation. Reconstitution experiments demonstrated that TROP2T256R confers SG resistance via defective plasma membrane localization and reduced cell-surface binding by hRS7. These findings highlight parallel genomic alterations in both antibody and payload targets associated with resistance to SG.
Significance:
These findings underscore TROP2 as a response determinant and reveal acquired SG resistance mechanisms involving the direct antibody and drug payload targets in distinct metastatic subclones of an individual patient. This study highlights the specificity of SG and illustrates how such mechanisms will inform therapeutic strategies to overcome ADC resistance.

This article is highlighted in the In This Issue feature, p. 2355
Data from CARM1 Inhibition Enables Immunotherapy of Resistant Tumors by Dual Action on Tumor Cells and T Cells
Authors: Kumar Sushil , Zeng Zexian , Bagati Archis , Tay Rong En , Sanz Lionel A. , Hartono Stella R. , Ito Yoshinaga , Abderazzaq Fieda , Hatchi Elodie , Jiang Peng , Cartwright Adam N.R. , Olawoyin Olamide , Mathewson Nathan D. , Pyrdol Jason W. , Li Mamie Z. , Doench John G. , Booker Matthew A. , Tolstorukov Michael Y. , Elledge Stephen J. , Chédin Frédéric , Liu X. Shirley , Wucherpfennig Kai W.
Published: March 2024

Abstract
A number of cancer drugs activate innate immune pathways in tumor cells but unfortunately also compromise antitumor immune function. We discovered that inhibition of CARM1, an epigenetic enzyme and cotranscriptional activator, elicited beneficial antitumor activity in both cytotoxic T cells and tumor cells. In T cells, Carm1 inactivation substantially enhanced their antitumor function and preserved memory-like populations required for sustained antitumor immunity. In tumor cells, Carm1 inactivation induced a potent type 1 interferon response that sensitized resistant tumors to cytotoxic T cells. Substantially increased numbers of dendritic cells, CD8 T cells, and natural killer cells were present in Carm1-deficient tumors, and infiltrating CD8 T cells expressed low levels of exhaustion markers. Targeting of CARM1 with a small molecule elicited potent antitumor immunity and sensitized resistant tumors to checkpoint blockade. Targeting of this cotranscriptional regulator thus offers an opportunity to enhance immune function while simultaneously sensitizing resistant tumor cells to immune attack.
Significance:
Resistance to cancer immunotherapy remains a major challenge. Targeting of CARM1 enables immunotherapy of resistant tumors by enhancing T-cell functionality and preserving memory-like T-cell populations within tumors. CARM1 inhibition also sensitizes resistant tumor cells to immune attack by inducing a tumor cell–intrinsic type 1 interferon response.

This article is highlighted in the In This Issue feature, p. 1861
Data from Cancer Cells Retrace a Stepwise Differentiation Program during Malignant Progression
Authors: Saghafinia Sadegh , Homicsko Krisztian , Di Domenico Annunziata , Wullschleger Stephan , Perren Aurel , Marinoni Ilaria , Ciriello Giovanni , Michael Iacovos P. , Hanahan Douglas
Published: March 2024

Abstract
Pancreatic neuroendocrine tumors (PanNET) comprise two molecular subtypes, relatively benign islet tumors (IT) and invasive, metastasis-like primary (MLP) tumors. Until now, the origin of aggressive MLP tumors has been obscure. Herein, using multi-omics approaches, we revealed that MLP tumors arise from IT via dedifferentiation following a reverse trajectory along the developmental pathway of islet β cells, which results in the acquisition of a progenitor-like molecular phenotype. Functionally, the miR-181cd cluster induces the IT-to-MLP transition by suppressing expression of the Meis2 transcription factor, leading to upregulation of a developmental transcription factor, Hmgb3. Notably, the IT-to-MLP transition constitutes a distinct step of tumorigenesis and is separable from the classic proliferation-associated hallmark, temporally preceding accelerated proliferation of cancer cells. Furthermore, patients with PanNET with elevated HMGB3 expression and an MLP transcriptional signature are associated with higher-grade tumors and worse survival. Overall, our results unveil a new mechanism that modulates cancer cell plasticity to enable malignant progression.
Significance:
Dedifferentiation has long been observed as a histopathologic characteristic of many cancers, albeit inseparable from concurrent increases in cell proliferation. Herein, we demonstrate that dedifferentiation is a mechanistically and temporally separable step in the multistage tumorigenesis of pancreatic islet cells, retracing the developmental lineage of islet β cells.

This article is highlighted in the In This Issue feature, p. 2355
Data from CDK4/6 Inhibition Promotes Antitumor Immunity through the Induction of T-cell Memory
Authors: Lelliott Emily J. , Kong Isabella Y. , Zethoven Magnus , Ramsbottom Kelly M. , Martelotto Luciano G. , Meyran Deborah , Zhu Joe Jiang , Costacurta Matteo , Kirby Laura , Sandow Jarrod J. , Lim Lydia , Dominguez Pilar M. , Todorovski Izabela , Haynes Nicole M. , Beavis Paul A. , Neeson Paul J. , Hawkins Edwin D. , McArthur Grant A. , Parish Ian A. , Johnstone Ricky W. , Oliaro Jane , Sheppard Karen E. , Kearney Conor J. , Vervoort Stephin J.
Published: March 2024

Abstract
Pharmacologic inhibitors of cyclin-dependent kinases 4 and 6 (CDK4/6) are an approved treatment for hormone receptor–positive breast cancer and are currently under evaluation across hundreds of clinical trials for other cancer types. The clinical success of these inhibitors is largely attributed to well-defined tumor-intrinsic cytostatic mechanisms, whereas their emerging role as immunomodulatory agents is less understood. Using integrated epigenomic, transcriptomic, and proteomic analyses, we demonstrated a novel action of CDK4/6 inhibitors in promoting the phenotypic and functional acquisition of immunologic T-cell memory. Short-term priming with a CDK4/6 inhibitor promoted long-term endogenous antitumor T-cell immunity in mice, enhanced the persistence and therapeutic efficacy of chimeric antigen receptor T cells, and induced a retinoblastoma-dependent T-cell phenotype supportive of favorable responses to immune checkpoint blockade in patients with melanoma. Together, these mechanistic insights significantly broaden the prospective utility of CDK4/6 inhibitors as clinical tools to boost antitumor T-cell immunity.
Significance:
Immunologic memory is critical for sustained antitumor immunity. Our discovery that CDK4/6 inhibitors drive T-cell memory fate commitment sheds new light on their clinical activity, which is essential for the design of clinical trial protocols incorporating these agents, particularly in combination with immunotherapy, for the treatment of cancer.

This article is highlighted in the In This Issue feature, p. 2355
Data from PTHrP Drives Pancreatic Cancer Growth and Metastasis and Reveals a New Therapeutic Vulnerability
Authors: Pitarresi Jason R. , Norgard Robert J. , Chiarella Anna M. , Suzuki Kensuke , Bakir Basil , Sahu Varun , Li Jinyang , Zhao Jun , Marchand Benoît , Wengyn Maximilian D. , Hsieh Antony , Kim Il-Kyu , Zhang Amy , Sellin Karine , Lee Vivian , Takano Shigetsugu , Miyahara Yoji , Ohtsuka Masayuki , Maitra Anirban , Notta Faiyaz , Kremer Richard , Stanger Ben Z. , Rustgi Anil K.
Published: March 2024

Abstract
Pancreatic cancer metastasis is a leading cause of cancer-related deaths, yet very little is understood regarding the underlying biology. As a result, targeted therapies to inhibit metastasis are lacking. Here, we report that the parathyroid hormone–related protein (PTHrP encoded by PTHLH) is frequently amplified as part of the KRAS amplicon in patients with pancreatic cancer. PTHrP upregulation drives the growth of both primary and metastatic tumors in mice and is highly enriched in pancreatic ductal adenocarcinoma metastases. Loss of PTHrP—either genetically or pharmacologically—dramatically reduces tumor burden, eliminates metastasis, and enhances overall survival. These effects are mediated in part through a reduction in epithelial-to-mesenchymal transition, which reduces the ability of tumor cells to initiate metastatic cascade. Spp1, which encodes osteopontin, is revealed to be a downstream effector of PTHrP. Our results establish a new paradigm in pancreatic cancer whereby PTHrP is a driver of disease progression and emerges as a novel therapeutic vulnerability.
Significance:
Pancreatic cancer often presents with metastases, yet no strategies exist to pharmacologically inhibit this process. Herein, we establish the oncogenic and prometastatic roles of PTHLH, a novel amplified gene in pancreatic ductal adenocarcinoma. We demonstrate that blocking PTHrP activity reduces primary tumor growth, prevents metastasis, and prolongs survival in mice.

This article is highlighted in the In This Issue feature, p. 1601
Data from High-Throughput Stability Screening of Neoantigen/HLA Complexes Improves Immunogenicity Predictions
Authors: Blaha Dylan T. , Anderson Scott D. , Yoakum Daniel M. , Hager Marlies V. , Zha Yuanyuan , Gajewski Thomas F. , Kranz David M.
Published: March 2024

Abstract
Mutated peptides (neoantigens) from a patient's cancer genome can serve as targets for T-cell immunity, but identifying which peptides can be presented by an MHC molecule and elicit T cells has been difficult. Although algorithms that predict MHC binding exist, they are not yet able to distinguish experimental differences in half-lives of the complexes (an immunologically relevant parameter, referred to here as kinetic stability). Improvement in determining actual neoantigen peptide/MHC stability could be important, as only a small fraction of peptides in most current vaccines are capable of eliciting CD8+ T-cell responses. Here, we used a rapid, high-throughput method to experimentally determine peptide/HLA thermal stability on a scale that will be necessary for analysis of neoantigens from thousands of patients. The method combined the use of UV-cleavable peptide/HLA class I complexes and differential scanning fluorimetry to determine the Tm values of neoantigen complexes. Measured Tm values were accurate and reproducible and were directly proportional to the half-lives of the complexes. Analysis of known HLA-A2–restricted immunogenic peptides showed that Tm values better correlated with immunogenicity than algorithm-predicted binding affinities. We propose that temperature stability information can be used as a guide for the selection of neoantigens in cancer vaccines in order to focus attention on those mutated peptides with the highest probability of being expressed on the cell surface.
Data from Regression of Metastatic Merkel Cell Carcinoma Following Transfer of Polyomavirus-Specific T Cells and Therapies Capable of Reinducing HLA Class-I
Authors: Chapuis Aude G. , Afanasiev Olga K. , Iyer Jayasri G. , Paulson Kelly G. , Parvathaneni Upendra , Hwang Joo Ha , Lai Ivy , Roberts Ilana M. , Sloan Heather L. , Bhatia Shailender , Shibuya Kendall C. , Gooley Ted , Desmarais Cindy , Koelle David M. , Yee Cassian , Nghiem Paul
Published: March 2024

Abstract
Merkel cell carcinoma (MCC) is an aggressive skin cancer that typically requires the persistent expression of Merkel cell polyomavirus (MCPyV) oncoproteins that can serve as ideal immunotherapeutic targets. Several immune evasion mechanisms are active in MCC, including downregulation of HLA class-I expression on tumor cells and dysfunctional endogenous MCPyV-specific CD8 T-cell responses. To overcome these obstacles, we combined local and systemic immune therapies in a 67-year-old man, who developed metastatic MCPyV-expressing MCC. Intralesional IFN-β-1b or targeted single-dose radiation was administered as a preconditioning strategy to reverse the downregulation of HLA-I expression noted in his tumors and to facilitate the subsequent recognition of tumor cells by T cells. This was followed by the adoptive transfer of ex vivo expanded polyclonal, polyomavirus-specific T cells as a source of reactive antitumor immunity. The combined regimen was well tolerated and led to persistent upregulation of HLA-I expression in the tumor and a durable complete response in two of three metastatic lesions. Relative to historical controls, the patient experienced a prolonged period without development of additional distant metastases (535 days compared with historic median of 200 days; 95% confidence interval, 154–260 days). The transferred CD8+ T cells preferentially accumulated in the tumor tissue, remained detectable and functional for more than 200 days, persisted with an effector phenotype, and exhibited evidence of recent in vivo activation and proliferation. The combination of local and systemic immune stimulatory therapies was well tolerated and may be a promising approach to overcome immune evasion in virus-driven cancers. Cancer Immunol Res; 2(1); 27–36. ©2013 AACR.
Data from CD16A Activation of NK Cells Promotes NK Cell Proliferation and Memory-Like Cytotoxicity against Cancer Cells
Authors: Pahl Jens H.W. , Koch Joachim , Götz Jana-Julia , Arnold Annette , Reusch Uwe , Gantke Thorsten , Rajkovic Erich , Treder Martin , Cerwenka Adelheid
Published: March 2024

Abstract
CD16A is a potent cytotoxicity receptor on human natural killer (NK) cells, which can be exploited by therapeutic bispecific antibodies. So far, the effects of CD16A-mediated activation on NK cell effector functions beyond classical antibody-dependent cytotoxicity have remained poorly elucidated. Here, we investigated NK cell responses after exposure to therapeutic antibodies such as the tetravalent bispecific antibody AFM13 (CD30/CD16A), designed for the treatment of Hodgkin lymphoma and other CD30+ lymphomas. Our results reveal that CD16A engagement enhanced subsequent IL2- and IL15-driven NK cell proliferation and expansion. This effect involved the upregulation of CD25 (IL2Rα) and CD132 (γc) on NK cells, resulting in increased sensitivity to low-dose IL2 or to IL15. CD16A engagement initially induced NK cell cytotoxicity. The lower NK cell reactivity observed 1 day after CD16A engagement could be recovered by reculture in IL2 or IL15. After reculture in IL2 or IL15, these CD16A-experienced NK cells exerted more vigorous IFNγ production upon restimulation with tumor cells or cytokines. Importantly, after reculture, CD16A-experienced NK cells also exerted increased cytotoxicity toward different tumor targets, mainly through the activating NK cell receptor NKG2D. Our findings uncover a role for CD16A engagement in priming NK cell responses to restimulation by cytokines and tumor cells, indicative of a memory-like functionality. Our study suggests that combination of AFM13 with IL2 or IL15 may boost NK cell antitumor activity in patients by expanding tumor-reactive NK cells and enhancing NK cell reactivity, even upon repeated tumor encounters. Cancer Immunol Res; 6(5); 517–27. ©2018 AACR.
Data from Rapid Construction of Antitumor T-cell Receptor Vectors from Frozen Tumors for Engineered T-cell Therapy
Authors: Tsuji Takemasa , Yoneda Akira , Matsuzaki Junko , Miliotto Anthony , Ryan Courtney , Koya Richard C. , Odunsi Kunle
Published: March 2024

Abstract
T cells genetically engineered with tumor antigen–specific T-cell receptor (TCR) genes have demonstrated therapeutic potential in patients with solid tumors. In order to achieve broader application, an efficient method to identify TCR genes for an array of tumor antigens and HLA restriction elements is required. Here, we have developed a method to construct a TCR-expression library from specimens, including frozen tumor biopsies, that contain antigen-specific T cells. TCR-expressing cassettes were constructed and cloned in a retroviral plasmid vector within 24 hours by unbiased PCR amplification of TCR α and β chain variable regions assembled with TCR constant regions. The method was validated by constructing TCR-expressing vectors from tumor antigen–specific T-cell clones and functionally assessing TCR gene–transduced T cells. We applied this method to frozen ovarian tumor specimens that were infiltrated by tumor antigen–specific T cells. The tumor-derived TCR libraries were expressed in peripheral T cells from healthy volunteers and screened for tumor antigen–specific TCR pairs with the use of an MHC/peptide tetramer reagent. Tumor antigen–specific TCR-expressing transgenes were recovered from isolated tetramer-positive T cells. Peripheral T cells that were engineered with library-derived TCR gene showed potent therapeutic antitumor effect in a tumor xenograft model. Our method can efficiently and rapidly provide tumor-specific TCR-expressing viral vectors for the manufacture of therapeutic and personalized antitumor T-cell products. Cancer Immunol Res; 6(5); 594–604. ©2018 AACR.
Data from Mapping the MHC Class I–Spliced Immunopeptidome of Cancer Cells
Authors: Liepe Juliane , Sidney John , Lorenz Felix K.M. , Sette Alessandro , Mishto Michele
Published: March 2024

Abstract
Anticancer immunotherapies demand optimal epitope targets, which could include proteasome-generated spliced peptides if tumor cells were to present them. Here, we show that spliced peptides are widely presented by MHC class I molecules of colon and breast carcinoma cell lines. The peptides derive from hot spots within antigens and enlarge the antigen coverage. Spliced peptides also represent a large number of antigens that would otherwise be neglected by patrolling T cells. These antigens tend to be long, hydrophobic, and basic. Thus, spliced peptides can be a key to identifying targets in an enlarged pool of antigens associated with cancer.
Data from Programmed Cell Death Ligand 1 (PD-L1) Signaling Regulates Macrophage Proliferation and Activation
Authors: Hartley Genevieve P. , Chow Lyndah , Ammons Dylan T. , Wheat William H. , Dow Steven W.
Published: March 2024

Abstract
Tumor-associated macrophages (TAMs) express programmed cell death ligand 1 (PD-L1) and contribute to the immune-suppressive tumor microenvironment. Although the role of the PD-L1 and PD-1 interaction to regulate T-cell suppression is established, less is known about PD-L1 signaling in macrophages and how these signals may affect the function of TAMs. We used in vitro and in vivo models to investigate PD-L1 signaling in macrophages and the effects of PD-L1 antibody treatment on TAM responses. Treatment of mouse and human macrophages with PD-L1 antibodies increased spontaneous macrophage proliferation, survival, and activation (costimulatory molecule expression, cytokine production). Similar changes were observed in macrophages incubated with soluble CD80 and soluble PD-1, and in PD-L1−/− macrophages. Macrophage treatment with PD-L1 antibodies upregulated mTOR pathway activity, and RNAseq analysis revealed upregulation of multiple macrophage inflammatory pathways. In vivo, treatment with PD-L1 antibody resulted in increased tumor infiltration with activated macrophages. In tumor-bearing RAG−/− mice, upregulated costimulatory molecule expression by TAMs and reduced tumor growth were observed. Combined PD-1/ PD-L1 antibody treatment of animals with established B16 melanomas cured half of the treated mice, whereas treatment with single antibodies had little therapeutic effect. These findings indicate that PD-L1 delivers a constitutive negative signal to macrophages, resulting in an immune-suppressive cell phenotype. Treatment with PD-L1 antibodies reverses this phenotype and triggers macrophage-mediated antitumor activity, suggesting a distinct effect of PD-L1, but not PD-1, antibody treatment. Cancer Immunol Res; 6(10); 1260–73. ©2018 AACR.
Data from Computational Immune Monitoring Reveals Abnormal Double-Negative T Cells Present across Human Tumor Types
Authors: Greenplate Allison R. , McClanahan Daniel D. , Oberholtzer Brian K. , Doxie Deon B. , Roe Caroline E. , Diggins Kirsten E. , Leelatian Nalin , Rasmussen Megan L. , Kelley Mark C. , Gama Vivian , Siska Peter J. , Rathmell Jeffrey C. , Ferrell P. Brent , Johnson Douglas B. , Irish Jonathan M.
Published: March 2024

Abstract
Advances in single-cell biology have enabled measurements of >40 protein features on millions of immune cells within clinical samples. However, the data analysis steps following cell population identification are susceptible to bias, time-consuming, and challenging to compare across studies. Here, an ensemble of unsupervised tools was developed to evaluate four essential types of immune cell information, incorporate changes over time, and address diverse immune monitoring challenges. The four complementary properties characterized were (i) systemic plasticity, (ii) change in population abundance, (iii) change in signature population features, and (iv) novelty of cellular phenotype. Three systems immune monitoring studies were selected to challenge this ensemble approach. In serial biopsies of melanoma tumors undergoing targeted therapy, the ensemble approach revealed enrichment of double-negative (DN) T cells. Melanoma tumor-resident DN T cells were abnormal and phenotypically distinct from those found in nonmalignant lymphoid tissues, but similar to those found in glioblastoma and renal cell carcinoma. Overall, ensemble systems immune monitoring provided a robust, quantitative view of changes in both the system and cell subsets, allowed for transparent review by human experts, and revealed abnormal immune cells present across multiple human tumor types.
Data from Human GUCY2C-Targeted Chimeric Antigen Receptor (CAR)-Expressing T Cells Eliminate Colorectal Cancer Metastases
Authors: Magee Michael S. , Abraham Tara S. , Baybutt Trevor R. , Flickinger John C. , Ridge Natalie A. , Marszalowicz Glen P. , Prajapati Priyanka , Hersperger Adam R. , Waldman Scott A. , Snook Adam E.
Published: March 2024

Abstract
One major hurdle to the success of adoptive T-cell therapy is the identification of antigens that permit effective targeting of tumors in the absence of toxicities to essential organs. Previous work has demonstrated that T cells engineered to express chimeric antigen receptors (CAR-T cells) targeting the murine homolog of the colorectal cancer antigen GUCY2C treat established colorectal cancer metastases, without toxicity to the normal GUCY2C-expressing intestinal epithelium, reflecting structural compartmentalization of endogenous GUCY2C to apical membranes comprising the intestinal lumen. Here, we examined the utility of a human-specific, GUCY2C-directed single-chain variable fragment as the basis for a CAR construct targeting human GUCY2C-expressing metastases. Human GUCY2C-targeted murine CAR-T cells promoted antigen-dependent T-cell activation quantified by activation marker upregulation, cytokine production, and killing of GUCY2C-expressing, but not GUCY2C-deficient, cancer cells in vitro. GUCY2C CAR-T cells provided long-term protection against lung metastases of murine colorectal cancer cells engineered to express human GUCY2C in a syngeneic mouse model. GUCY2C murine CAR-T cells recognized and killed human colorectal cancer cells endogenously expressing GUCY2C, providing durable survival in a human xenograft model in immunodeficient mice. Thus, we have identified a human GUCY2C-specific CAR-T cell therapy approach that may be developed for the treatment of GUCY2C-expressing metastatic colorectal cancer. Cancer Immunol Res; 6(5); 509–16. ©2018 AACR.
Data from Phase I Trial of Autologous CAR T Cells Targeting NKG2D Ligands in Patients with AML/MDS and Multiple Myeloma
Authors: Baumeister Susanne H. , Murad Joana , Werner Lillian , Daley Heather , Trebeden-Negre Helene , Gicobi Joanina K. , Schmucker Adam , Reder Jake , Sentman Charles L. , Gilham David E. , Lehmann Frédéric F. , Galinsky Ilene , DiPietro Heidi , Cummings Kristen , Munshi Nikhil C. , Stone Richard M. , Neuberg Donna S. , Soiffer Robert , Dranoff Glenn , Ritz Jerome , Nikiforow Sarah
Published: March 2024

Abstract
NKG2D ligands are widely expressed in solid and hematologic malignancies but absent or poorly expressed on healthy tissues. We conducted a phase I dose-escalation study to evaluate the safety and feasibility of a single infusion of NKG2D-chimeric antigen receptor (CAR) T cells, without lymphodepleting conditioning in subjects with acute myeloid leukemia/myelodysplastic syndrome or relapsed/refractory multiple myeloma. Autologous T cells were transfected with a γ-retroviral vector encoding a CAR fusing human NKG2D with the CD3ζ signaling domain. Four dose levels (1 × 106–3 × 107 total viable T cells) were evaluated. Twelve subjects were infused [7 acute myeloid leukemia (AML) and 5 multiple myeloma]. NKG2D-CAR products demonstrated a median 75% vector-driven NKG2D expression on CD3+ T cells. No dose-limiting toxicities, cytokine release syndrome, or CAR T cell–related neurotoxicity was observed. No significant autoimmune reactions were noted, and none of the ≥ grade 3 adverse events were attributable to NKG2D-CAR T cells. At the single injection of low cell doses used in this trial, no objective tumor responses were observed. However, hematologic parameters transiently improved in one subject with AML at the highest dose, and cases of disease stability without further therapy or on subsequent treatments were noted. At 24 hours, the cytokine RANTES increased a median of 1.9-fold among all subjects and 5.8-fold among six AML patients. Consistent with preclinical studies, NKG2D-CAR T cell–expansion and persistence were limited. Manufactured NKG2D-CAR T cells exhibited functional activity against autologous tumor cells in vitro, but modifications to enhance CAR T-cell expansion and target density may be needed to boost clinical activity.
Data from TGFβ Inhibition Prior to Hypofractionated Radiation Enhances Efficacy in Preclinical Models
Authors: Young Kristina H. , Newell Pippa , Cottam Benjamin , Friedman David , Savage Talicia , Baird Jason R. , Akporiaye Emmanuel , Gough Michael J. , Crittenden Marka
Published: March 2024

Abstract
The immune infiltrate in colorectal cancer has been correlated with outcome, such that individuals with higher infiltrations of T cells have increased survival independent of the disease stage. For patients with lower immune infiltrates, overall survival is limited. Because the patients with colorectal cancer studied have received conventional cancer therapies, these data may indicate that the pretreatment tumor environment increases the efficacy of treatments such as chemotherapy, surgery, and radiotherapy. This study was designed to test the hypothesis that an improved immune environment in the tumor at the time of treatment will increase the efficacy of radiotherapy. We demonstrate that inhibition of TGFβ using the orally available small-molecule inhibitor SM16 improved the immune environment of tumors in mice and significantly improved the efficacy of subsequent radiotherapy. This effect was not due to changes in radiosensitivity, epithelial–mesenchymal transition, or changes in vascular function in the tumor; rather, this effect was dependent on adaptive immunity and resulted in long-term protective immunity in cured mice. These data demonstrate that immunotherapy is an option to improve the immune status of patients with poor tumor infiltrates and that pretreatment improves the efficacy of radiotherapy. Cancer Immunol Res; 2(10); 1011–22. ©2014 AACR.
Data from Lysophosphatidic Acid Inhibits CD8 T-cell Activation and Control of Tumor Progression
Authors: Oda Shannon K. , Strauch Pamela , Fujiwara Yuko , Al-Shami Amin , Oravecz Tamas , Tigyi Gabor , Pelanda Roberta , Torres Raul M.
Published: March 2024

Abstract
CD8 T lymphocytes are able to eliminate nascent tumor cells through a process referred to as immunosurveillance. However, multiple inhibitory mechanisms within the tumor microenvironment have been described that impede tumor rejection by CD8 T cells, including increased signaling by inhibitory receptors. Lysophosphatidic acid (LPA) is a bioactive lysophospholipid that has been shown repeatedly to promote diverse cellular processes benefiting tumorigenesis. Accordingly, the exaggerated expression of LPA and LPA receptors is a common feature of diverse tumor cell lineages and can result in elevated systemic LPA levels. LPA is recognized by at least six distinct G protein–coupled receptors, several of which are expressed by T cells, although the precise function of LPA signaling in CD8 T-cell activation and function has not been defined. Here, we show that LPA signaling via the LPA5 receptor expressed by CD8 T cells suppresses antigen receptor signaling, cell activation, and proliferation in vitro and in vivo. Importantly, in a mouse melanoma model tumor-specific CD8 T cells that are LPA5-deficient are able to control tumor growth significantly better than wild-type tumor-specific CD8 T cells. Together, these data suggest that the production of LPA by tumors serves not only in an autocrine manner to promote tumorigenesis, but also as a mechanism to suppress adaptive immunity and highlights a potential novel target for cancer treatment. Cancer Immunol Res; 1(4); 245–55. ©2013 AACR.
Data from Influenza Virus Infection Elicits Protective Antibodies and T Cells Specific for Host Cell Antigens Also Expressed as Tumor-Associated Antigens: A New View of Cancer Immunosurveillance
Authors: Iheagwara Uzoma K. , Beatty Pamela L. , Van Phu T. , Ross Ted M. , Minden Jonathan S. , Finn Olivera J.
Published: March 2024

Abstract
Most tumor-associated antigens (TAA) are self-molecules that are abnormally expressed in cancer cells and become targets of antitumor immune responses. Antibodies and T cells specific for some TAAs have been found in healthy individuals and are associated with lowered lifetime risk for developing cancer. Lower risk for cancer has also been associated with a history of febrile viral diseases. We hypothesized that virus infections could lead to transient expression of abnormal forms of self-molecules, some of which are TAAs; facilitated by the adjuvant effects of infection and inflammation, these molecules could elicit specific antibodies, T cells, and lasting immune memory simultaneously with immunity against viral antigens. Such infection-induced immune memory for TAA would be expected to provide life-long immune surveillance of cancer. Using influenza virus infection in mice as a model system, we tested this hypothesis and demonstrated that influenza-experienced mice control 3LL mouse lung tumor challenge better than infection-naive control mice. Using 2D-difference gel electrophoresis and mass spectrometry, we identified numerous molecules, some of which are known TAAs, on the 3LL tumor cells recognized by antibodies elicited by two successive influenza infections. We studied in detail immune responses against glyceraldehyde-3-phosphate dehydrogenase (GAPDH), histone H4, HSP90, malate dehydrogenase 2, and annexin A2, all of which were overexpressed in influenza-infected lungs and in tumor cells. Finally, we show that immune responses generated through vaccination against peptides derived from these antigens correlated with improved tumor control. Cancer Immunol Res; 2(3); 263–73. ©2013 AACR.
Data from Plasmacytoid Dendritic Cells Support Melanoma Progression by Promoting Th2 and Regulatory Immunity through OX40L and ICOSL
Authors: Aspord Caroline , Leccia Marie-Therese , Charles Julie , Plumas Joel
Published: March 2024

Abstract
Even though melanoma is considered to be one of the most immunogenic solid tumors, handling its development remains a challenge. The basis for such escape from antitumor immune control has not yet been documented. Plasmacytoid dendritic cells (pDC) are emerging as crucial but still enigmatic cells in cancer. In melanoma, the function of tumor-infiltrating pDCs remains poorly explored. We investigated the pathophysiologic role of pDCs in melanoma, both ex vivo from a large cohort of melanoma patients and in vivo in melanoma-bearing humanized mice. pDCs were found in high proportions in cutaneous melanoma and tumor-draining lymph nodes, yet associated with poor clinical outcome. We showed that pDCs migrating to the tumor microenvironment displayed particular features, subsequently promoting proinflammatory Th2 and regulatory immune profiles through OX40L and ICOSL expression. Elevated frequencies of interleukin (IL)-5-, IL-13- and IL-10–producing T cells in patients with melanoma correlated with high proportions of OX40L- and ICOSL-expressing pDCs. Strikingly TARC/CCL17, MDC/CCL22, and MMP-2 found in the melanoma microenvironment were associated with pDC accumulation, OX40L and ICOSL modulation, and/or early relapse. Thus, melanoma actively exploits pDC plasticity to promote its progression. By identifying novel insights into the mechanism of hijacking of immunity by melanoma, our study exposes potential for new therapeutic opportunities. Cancer Immunol Res; 1(6); 402–15. ©2013 AACR.