bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Anti–PD-L1 Treatment Results in Functional Remodeling of the Macrophage Compartment
Authors: Xiong Huizhong , Mittman Stephanie , Rodriguez Ryan , Moskalenko Marina , Pacheco-Sanchez Patricia , Yang Yagai , Nickles Dorothee , Cubas Rafael
Published: March 2023

Abstract
Checkpoint inhibitors like anti-PD1/PD-L1 have demonstrated significant therapeutic efficacy in a subset of patients partly through reinvigoration of CD8 T cells. However, their impact on myeloid cells remains largely unknown. Here, we report that anti–PD-L1 treatment favorably impacts the phenotype and function of tumor macrophages by polarizing the macrophage compartment toward a more proinflammatory phenotype. This phenotype was characterized by a decrease in Arginase-I (ARG1) expression and an increase in iNOS, MHCII, and CD40 expression. Whole-transcriptome profiling further confirmed extensive polarization of both tumor monocytes and macrophages from a suppressive to a proinflammatory, immunostimulatory phenotype. This polarization was driven mainly through IFNγ and was associated with enhanced T-cell activity. Transfer of monocytes into anti–PD-L1–treated tumor-bearing mice led to macrophage differentiation into a more proinflammatory phenotype, with an increase in CD8 T cells expressing granzyme-B and an increase in the CD8/Treg ratio compared with control-treated mice. Although in responsive tumor models, anti–PD-L1 treatment remodeled the macrophage compartment with beneficial effects on T cells, both macrophage reprogramming and depletion were needed to maximize anti–PD-L1 responses in a tumor immune contexture with high macrophage burden. Our results demonstrate that anti–PD-L1 treatment can favorably remodel the macrophage compartment in responsive tumor models toward a more proinflammatory phenotype, mainly through increased IFNγ levels. They also suggest that directly targeting these cells with reprogramming and depleting agents may further augment the breadth and depth of response to anti–PD-L1 treatment in less responsive or more macrophage-dense tumor microenvironments.
Significance:
This work demonstrates that increased IFNγ signaling following anti–PD-L1 treatment can remodel the macrophage compartment to enhance T-cell responses.
Data from Radiosensitivity Is an Acquired Vulnerability of PARPi-Resistant BRCA1-Deficient Tumors
Authors: Barazas Marco , Gasparini Alessia , Huang Yike , Küçükosmanoğlu Asli , Annunziato Stefano , Bouwman Peter , Sol Wendy , Kersbergen Ariena , Proost Natalie , de Korte-Grimmerink Renske , van de Ven Marieke , Jonkers Jos , Borst Gerben R. , Rottenberg Sven
Published: March 2023

Abstract
The defect in homologous recombination (HR) found in BRCA1-associated cancers can be therapeutically exploited by treatment with DNA-damaging agents and PARP inhibitors. We and others previously reported that BRCA1-deficient tumors are initially hypersensitive to the inhibition of topoisomerase I/II and PARP, but acquire drug resistance through restoration of HR activity by the loss of end-resection antagonists of the 53BP1/RIF1/REV7/Shieldin/CST pathway. Here, we identify radiotherapy as an acquired vulnerability of 53BP1;BRCA1-deficient cells in vitro and in vivo. In contrast to the radioresistance caused by HR restoration through BRCA1 reconstitution, HR restoration by 53BP1 pathway inactivation further increases radiosensitivity. This highlights the relevance of this pathway for the repair of radiotherapy-induced damage. Moreover, our data show that BRCA1-mutated tumors that acquire drug resistance due to BRCA1-independent HR restoration can be targeted by radiotherapy.
Significance:
These findings uncover radiosensitivity as a novel, therapeutically viable vulnerability of BRCA1-deficient mouse mammary cells that have acquired drug resistance due to the loss of the 53BP1 pathway.
Data from RNF168-Mediated Ubiquitin Signaling Inhibits the Viability of BRCA1-Null Cancers
Authors: Krais John J. , Wang Yifan , Bernhardy Andrea J. , Clausen Emma , Miller Jessica A. , Cai Kathy Q. , Scott Clare L. , Johnson Neil
Published: March 2023

Abstract
BRCA1 gene mutations impair homologous recombination (HR) DNA repair, resulting in cellular senescence and embryonic lethality in mice. Therefore, BRCA1-deficient cancers require adaptations that prevent excessive genomic alterations from triggering cell death. RNF168-mediated ubiquitination of γH2AX at K13/15 (ub-H2AX) serves as a recruitment module for the localization of 53BP1 to DNA break sites. Here, we found multiple BRCA1-mutant cancer cell lines and primary tumors with low levels of RNF168 protein expression. Overexpression of ectopic RNF168 or a ub-H2AX fusion protein induced cell death and delayed BRCA1-mutant tumor formation. Cell death resulted from the recruitment of 53BP1 to DNA break sites and inhibition of DNA end resection. Strikingly, reintroduction of BRCA1 or 53BP1 depletion restored HR and rescued the ability of cells to maintain RNF168 and ub-H2AX overexpression. Thus, downregulation of RNF168 protein expression is a mechanism for providing BRCA1-null cancer cell lines with a residual level of HR that is essential for viability. Overall, our work identifies loss of RNF168 ubiquitin signaling as a proteomic alteration that supports BRCA1-mutant carcinogenesis. We propose that restoring RNF168-ub-H2AX signaling, potentially through inhibition of deubiquitinases, could represent a new therapeutic approach.
Significance:
This study explores the concept that homologous recombination DNA repair is not an all-or-nothing concept, but a spectrum, and that where a tumor stands on this spectrum may have therapeutic relevance.

See related commentary by Wang and Wulf, p. 2720
Data from ZNF143-Mediated H3K9 Trimethylation Upregulates CDC6 by Activating MDIG in Hepatocellular Carcinoma
Authors: Zhang Lili , Huo Qi , Ge Chao , Zhao Fangyu , Zhou Qingqing , Chen Xiaoxia , Tian Hua , Chen Taoyang , Xie Haiyang , Cui Ying , Yao Ming , Li Hong , Li Jinjun
Published: March 2023

Abstract
Zinc finger protein 143 (ZNF143) belongs to the zinc finger protein family and possesses transcription factor activity by binding sequence-specific DNA. The exact biological role of ZNF143 in hepatocellular carcinoma (HCC) has not been investigated. Here we report that ZNF143 is overexpressed in HCC tissues and its overexpression correlates with poor prognosis. Gain- and loss-of-function experiments showed that ZNF143 promoted HCC cell proliferation, colony formation, and tumor growth in vitro and in vivo. ZNF143 accelerated HCC cell-cycle progression by activating cell division cycle 6 (CDC6). Mechanistically, ZNF143 promoted expression of CDC6 by directly activating transcription of histone demethylase mineral dust–induced gene (MDIG), which in turn reduced H3K9me3 enrichment in the CDC6 promoter region. Consistently, ZNF143 expression correlated significantly with MDIG and CDC6 expression in HCC. Collectively, we propose a model for a ZNF143–MDIG–CDC6 oncoprotein axis that provides novel insight into ZNF143, which may serve as a therapeutic target in HCC.
Significance:
These findings describe the mechanism by which ZNF143 promotes HCC proliferation and provide important clues for exploring new targets and strategies for clinical treatment of human liver cancer.
Data from Pomalidomide Alters Pancreatic Macrophage Populations to Generate an Immune-Responsive Environment at Precancerous and Cancerous Lesions
Authors: Bastea Ligia I. , Liou Geou-Yarh , Pandey Veethika , Fleming Alicia K. , von Roemeling Christina A. , Doeppler Heike , Li Zhimin , Qiu Yushi , Edenfield Brandy , Copland John A. , Tun Han W. , Storz Peter
Published: March 2023

Abstract
During development of pancreatic cancer, alternatively activated macrophages contribute to fibrogenesis, pancreatic intraepithelial neoplasia (PanIN) lesion growth, and generation of an immunosuppressive environment. Here, we show that the immunomodulatory agent pomalidomide depletes pancreatic lesion areas of alternatively activated macrophage populations. Pomalidomide treatment resulted in downregulation of interferon regulatory factor 4, a transcription factor for M2 macrophage polarization. Pomalidomide-induced absence of alternatively activated macrophages led to a decrease in fibrosis at PanIN lesions and in syngeneic tumors; this was due to generation of an inflammatory, immune-responsive environment with increased expression of IL1α and presence of activated (IFNγ-positive) CD4+ and CD8+ T-cell populations. Our results indicate that pomalidomide could be used to decrease fibrogenesis in pancreatic cancer and may be ideal as a combination treatment with chemotherapeutic drugs or other immunotherapies.
Significance:
These findings reveal new insights into how macrophage populations within the pancreatic cancer microenvironment can be modulated, providing the means to turn the microenvironment from immunosuppressive to immune-responsive.
Data from Myc and Loss of p53 Cooperate to Drive Formation of Choroid Plexus Carcinoma
Authors: Wang Jun , Merino Diana M. , Light Nicholas , Murphy Brian L. , Wang Yong-Dong , Guo Xiaohui , Hodges Andrew P. , Chau Lianne Q. , Liu Kun-Wei , Dhall Girish , Asgharzadeh Shahab , Kiehna Erin N. , Shirey Ryan J. , Janda Kim D. , Taylor Michael D. , Malkin David , Ellison David W. , VandenBerg Scott R. , Eberhart Charles G. , Sears Rosalie C. , Roussel Martine F. , Gilbertson Richard J. , Wechsler-Reya Robert J.
Published: March 2023

Abstract
Choroid plexus carcinoma (CPC) is a rare brain tumor that occurs most commonly in very young children and has a dismal prognosis despite intensive therapy. Improved outcomes for patients with CPC depend on a deeper understanding of the mechanisms underlying the disease. Here we developed transgenic models of CPCs by activating the Myc oncogene and deleting the Trp53 tumor suppressor gene in murine neural stem cells or progenitors. Murine CPC resembled their human counterparts at a histologic level, and like the hypodiploid subset of human CPC, exhibited multiple whole-chromosome losses, particularly of chromosomes 8, 12, and 19. Analysis of murine and human CPC gene expression profiles and copy number changes revealed altered expression of genes involved in cell cycle, DNA damage response, and cilium function. High-throughput drug screening identified small molecule inhibitors that decreased the viability of CPC. These models will be valuable tools for understanding the biology of choroid plexus tumors and for testing novel approaches to therapy.
Significance:
This study describes new mouse models of choroid plexus carcinoma and uses them to investigate the biology and therapeutic responsiveness of this highly malignant pediatric brain tumor.
Data from H2A Monoubiquitination Links Glucose Availability to Epigenetic Regulation of the Endoplasmic Reticulum Stress Response and Cancer Cell Death
Authors: Zhang Yilei , Shi Jiejun , Liu Xiaoguang , Xiao Zhenna , Lei Guang , Lee Hyemin , Koppula Pranavi , Cheng Weijie , Mao Chao , Zhuang Li , Ma Li , Li Wei , Gan Boyi
Published: March 2023

Abstract
Epigenetic regulation of gene transcription has been shown to coordinate with nutrient availability, yet the mechanisms underlying this coordination remain incompletely understood. Here, we show that glucose starvation suppresses histone 2A K119 monoubiquitination (H2Aub), a histone modification that correlates with gene repression. Glucose starvation suppressed H2Aub levels independently of energy stress–mediated AMP-activated protein kinase activation and possibly through NADPH depletion and subsequent inhibition of BMI1, an integral component of polycomb-repressive complex 1 (PRC1) that catalyzes H2Aub on chromatin. Integrated transcriptomic and epigenomic analyses linked glucose starvation–mediated H2Aub repression to the activation of genes involved in the endoplasmic reticulum (ER) stress response. We further showed that this epigenetic mechanism has a role in glucose starvation–induced cell death and that pharmacologic inhibition of glucose transporter 1 and PRC1 synergistically promoted ER stress and suppressed tumor growth in vivo. Together, these results reveal a hitherto unrecognized epigenetic mechanism coupling glucose availability to the ER stress response.
Significance:
These findings link glucose deprivation and H2A ubiquitination to regulation of the ER stress response in tumor growth and demonstrate pharmacologic susceptibility to inhibition of polycomb and glucose transporters.
Data from Platelet-Specific PDGFB Ablation Impairs Tumor Vessel Integrity and Promotes Metastasis
Authors: Zhang Yanyu , Cedervall Jessica , Hamidi Anahita , Herre Melanie , Viitaniemi Kati , D'Amico Gabriela , Miao Zuoxiu , Unnithan Ragaseema Valsala Madhavan , Vaccaro Alessandra , van Hooren Luuk , Georganaki Maria , Thulin Åsa , Qiao Qi , Andrae Johanna , Siegbahn Agneta , Heldin Carl-Henrik , Alitalo Kari , Betsholtz Christer , Dimberg Anna , Olsson Anna-Karin
Published: March 2023

Abstract
Platelet-derived growth factor B (PDGFB) plays a crucial role in recruitment of PDGF receptor β–positive pericytes to blood vessels. The endothelium is an essential source of PDGFB in this process. Platelets constitute a major reservoir of PDGFB and are continuously activated in the tumor microenvironment, exposing tumors to the plethora of growth factors contained in platelet granules. Here, we show that tumor vascular function, as well as pericyte coverage is significantly impaired in mice with conditional knockout of PDGFB in platelets. A lack of PDGFB in platelets led to enhanced hypoxia and epithelial-to-mesenchymal transition in the primary tumors, elevated levels of circulating tumor cells, and increased spontaneous metastasis to the liver or lungs in two mouse models. These findings establish a previously unknown role for platelet-derived PDGFB, whereby it promotes and maintains vascular integrity in the tumor microenvironment by contributing to the recruitment of pericytes.
Significance:
Conditional knockout of PDGFB in platelets demonstrates its previously unknown role in the maintenance of tumor vascular integrity and host protection against metastasis.
Data from Host Wnt5a Potentiates Microenvironmental Regulation of Ovarian Cancer Metastasis
Authors: Asem Marwa , Young Allison M. , Oyama Carlysa , Claure De La Zerda Alejandro , Liu Yueying , Yang Jing , Hilliard Tyvette S. , Johnson Jeffery , Harper Elizabeth I. , Guldner Ian , Zhang Siyuan , Page-Mayberry Toni , Kaliney William J. , Stack M. Sharon
Published: March 2023

Abstract
The noncanonical Wnt ligand Wnt5a is found in high concentrations in ascites of women with ovarian cancer. In this study, we elucidated the role of Wnt5a in ovarian cancer metastasis. Wnt5a promoted ovarian tumor cell adhesion to peritoneal mesothelial cells as well as migration and invasion, leading to colonization of peritoneal explants. Host components of the ovarian tumor microenvironment, notably peritoneal mesothelial cells and visceral adipose, secreted Wnt5a. Conditional knockout of host WNT5A significantly reduced peritoneal metastatic tumor burden. Tumors formed in WNT5A knockout mice had elevated cytotoxic T cells, increased M1 macrophages, and decreased M2 macrophages, indicating that host Wnt5a promotes an immunosuppressive microenvironment. The Src family kinase Fgr was identified as a downstream effector of Wnt5a. These results highlight a previously unreported role for host-expressed Wnt5a in ovarian cancer metastasis and suggest Fgr as a novel target for inhibition of ovarian cancer metastatic progression.

Significance: This study establishes host-derived Wnt5a, expressed by peritoneal mesothelial cells and adipocytes, as a primary regulator of ovarian cancer intraperitoneal metastatic dissemination and identifies Fgr kinase as novel target for inhibition of metastasis.
Data from A miRNA Expression Signature in Breast Tumor Tissue Is Associated with Risk of Distant Metastasis
Authors: Rohan Thomas E. , Wang Tao , Weinmann Sheila , Wang Yihong , Lin Juan , Ginsberg Mindy , Loudig Olivier
Published: March 2023

Abstract
Dysregulation of miRNA expression may influence breast cancer progression, and experimental evidence suggests that miRNA silencing might suppress breast cancer metastasis. However, the relationship between miRNA and metastasis must be confirmed before this approach can be applied in the clinic. To this end, we conducted a two-stage study in a cohort of 3,760 patients with breast cancer to first identify and then validate the association between miRNA expression and risk of distant metastasis. The first stage (discovery) entailed miRNA sequencing of 126 case–control pairs; qPCR was used to validate the findings in a separate set of 80 case–control pairs. The 13 miRNAs most differentially expressed between cases and controls were combined into an miRNA score that was significantly associated with risk of distant metastasis in a logistic regression model that also included clinical variables (tumor size and number of positive lymph nodes) (ORper unit increase in score = 1.30; 95% confidence interval, 1.03–1.66). The results of this study suggest that in women with invasive breast cancer, a miRNA score that incorporates both clinical variables and miRNA expression levels in breast tumor tissue is moderately predictive of risk of subsequent distant metastasis.
Significance:
A novel predictive scoring system for patients with breast cancer includes clinical variables and the expression levels of 13 miRNAs and may help to identify those at increased risk of distant metastasis.
Data from Loss of a Negative Feedback Loop between IRF8 and AR Promotes Prostate Cancer Growth and Enzalutamide Resistance
Authors: Wu Hongxi , You Linjun , Li Yan , Zhao Zhili , Shi Guangjiang , Chen Zhen , Wang Zhuo , Li Xianjing , Du Shijia , Ye Wanli , Gao Xiaofang , Duan Jingjing , Cheng Yan , Tao Weiyan , Bian Jinsong , Zhou Jin-Rong , Zhu Qingyi , Yang Yong
Published: March 2023

Abstract
In incurable castration-resistant prostate cancer (CRPC), resistance to the novel androgen receptor (AR) antagonist enzalutamide is driven mainly by AR overexpression. Here we report that the expression of interferon regulatory factor 8 (IRF8) is increased in primary prostate cancer but decreased in CRPC compared with normal prostate tissue. Decreased expression of IRF8 positively associated with CRPC progression and enzalutamide resistance. IRF8 interacted with AR and promoted its degradation via activation of the ubiquitin/proteasome systems. Epigenetic knockdown of IRF8 promoted AR-mediated prostate cancer progression and enzalutamide resistance in vitro and in vivo. Furthermore, IFNα increased expression of IRF8 and improved the efficacy of enzalutamide in CRPC by targeting the IRF8–AR axis. We also provide preliminary evidence for the efficacy of IFNα with hormonotherapy in a clinical study. Collectively, this study identifies IRF8 both as a tumor suppressor in prostate cancer pathogenesis and a potential alternative therapeutic option to overcome enzalutamide resistance.
Significance:
These findings identify IRF8-mediated AR degradation as a mechanism of resistance to AR-targeted therapy, highlighting the therapeutic potential of IFNα in targeting IRF8–AR axis in CRPC.
Data from Cancer Cell–Derived Matrisome Proteins Promote Metastasis in Pancreatic Ductal Adenocarcinoma
Authors: Tian Chenxi , Öhlund Daniel , Rickelt Steffen , Lidström Tommy , Huang Ying , Hao Liangliang , Zhao Renee T. , Franklin Oskar , Bhatia Sangeeta N. , Tuveson David A. , Hynes Richard O.
Published: March 2023

Abstract
The prognosis for pancreatic ductal adenocarcinoma (PDAC) remains poor despite decades of effort. The abundant extracellular matrix (ECM) in PDAC comprises a major fraction of the tumor mass and plays various roles in promoting resistance to therapies. However, nonselective depletion of ECM has led to poor patient outcomes. Consistent with that observation, we previously showed that individual matrisome proteins derived from stromal cells correlate with either long or short patient survival. In marked contrast, those derived from cancer cells correlate strongly with poor survival. Here, we studied three cancer cell–derived matrisome proteins that are significantly overrepresented during PDAC progression, AGRN (agrin), SERPINB5 (serine protease inhibitor B5), and CSTB (cystatin B). Using both overexpression and knockdown experiments, we demonstrate that all three are promoters of PDAC metastasis. Furthermore, these proteins operate at different metastatic steps. AGRN promoted epithelial-to-mesenchymal transition in primary tumors, whereas SERPINB5 and CSTB enhanced late steps in the metastatic cascade by elevating invadopodia formation and in vivo extravasation. All three genes were associated with a poor prognosis in human patients and high levels of SERPINB5, secreted by cancer cells and deposited in the ECM, correlated with poor patient prognosis. This study provides strong evidence that cancer cell–derived matrisome proteins can be causal in promoting tumorigenesis and metastasis and lead to poor patient survival. Therefore, compared with the bulk matrix, mostly made by stromal cells, precise interventions targeting cancer cell–derived matrisome proteins, such as AGRN, SERPINB5, and CSTB, may represent preferred potential therapeutic targets.
Significance:
This study provides insights into the biological roles of cancer cell–derived matrisome proteins in PDAC and supports the notion that these proteins are protumorigenic and better therapeutic targets.
Data from Senescent Stromal Cells Promote Cancer Resistance through SIRT1 Loss-Potentiated Overproduction of Small Extracellular Vesicles
Authors: Han Liu , Long Qilai , Li Shenjun , Xu Qixia , Zhang Boyi , Dou Xuefeng , Qian Min , Jiramongkol Yannasittha , Guo Jianming , Cao Liu , Chin Y. Eugene , Lam Eric W.-F. , Jiang Jing , Sun Yu
Published: March 2023

Abstract
Cellular senescence is a potent tumor-suppressive program that prevents neoplastic events. Paradoxically, senescent cells develop an inflammatory secretome, termed the senescence-associated secretory phenotype, which is implicated in age-related pathologies including cancer. Here, we report that senescent cells actively synthesize and release small extracellular vesicles (sEV) with a distinctive size distribution. Mechanistically, SIRT1 loss supported accelerated sEV production despite enhanced proteome-wide ubiquitination, a process correlated with ATP6V1A downregulation and defective lysosomal acidification. Once released, senescent stromal sEVs significantly altered the expression profile of recipient cancer cells and enhanced their aggressiveness, specifically drug resistance mediated by expression of ATP-binding cassette subfamily B member 4 (ABCB4). Targeting SIRT1 with agonist SRT2104 prevented development of cancer resistance by restraining sEV production by senescent stromal cells. In clinical oncology, sEVs in peripheral blood of posttreatment cancer patients were readily detectable by routine biotechniques, presenting an exploitable biomarker to monitor therapeutic efficacy and predict long-term outcome. Together, this study identifies a distinct mechanism supporting pathologic activities of senescent cells and provides a potent avenue to circumvent advanced human malignancies by cotargeting cancer cells and their surrounding microenvironment, which contributes to drug resistance via secretion of sEVs from senescent stromal cells.
Significance:
Senescent stromal cells produce a large number of sEVs to promote cancer resistance in therapeutic settings, a process driven by SIRT1 decline in stromal cells and ABCB4 augmentation in cancer cells.

See related commentary by Wiley, p. 3193
Data from Autofluorescence Imaging of 3D Tumor–Macrophage Microscale Cultures Resolves Spatial and Temporal Dynamics of Macrophage Metabolism
Authors: Heaster Tiffany M. , Humayun Mouhita , Yu Jiaquan , Beebe David J. , Skala Melissa C.
Published: March 2023

Abstract
Macrophages within the tumor microenvironment (TME) exhibit a spectrum of protumor and antitumor functions, yet it is unclear how the TME regulates this macrophage heterogeneity. Standard methods to measure macrophage heterogeneity require destructive processing, limiting spatiotemporal studies of function within the live, intact 3D TME. Here, we demonstrate two-photon autofluorescence imaging of NAD(P)H and FAD to nondestructively resolve spatiotemporal metabolic heterogeneity of individual macrophages within 3D microscale TME models. Fluorescence lifetimes and intensities of NAD(P)H and FAD were acquired at 24, 48, and 72 hours poststimulation for mouse macrophages (RAW264.7) stimulated with IFNγ or IL4 plus IL13 in 2D culture, confirming that autofluorescence measurements capture known metabolic phenotypes. To quantify metabolic dynamics of macrophages within the TME, mouse macrophages or human monocytes (RAW264.7 or THP-1) were cultured alone or with breast cancer cells (mouse polyoma-middle T virus or primary human IDC) in 3D microfluidic platforms. Human monocytes and mouse macrophages in tumor cocultures exhibited significantly different FAD mean lifetimes and greater migration than monocultures at 24, 48, and 72 hours postseeding. In cocultures with primary human cancer cells, actively migrating monocyte-derived macrophages had greater redox ratios [NAD(P)H/FAD intensity] compared with passively migrating monocytes at 24 and 48 hours postseeding, reflecting metabolic heterogeneity in this subpopulation of monocytes. Genetic analyses further confirmed this metabolic heterogeneity. These results establish label-free autofluorescence imaging to quantify dynamic metabolism, polarization, and migration of macrophages at single-cell resolution within 3D microscale models. This combined culture and imaging system provides unique insights into spatiotemporal tumor–immune cross-talk within the 3D TME.
Significance:
Label-free metabolic imaging and microscale culture technologies enable monitoring of single-cell macrophage metabolism, migration, and function in the 3D tumor microenvironment.
Data from FGFR1 Is Critical for RBL2 Loss–Driven Tumor Development and Requires PLCG1 Activation for Continued Growth of Small Cell Lung Cancer
Authors: Kim Kee-Beom , Kim Youngchul , Rivard Christopher J. , Kim Dong-Wook , Park Kwon-Sik
Published: March 2023

Abstract
Small cell lung cancer (SCLC) remains a recalcitrant disease where limited therapeutic options have not improved overall survival, and approved targeted therapies are lacking. Amplification of the tyrosine kinase receptor FGFR1 (fibroblast growth factor receptor 1) is one of the few actionable alterations found in the SCLC genome. However, efforts to develop targeted therapies for FGFR1-amplified SCLC are hindered by critical gaps in knowledge around the molecular origins and mediators of FGFR1-driven signaling as well as the physiologic impact of targeting FGFR1. Here we show that increased FGFR1 promotes tumorigenic progression in precancerous neuroendocrine cells and is required for SCLC development in vivo. Notably, Fgfr1 knockout suppressed tumor development in a mouse model lacking the retinoblastoma-like protein 2 (Rbl2) tumor suppressor gene but did not affect a model with wild-type Rbl2. In support of a functional interaction between these two genes, loss of RBL2 induced FGFR1 expression and restoration of RBL2 repressed it, suggesting a novel role for RBL2 as a regulator of FGFR1 in SCLC. Additionally, FGFR1 activated phospholipase C gamma 1 (PLCG1), whereas chemical inhibition of PLCG1 suppressed SCLC growth, implicating PLCG1 as an effector of FGFR1 signaling in SCLC. Collectively, this study uncovers mechanisms underlying FGFR1-driven SCLC that involve RBL2 upstream and PLCG1 downstream, thus providing potential biomarkers for anti-FGFR1 therapy.
Significance:
This study identifies RBL2 and PLCG1 as critical components of amplified FGFR1 signaling in SCLC, thus representing potential targets for biomarker analysis and therapeutic development in this disease.
Data from The Functional Landscape of Patient-Derived RNF43 Mutations Predicts Sensitivity to Wnt Inhibition
Authors: Yu Jia , Yusoff Permeen A. Mohamed , Woutersen Daniëlle T.J. , Goh Pamela , Harmston Nathan , Smits Ron , Epstein David M. , Virshup David M. , Madan Babita
Published: March 2023

Abstract
A subset of Wnt-addicted cancers are sensitive to targeted therapies that block Wnt secretion or receptor engagement. RNF43 loss-of-function (LOF) mutations that increase cell surface Wnt receptor abundance cause sensitivity to Wnt inhibitors. However, it is not clear which of the clinically identified RNF43 mutations affect its function in vivo. We assayed 119 missense and 45 truncating RNF43 mutations found in human cancers using a combination of cell-based reporter assays, genome editing, flow cytometry, and immunofluorescence microscopy. Five common germline variants of RNF43 exhibited wild-type activity. Cancer-associated missense mutations in the RING ubiquitin ligase domain and a subset of mutations in the extracellular domain hyperactivate Wnt/β-catenin signaling through formation of inactive dimers with endogenous RNF43 or ZNRF3. RNF43 C-terminal truncation mutants, including the common G659fs mutant are LOF specifically when endogenous mutations are examined, unlike their behavior in transient transfection assays. Patient-derived xenografts and cell lines with C-terminal truncations showed increased cell surface Frizzled and Wnt/β-catenin signaling and were responsive to porcupine (PORCN) inhibition in vivo, providing clear evidence of RNF43 impairment. Our study provides potential guidelines for patient assignment, as virtually all RNF43 nonsense and frameshift mutations, including those in the C-terminal domain and a large number of patient-associated missense mutations in the RING domain and N-terminal region compromise its activity, and therefore predict response to upstream Wnt inhibitors in cancers without microsatellite instability. This study expands the landscape of actionable RNF43 mutations, extending the benefit of these therapies to additional patients.
Significance:
Systematic examination of patient-derived RNF43 mutations identifies rules to guide patient selection, including that truncation or point mutations in well-defined functional domains sensitize cancers to PORCN inhibitors.
Data from Oncogenic BRAFV600E Governs Regulatory T-cell Recruitment during Melanoma Tumorigenesis
Authors: Shabaneh Tamer B. , Molodtsov Aleksey K. , Steinberg Shannon M. , Zhang Peisheng , Torres Gretel M. , Mohamed Gadisti A. , Boni Andrea , Curiel Tyler J. , Angeles Christina V. , Turk Mary Jo
Published: March 2023

Abstract
Regulatory T cells (Treg) are critical mediators of immunosuppression in established tumors, although little is known about their role in restraining immunosurveillance during tumorigenesis. Here, we employ an inducible autochthonous model of melanoma to investigate the earliest Treg and CD8 effector T-cell responses during oncogene-driven tumorigenesis. Induction of oncogenic BRAFV600E and loss of Pten in melanocytes led to localized accumulation of FoxP3+ Tregs, but not CD8 T cells, within 1 week of detectable increases in melanocyte differentiation antigen expression. Melanoma tumorigenesis elicited early expansion of shared tumor/self-antigen–specific, thymically derived Tregs in draining lymph nodes, and induced their subsequent recruitment to sites of tumorigenesis in the skin. Lymph node egress of tumor-activated Tregs was required for their C-C chemokine receptor 4 (Ccr4)–dependent homing to nascent tumor sites. Notably, BRAFV600E signaling controlled expression of Ccr4-cognate chemokines and governed recruitment of Tregs to tumor-induced skin sites. BRAFV600E expression alone in melanocytes resulted in nevus formation and associated Treg recruitment, indicating that BRAFV600E signaling is sufficient to recruit Tregs. Treg depletion liberated immunosurveillance, evidenced by CD8 T-cell responses against the tumor/self-antigen gp100, which was concurrent with the formation of microscopic neoplasia. These studies establish a novel role for BRAFV600E as a tumor cell–intrinsic mediator of immune evasion and underscore the critical early role of Treg-mediated suppression during autochthonous tumorigenesis.

Significance: This work provides new insights into the mechanisms by which oncogenic pathways impact immune regulation in the nascent tumor microenvironment. Cancer Res; 78(17); 5038–49. ©2018 AACR.
Data from Functional Precision Medicine Identifies New Therapeutic Candidates for Medulloblastoma
Authors: Rusert Jessica M. , Juarez Edwin F. , Brabetz Sebastian , Jensen James , Garancher Alexandra , Chau Lianne Q. , Tacheva-Grigorova Silvia K. , Wahab Sameerah , Udaka Yoko T. , Finlay Darren , Seker-Cin Huriye , Reardon Brendan , Gröbner Susanne , Serrano Jonathan , Ecker Jonas , Qi Lin , Kogiso Mari , Du Yuchen , Baxter Patricia A. , Henderson Jacob J. , Berens Michael E. , Vuori Kristiina , Milde Till , Cho Yoon-Jae , Li Xiao-Nan , Olson James M. , Reyes Iris , Snuderl Matija , Wong Terence C. , Dimmock David P. , Nahas Shareef A. , Malicki Denise , Crawford John R. , Levy Michael L. , Van Allen Eliezer M. , Pfister Stefan M. , Tamayo Pablo , Kool Marcel , Mesirov Jill P. , Wechsler-Reya Robert J.
Published: March 2023

Abstract
Medulloblastoma is among the most common malignant brain tumors in children. Recent studies have identified at least four subgroups of the disease that differ in terms of molecular characteristics and patient outcomes. Despite this heterogeneity, most patients with medulloblastoma receive similar therapies, including surgery, radiation, and intensive chemotherapy. Although these treatments prolong survival, many patients still die from the disease and survivors suffer severe long-term side effects from therapy. We hypothesize that each patient with medulloblastoma is sensitive to different therapies and that tailoring therapy based on the molecular and cellular characteristics of patients' tumors will improve outcomes. To test this, we assembled a panel of orthotopic patient-derived xenografts (PDX) and subjected them to DNA sequencing, gene expression profiling, and high-throughput drug screening. Analysis of DNA sequencing revealed that most medulloblastomas do not have actionable mutations that point to effective therapies. In contrast, gene expression and drug response data provided valuable information about potential therapies for every tumor. For example, drug screening demonstrated that actinomycin D, which is used for treatment of sarcoma but rarely for medulloblastoma, was active against PDXs representing Group 3 medulloblastoma, the most aggressive form of the disease. Functional analysis of tumor cells was successfully used in a clinical setting to identify more treatment options than sequencing alone. These studies suggest that it should be possible to move away from a one-size-fits-all approach and begin to treat each patient with therapies that are effective against their specific tumor.
Significance:
These findings show that high-throughput drug screening identifies therapies for medulloblastoma that cannot be predicted by genomic or transcriptomic analysis.
Data from The Synthetic Small Molecule FL3 Combats Intestinal Tumorigenesis via Axin1-Mediated Inhibition of Wnt/β-Catenin Signaling
Authors: Jackson Dakota N. , Alula Kibrom M. , Delgado-Deida Yaritza , Tabti Redouane , Turner Kevin , Wang Xuan , Venuprasad K. , Souza Rhonda F. , Désaubry Laurent , Theiss Arianne L.
Published: March 2023

Abstract
Colorectal cancer exhibits aberrant activation of Wnt/β-catenin signaling. Many inhibitors of the Wnt/β-catenin pathway have been tested for Wnt-dependent cancers including colorectal cancer, but are unsuccessful due to severe adverse reactions. FL3 is a synthetic derivative of natural products called flavaglines, which exhibit anti-inflammatory and cytoprotective properties in intestinal epithelial cells, but has not been previously tested in cell or preclinical models of intestinal tumorigenesis. In vitro studies suggest that flavaglines target prohibitin 1 (PHB1) as a ligand, but this has not been established in the intestine. PHB1 is a highly conserved protein with diverse functions that depend on its posttranslational modifications and subcellular localization. Here, we demonstrate that FL3 combats intestinal tumorigenesis in the azoxymethane-dextran sodium sulfate and ApcMin/+ mouse models and in human colorectal cancer tumor organoids (tumoroids) by inhibiting Wnt/β-catenin signaling via induction of Axin1 expression. FL3 exhibited no change in cell viability in normal intestinal epithelial cells or human matched-normal colonoids. FL3 response was diminished in colorectal cancer cell lines and human colorectal cancer tumoroids harboring a mutation at S45 of β-catenin. PHB1 deficiency in mice or in human colorectal cancer tumoroids abolished FL3-induced expression of Axin1 and drove tumoroid death. In colorectal cancer cells, FL3 treatment blocked phosphorylation of PHB1 at Thr258, resulting in its nuclear translocation and binding to the Axin1 promoter. These results suggest that FL3 inhibits Wnt/β-catenin signaling via PHB1-dependent activation of Axin1. FL3, therefore, represents a novel compound that combats Wnt pathway–dependent cancers, such as colorectal cancer.
Significance:
Targeting of PHB1 by FL3 provides a novel mechanism to combat Wnt-driven cancers, with limited intestinal toxicity.
Data from Enhanced Efficacy of Simultaneous PD-1 and PD-L1 Immune Checkpoint Blockade in High-Grade Serous Ovarian Cancer
Authors: Wan Changxin , Keany Matthew P. , Dong Han , Al-Alem Linah F. , Pandya Unnati M. , Lazo Suzan , Boehnke Karsten , Lynch Katherine N. , Xu Rui , Zarrella Dominique T. , Gu Shengqing , Cejas Paloma , Lim Klothilda , Long Henry W. , Elias Kevin M. , Horowitz Neil S. , Feltmate Colleen M. , Muto Michael G. , Worley Michael J. , Berkowitz Ross S. , Matulonis Ursula A. , Nucci Marisa R. , Crum Christopher P. , Rueda Bo R. , Brown Myles , Liu Xiaole Shirley , Hill Sarah J.
Published: March 2023

Abstract
Immune therapies have had limited efficacy in high-grade serous ovarian cancer (HGSC), as the cellular targets and mechanism(s) of action of these agents in HGSC are unknown. Here we performed immune functional and single-cell RNA sequencing transcriptional profiling on novel HGSC organoid/immune cell co-cultures treated with a unique bispecific anti-programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) antibody compared with monospecific anti-PD-1 or anti-PD-L1 controls. Comparing the functions of these agents across all immune cell types in real time identified key immune checkpoint blockade (ICB) targets that have eluded currently available monospecific therapies. The bispecific antibody induced superior cellular state changes in both T and natural killer (NK) cells. It uniquely induced NK cells to transition from inert to more active and cytotoxic phenotypes, implicating NK cells as a key missing component of the current ICB-induced immune response in HGSC. It also induced a subset of CD8 T cells to transition from naïve to more active and cytotoxic progenitor-exhausted phenotypes post-treatment, revealing the small, previously uncharacterized population of CD8 T cells responding to ICB in HGSC. These state changes were driven partially through bispecific antibody-induced downregulation of the bromodomain-containing protein BRD1. Small-molecule inhibition of BRD1 induced similar state changes in vitro and demonstrated efficacy in vivo, validating the co-culture results. Our results demonstrate that state changes in both NK and a subset of T cells may be critical in inducing an effective anti-tumor immune response and suggest that immune therapies able to induce such cellular state changes, such as BRD1 inhibitors, may have increased efficacy in HGSC.
Significance:
This study indicates that increased efficacy of immune therapies in ovarian cancer is driven by state changes of NK and small subsets of CD8 T cells into active and cytotoxic states.
Data from The Cancer-Associated ATM R3008H Mutation Reveals the Link between ATM Activation and Its Exchange
Authors: Milanovic Maja , Houghton Lisa M. , Menolfi Demis , Lee Ji-Hoon , Yamamoto Kenta , Li Yang , Lee Brian J. , Xu Jun , Estes Verna M. , Wang Dong , Mckinnon Peter J. , Paull Tanya T. , Zha Shan
Published: March 2023

Abstract
ATM kinase is a tumor suppressor and a master regulator of the DNA damage response. Most cancer-associated alterations to ATM are missense mutations at the PI3-kinase regulatory domain (PRD) or the kinase domain. Expression of kinase-dead (KD) ATM protein solely accelerates lymphomagenesis beyond ATM loss. To understand how PRD suppresses lymphomagenesis, we introduced the cancer-associated PRD mutation R3008H (R3016 in mouse) into mice. R3008H abrogated DNA damage- and oxidative stress-induced activation of ATM without consistently affecting ATM protein stability and recruitment. In contrast to the early embryonic lethality of AtmKD/KD mice, AtmR3016H (AtmR/R) mice were viable, immunodeficient, and displayed spontaneous craniofacial abnormalities and delayed lymphomagenesis compared with Atm−/− controls. Mechanistically, R3008H rescued the tardy exchange of ATM-KD at DNA damage foci, indicating that PRD coordinates ATM activation with its exchange at DNA-breaks. Taken together, our results reveal a unique tumorigenesis profile for PRD mutations that is distinct from null or KD mutations.
Significant:
This study functionally characterizes the most common ATM missense mutation R3008H in cancer and identifies a unique role of PI3-kinase regulatory domain in ATM activation.
Data from Frizzled-7 Identifies Platinum-Tolerant Ovarian Cancer Cells Susceptible to Ferroptosis
Authors: Wang Yinu , Zhao Guangyuan , Condello Salvatore , Huang Hao , Cardenas Horacio , Tanner Edward J. , Wei JianJun , Ji Yanrong , Li Junjie , Tan Yuying , Davuluri Ramana V. , Peter Marcus E. , Cheng Ji-Xin , Matei Daniela
Published: March 2023

Abstract
Defining traits of platinum-tolerant cancer cells could expose new treatment vulnerabilities. Here, new markers associated with platinum-tolerant cells and tumors were identified using in vitro and in vivo ovarian cancer models treated repetitively with carboplatin and validated in human specimens. Platinum-tolerant cells and tumors were enriched in ALDH+ cells, formed more spheroids, and expressed increased levels of stemness-related transcription factors compared with parental cells. Additionally, platinum-tolerant cells and tumors exhibited expression of the Wnt receptor Frizzled-7 (FZD7). Knockdown of FZD7 improved sensitivity to platinum, decreased spheroid formation, and delayed tumor initiation. The molecular signature distinguishing FZD7+ from FZD7− cells included epithelial-to-mesenchymal (EMT), stemness, and oxidative phosphorylation-enriched gene sets. Overexpression of FZD7 activated the oncogenic factor Tp63, driving upregulation of glutathione metabolism pathways, including glutathione peroxidase 4 (GPX4), which protected cells from chemotherapy-induced oxidative stress. FZD7+ platinum-tolerant ovarian cancer cells were more sensitive and underwent ferroptosis after treatment with GPX4 inhibitors. FZD7, Tp63, and glutathione metabolism gene sets were strongly correlated in the ovarian cancer Tumor Cancer Genome Atlas (TCGA) database and in residual human ovarian cancer specimens after chemotherapy. These results support the existence of a platinum-tolerant cell population with partial cancer stem cell features, characterized by FZD7 expression and dependent on the FZD7–β-catenin–Tp63–GPX4 pathway for survival. The findings reveal a novel therapeutic vulnerability of platinum-tolerant cancer cells and provide new insight into a potential “persister cancer cell” phenotype.
Significance:
Frizzled-7 marks platinum-tolerant cancer cells harboring stemness features and altered glutathione metabolism that depend on GPX4 for survival and are highly susceptible to ferroptosis.
Data from KDM5B Is Essential for the Hyperactivation of PI3K/AKT Signaling in Prostate Tumorigenesis
Authors: Li Guoliang , Kanagasabai Thanigaivelan , Lu Wenfu , Zou Mike R. , Zhang Shang-Min , Celada Sherly I. , Izban Michael G. , Liu Qi , Lu Tao , Ballard Billy R. , Zhou Xinchun , Adunyah Samuel E. , Matusik Robert J. , Yan Qin , Chen Zhenbang
Published: March 2023

Abstract
KDM5B (lysine[K]-specific demethylase 5B) is frequently upregulated in various human cancers including prostate cancer. KDM5B controls H3K4me3/2 levels and regulates gene transcription and cell differentiation, yet the contributions of KDM5B to prostate cancer tumorigenesis remain unknown. In this study, we investigated the functional role of KDM5B in epigenetic dysregulation and prostate cancer progression in cultured cells and in mouse models of prostate epithelium–specific mutant Pten/Kdm5b. Kdm5b deficiency resulted in a significant delay in the onset of prostate cancer in Pten-null mice, whereas Kdm5b loss alone caused no morphologic abnormalities in mouse prostates. At 6 months of age, the prostate weight of Pten/Kdm5b mice was reduced by up to 70% compared with that of Pten mice. Pathologic analysis revealed Pten/Kdm5b mice displayed mild morphologic changes with hyperplasia in prostates, whereas age-matched Pten littermates developed high-grade prostatic intraepithelial neoplasia and prostate cancer. Mechanistically, KDM5B governed PI3K/AKT signaling in prostate cancer in vitro and in vivo. KDM5B directly bound the PIK3CA promoter, and KDM5B knockout resulted in a significant reduction of P110α and PIP3 levels and subsequent decrease in proliferation of human prostate cancer cells. Conversely, KDM5B overexpression resulted in increased PI3K/AKT signaling. Loss of Kdm5b abrogated the hyperactivation of AKT signaling by decreasing P110α/P85 levels in Pten/Kdm5b mice. Taken together, our findings reveal that KDM5B acts as a key regulator of PI3K/AKT signaling; they also support the concept that targeting KDM5B is a novel and effective therapeutic strategy against prostate cancer.
Significance:
This study demonstrates that levels of histone modification enzyme KDM5B determine hyperactivation of PI3K/AKT signaling in prostate cancer and that targeting KDM5B could be a novel strategy against prostate cancer.
Data from Misannotated Multi-Nucleotide Variants in Public Cancer Genomics Datasets Lead to Inaccurate Mutation Calls with Significant Implications
Authors: Srinivasan Sujaya , Kalinava Natallia , Aldana Rafael , Li Zhipan , van Hagen Sjoerd , Rodenburg Sander Y.A. , Wind-Rotolo Megan , Qian Xiaozhong , Sasson Ariella S. , Tang Hao , Kirov Stefan
Published: March 2023

Abstract
Although next-generation sequencing is widely used in cancer to profile tumors and detect variants, most somatic variant callers used in these pipelines identify variants at the lowest possible granularity, single-nucleotide variants (SNV). As a result, multiple adjacent SNVs are called individually instead of as a multi-nucleotide variants (MNV). With this approach, the amino acid change from the individual SNV within a codon could be different from the amino acid change based on the MNV that results from combining SNV, leading to incorrect conclusions about the downstream effects of the variants. Here, we analyzed 10,383 variant call files (VCF) from the Cancer Genome Atlas (TCGA) and found 12,141 incorrectly annotated MNVs. Analysis of seven commonly mutated genes from 178 studies in cBioPortal revealed that MNVs were consistently missed in 20 of these studies, whereas they were correctly annotated in 15 more recent studies. At the BRAF V600 locus, the most common example of MNV, several public datasets reported separate BRAF V600E and BRAF V600M variants instead of a single merged V600K variant. VCFs from the TCGA Mutect2 caller were used to develop a solution to merge SNV to MNV. Our custom script used the phasing information from the SNV VCF and determined whether SNVs were at the same codon and needed to be merged into MNV before variant annotation. This study shows that institutions performing NGS sequencing for cancer genomics should incorporate the step of merging MNV as a best practice in their pipelines.
Significance:
Identification of incorrect mutation calls in TCGA, including clinically relevant BRAF V600 and KRAS G12, will influence research and potentially clinical decisions.