bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Differential Gain-of-Function Activity of Three p53 Hotspot Mutants In Vivo
Authors: Xiong Shunbin , Chachad Dhruv , Zhang Yun , Gencel-Augusto Jovanka , Sirito Mario , Pant Vinod , Yang Peirong , Sun Chang , Chau Gilda , Qi Yuan , Su Xiaoping , Whitley Elizabeth M. , El-Naggar Adel K. , Lozano Guillermina
Published: March 2023

Abstract
The majority of TP53 missense mutations identified in cancer patients are in the DNA-binding domain and are characterized as either structural or contact mutations. These missense mutations exhibit inhibitory effects on wild-type p53 activity. More importantly, these mutations also demonstrate gain-of-function (GOF) activities characterized by increased metastasis, poor prognosis, and drug resistance. To better understand the activities by which TP53 mutations, identified in Li–Fraumeni syndrome, contribute to tumorigenesis, we generated mice harboring a novel germline Trp53R245W allele (contact mutation) and compared them with existing models with Trp53R172H (structural mutation) and Trp53R270H (contact mutation) alleles. Thymocytes from heterozygous mice showed that all three hotspot mutations exhibited similar inhibitory effects on wild-type p53 transcription in vivo, and tumors from these mice had similar levels of loss of heterozygosity. However, the overall survival of Trp53R245W/+ and Trp53R270H/+ mice, but not Trp53R172H/+ mice, was significantly shorter than that of Trp53+/− mice, providing strong evidence for p53-mutant–specific GOF contributions to tumor development. Furthermore, Trp53R245W/+ and Trp53R270H/+ mice had more osteosarcoma metastases than Trp53R172H/+ mice, suggesting that these two contact mutants have stronger GOF in driving osteosarcoma metastasis. Transcriptomic analyses using RNA sequencing data from Trp53R172H/+, Trp53R245W/+, and Trp53R270H/+ primary osteosarcomas in comparison with Trp53+/– indicated that GOF of the three mutants was mediated by distinct pathways. Thus, both the inhibitory effect of mutant over wild-type p53 and GOF activities of mutant p53 contributed to tumorigenesis in vivo. Targeting p53 mutant–specific pathways may be important for therapeutic outcomes in osteosarcoma.
Significance:
p53 hotspot mutants inhibit wild-type p53 similarly but differ in their GOF activities, with stronger tumor-promoting activity in contact mutants and distinct protein partners of each mutant driving tumorigenesis and metastasis.
Data from Reorganization of the 3D Genome Pinpoints Noncoding Drivers of Primary Prostate Tumors
Authors: Hawley James R. , Zhou Stanley , Arlidge Christopher , Grillo Giacomo , Kron Ken J. , Hugh-White Rupert , van der Kwast Theodorus H. , Fraser Michael , Boutros Paul C. , Bristow Robert G. , Lupien Mathieu
Published: March 2023

Abstract
Prostate cancer is a heterogeneous disease whose progression is linked to genome instability. However, the impact of this instability on the noncoding genome and its three-dimensional organization to aid progression is unclear. Using primary benign and tumor tissue, we find a high concordance in higher-order three-dimensional genome organization. This concordance argues for constraints to the topology of prostate tumor genomes. Nonetheless, we identified changes in focal chromatin interactions, typical of loops bridging noncoding cis-regulatory elements, and showed how structural variants can induce these changes to guide cis-regulatory element hijacking. Such events resulted in opposing differential expression of genes found at antipodes of rearrangements. Collectively, these results argue that changes to focal chromatin interactions, as opposed to higher-order genome organization, allow for aberrant gene regulation and are repeatedly mediated by structural variants in primary prostate cancer.
Significance:
This work showcases how the noncoding genome can be hijacked by focal insults to its three-dimensional organization that contribute to prostate cancer oncogenesis.
Data from Inhibition of PIM Kinases in DLBCL Targets MYC Transcriptional Program and Augments the Efficacy of Anti-CD20 Antibodies
Authors: Szydłowski Maciej , Garbicz Filip , Jabłońska Ewa , Górniak Patryk , Komar Dorota , Pyrzyńska Beata , Bojarczuk Kamil , Prochorec-Sobieszek Monika , Szumera-Ciećkiewicz Anna , Rymkiewicz Grzegorz , Cybulska Magdalena , Statkiewicz Małgorzata , Gajewska Marta , Mikula Michał , Gołas Aniela , Domagała Joanna , Winiarska Magdalena , Graczyk-Jarzynka Agnieszka , Białopiotrowicz Emilia , Polak Anna , Barankiewicz Joanna , Puła Bartosz , Pawlak Michał , Nowis Dominika , Golab Jakub , Tomirotti Andrea M. , Brzózka Krzysztof , Pacheco-Blanco Mariana , Kupcova Kristyna , Green Michael R. , Havranek Ondrej , Chapuy Bjoern , Juszczyński Przemysław
Published: March 2023

Abstract
The family of PIM serine/threonine kinases includes three highly conserved oncogenes, PIM1, PIM2, and PIM3, which regulate multiple prosurvival pathways and cooperate with other oncogenes such as MYC. Recent genomic CRISPR-Cas9 screens further highlighted oncogenic functions of PIMs in diffuse large B-cell lymphoma (DLBCL) cells, justifying the development of small-molecule PIM inhibitors and therapeutic targeting of PIM kinases in lymphomas. However, detailed consequences of PIM inhibition in DLBCL remain undefined. Using chemical and genetic PIM blockade, we comprehensively characterized PIM kinase–associated prosurvival functions in DLBCL and the mechanisms of PIM inhibition–induced toxicity. Treatment of DLBCL cells with SEL24/MEN1703, a pan-PIM inhibitor in clinical development, decreased BAD phosphorylation and cap-dependent protein translation, reduced MCL1 expression, and induced apoptosis. PIM kinases were tightly coexpressed with MYC in diagnostic DLBCL biopsies, and PIM inhibition in cell lines and patient-derived primary lymphoma cells decreased MYC levels as well as expression of multiple MYC-dependent genes, including PLK1. Chemical and genetic PIM inhibition upregulated surface CD20 levels in an MYC-dependent fashion. Consistently, MEN1703 and other clinically available pan-PIM inhibitors synergized with the anti-CD20 monoclonal antibody rituximab in vitro, increasing complement-dependent cytotoxicity and antibody-mediated phagocytosis. Combined treatment with PIM inhibitor and rituximab suppressed tumor growth in lymphoma xenografts more efficiently than either drug alone. Taken together, these results show that targeting PIM in DLBCL exhibits pleiotropic effects that combine direct cytotoxicity with potentiated susceptibility to anti-CD20 antibodies, justifying further clinical development of such combinatorial strategies.
Significance:
These findings demonstrate that inhibition of PIM induces DLBCL cell death via MYC-dependent and -independent mechanisms and enhances the therapeutic response to anti-CD20 antibodies by increasing CD20 expression.
Data from Neoadjuvant Intratumoral Immunotherapy with TLR9 Activation and Anti-OX40 Antibody Eradicates Metastatic Cancer
Authors: Hong Wan Xing , Sagiv-Barfi Idit , Czerwinski Debra K. , Sallets Adrienne , Levy Ronald
Published: March 2023

Abstract
The combination of the synthetic TLR9 ligand CpG and agnostic OX40 antibody can trigger systemic antitumor immune responses upon co-injection into the tumor microenvironment, eradicating simultaneous untreated sites of metastatic disease. Here we explore the application of this in situ immunotherapy to the neoadjuvant setting. Current neoadjuvant checkpoint blockade therapy is delivered systemically, resulting in off-target adverse effects. In contrast, intratumoral immunotherapy minimizes the potential for toxicities and allows for greater development of combination therapies. In two metastatic solid tumor models, neoadjuvant intratumoral immunotherapy generated a local T-cell antitumor response that then acted systemically to attack cancer throughout the body. In addition, the importance of timing between neoadjuvant immunotherapy and surgical resection was established, as well as the increased therapeutic power of adding systemic anti-PD1 antibody. The combination of local and systemic immunotherapy generated an additional survival benefit due to synergistic inhibitory effect on tumor-associated macrophages. These results provide a strong rationale for translating this neoadjuvant intratumoral immunotherapy to the clinical setting, especially in conjunction with established checkpoint inhibitors.
Significance:
This work demonstrates the ability of neoadjuvant intratumoral immunotherapy to target local and distant metastatic disease and consequently improve survival.
Data from Dissecting the Oncogenic Roles of Keratin 17 in the Hallmarks of Cancer
Authors: Baraks Gabriella , Tseng Robert , Pan Chun-Hao , Kasliwal Saumya , Leiton Cindy V. , Shroyer Kenneth R. , Escobar-Hoyos Luisa F.
Published: March 2023

Abstract
There is an unmet need to identify and validate tumor-specific therapeutic targets to enable more effective treatments for cancer. Heterogeneity in patient clinical characteristics as well as biological and genetic features of tumors present major challenges for the optimization of therapeutic interventions, including the development of novel and more effective precision medicine. The expression of keratin 17 (K17) is a hallmark of the most aggressive forms of cancer across a wide range of anatomical sites and histological types. K17 correlates with shorter patient survival, predicts resistance to specific chemotherapeutic agents, and harbors functional domains that suggest it could be therapeutically targeted. Here, we explore the role of K17 in the hallmarks of cancer and summarize evidence to date for K17-mediated mechanisms involved in each hallmark, elucidating functional roles that warrant further investigation to guide the development of novel therapeutic strategies.
Data from Antibody-Peptide Epitope Conjugates for Personalized Cancer Therapy
Authors: Zhang Songfa , Yan Chuan , Millar David G. , Yang Qiqi , Heather James M. , Langenbucher Adam , Morton Laura T. , Sepulveda Sean , Alpert Eric , Whelton Lauren R. , Zarrella Dominique T. , Guo Mei , Minogue Eleanor , Lawrence Michael S. , Rueda Bo R. , Spriggs David R. , Lu Weiguo , Langenau David M. , Cobbold Mark
Published: March 2023

Abstract
Antibody–peptide epitope conjugates (APEC) are a new class of modified antibody–drug conjugates that redirect T-cell viral immunity against tumor cells. APECs contain a tumor-specific protease cleavage site linked to a patient-specific viral epitope, resulting in presentation of viral epitopes on cancer cells and subsequent recruitment and killing by CD8+ T cells. Here we developed an experimental pipeline to create patient-specific APECs and identified new preclinical therapies for ovarian carcinoma. Using functional assessment of viral peptide antigen responses to common viruses like cytomegalovirus (CMV) in patients with ovarian cancer, a library of 192 APECs with distinct protease cleavage sequences was created using the anti-epithelial cell adhesion molecule (EpCAM) antibody. Each APEC was tested for in vitro cancer cell killing, and top candidates were screened for killing xenograft tumors grown in zebrafish and mice. These preclinical modeling studies identified EpCAM-MMP7-CMV APEC (EpCAM-MC) as a potential new immunotherapy for ovarian carcinoma. Importantly, EpCAM-MC also demonstrated robust T-cell responses in primary ovarian carcinoma patient ascites samples. This work highlights a robust, customizable platform to rapidly develop patient-specific APECs.
Significance:
This study develops a high-throughput preclinical platform to identify patient-specific antibody–peptide epitope conjugates that target cancer cells and demonstrates the potential of this immunotherapy approach for treating ovarian carcinoma.
Data from Multiomics Analysis of Spatially Distinct Stromal Cells Reveals Tumor-Induced O-Glycosylation of the CDK4–pRB Axis in Fibroblasts at the Invasive Tumor Edge
Authors: Bouchard Gina , Garcia-Marques Fernando Jose , Karacosta Loukia Georgiou , Zhang Weiruo , Bermudez Abel , Riley Nicholas McIlvain , Varma Sushama , Mehl Lindsey Catherine , Benson Jalen Anthony , Shrager Joseph B. , Bertozzi Carolyn Ruth , Pitteri Sharon J. , Giaccia Amato J. , Plevritis Sylvia Katina
Published: March 2023

Abstract
The invasive leading edge represents a potential gateway for tumor metastasis. The role of fibroblasts from the tumor edge in promoting cancer invasion and metastasis has not been comprehensively elucidated. We hypothesize that cross-talk between tumor and stromal cells within the tumor microenvironment results in activation of key biological pathways depending on their position in the tumor (edge vs. core). Here we highlight phenotypic differences between tumor-adjacent-fibroblasts (TAF) from the invasive edge and tumor core fibroblasts from the tumor core, established from human lung adenocarcinomas. A multiomics approach that includes genomics, proteomics, and O-glycoproteomics was used to characterize cross-talk between TAFs and cancer cells. These analyses showed that O-glycosylation, an essential posttranslational modification resulting from sugar metabolism, alters key biological pathways including the cyclin-dependent kinase 4 (CDK4) and phosphorylated retinoblastoma protein axis in the stroma and indirectly modulates proinvasive features of cancer cells. In summary, the O-glycoproteome represents a new consideration for important biological processes involved in tumor–stroma cross-talk and a potential avenue to improve the anticancer efficacy of CDK4 inhibitors.
Significance:
A multiomics analysis of spatially distinct fibroblasts establishes the importance of the stromal O-glycoproteome in tumor–stroma interactions at the leading edge and provides potential strategies to improve cancer treatment.

See related commentary by De Wever, p. 537
Data from p16 Represses DNA Damage Repair via a Novel Ubiquitin-Dependent Signaling Cascade
Authors: Molkentine David P. , Molkentine Jessica M. , Bridges Kathleen A. , Valdecanas David R. , Dhawan Annika , Bahri Reshub , Hefner Andrew J. , Kumar Manish , Yang Liangpeng , Abdelhakiem Mohamed , Pifer Phillip M. , Sandulache Vlad , Sheth Aakash , Beadle Beth M. , Thames Howard D. , Mason Kathryn A. , Pickering Curtis R. , Meyn Raymond E. , Skinner Heath D.
Published: March 2023

Abstract
Squamous cell carcinoma driven by human papillomavirus (HPV) is more sensitive to DNA-damaging therapies than its HPV-negative counterpart. Here, we show that p16, the clinically used surrogate for HPV positivity, renders cells more sensitive to radiotherapy via a ubiquitin-dependent signaling pathway, linking high levels of this protein to increased activity of the transcription factor SP1, increased HUWE1 transcription, and degradation of ubiquitin-specific protease 7 (USP7) and TRIP12. Activation of this pathway in HPV-positive disease led to decreased homologous recombination and improved response to radiotherapy, a phenomenon that can be recapitulated in HPV-negative disease using USP7 inhibitors in clinical development. This p16-driven axis induced sensitivity to PARP inhibition and potentially leads to “BRCAness” in head and neck squamous cell carcinoma (HNSCC) cells. Thus, these findings support a functional role for p16 in HPV-positive tumors in driving response to DNA damage, which can be exploited to improve outcomes in both patients with HPV-positive and HPV-negative HNSCC.
Significance:
In HPV-positive tumors, a previously undiscovered pathway directly links p16 to DNA damage repair and sensitivity to radiotherapy via a clinically relevant and pharmacologically targetable ubiquitin-mediated degradation pathway.
Data from Lactate Rewires Lipid Metabolism and Sustains a Metabolic–Epigenetic Axis in Prostate Cancer
Authors: Ippolito Luigi , Comito Giuseppina , Parri Matteo , Iozzo Marta , Duatti Assia , Virgilio Francesca , Lorito Nicla , Bacci Marina , Pardella Elisa , Sandrini Giada , Bianchini Francesca , Damiano Roberta , Ferrone Lavinia , la Marca Giancarlo , Serni Sergio , Spatafora Pietro , Catapano Carlo V. , Morandi Andrea , Giannoni Elisa , Chiarugi Paola
Published: March 2023

Abstract
Lactate is an abundant oncometabolite in the tumor environment. In prostate cancer, cancer-associated fibroblasts (CAF) are major contributors of secreted lactate, which can be taken up by cancer cells to sustain mitochondrial metabolism. However, how lactate impacts transcriptional regulation in tumors has yet to be fully elucidated. Here, we describe a mechanism by which CAF-secreted lactate is able to increase the expression of genes involved in lipid metabolism in prostate cancer cells. This regulation enhanced intracellular lipid accumulation in lipid droplets (LD) and provided acetyl moieties for histone acetylation, establishing a regulatory loop between metabolites and epigenetic modification. Inhibition of this loop by targeting the bromodomain and extraterminal protein family of histone acetylation readers suppressed the expression of perilipin 2 (PLIN2), a crucial component of LDs, disrupting lactate-dependent lipid metabolic rewiring. Inhibition of this CAF-induced metabolic–epigenetic regulatory loop in vivo reduced growth and metastasis of prostate cancer cells, demonstrating its translational relevance as a therapeutic target in prostate cancer. Clinically, PLIN2 expression was elevated in tumors with a higher Gleason grade and in castration-resistant prostate cancer compared with primary prostate cancer. Overall, these findings show that lactate has both a metabolic and an epigenetic role in promoting prostate cancer progression.
Significance:
This work shows that stromal-derived lactate induces accumulation of lipid droplets, stimulates epigenetic rewiring, and fosters metastatic potential in prostate cancer.
Data from Comprehensive Analysis of Metabolic Isozyme Targets in Cancer
Authors: Marczyk Michal , Gunasekharan Vignesh , Casadevall David , Qing Tao , Foldi Julia , Sehgal Raghav , Shan Naing Lin , Blenman Kim R.M. , O'Meara Tess A. , Umlauf Sheila , Surovtseva Yulia V. , Muthusamy Viswanathan , Rinehart Jesse , Perry Rachel J. , Kibbey Richard , Hatzis Christos , Pusztai Lajos
Published: March 2023

Abstract
Metabolic reprogramming is a hallmark of malignant transformation, and loss of isozyme diversity (LID) contributes to this process. Isozymes are distinct proteins that catalyze the same enzymatic reaction but can have different kinetic characteristics, subcellular localization, and tissue specificity. Cancer-dominant isozymes that catalyze rate-limiting reactions in critical metabolic processes represent potential therapeutic targets. Here, we examined the isozyme expression patterns of 1,319 enzymatic reactions in 14 cancer types and their matching normal tissues using The Cancer Genome Atlas mRNA expression data to identify isozymes that become cancer-dominant. Of the reactions analyzed, 357 demonstrated LID in at least one cancer type. Assessment of the expression patterns in over 600 cell lines in the Cancer Cell Line Encyclopedia showed that these reactions reflect cellular changes instead of differences in tissue composition; 50% of the LID-affected isozymes showed cancer-dominant expression in the corresponding cell lines. The functional importance of the cancer-dominant isozymes was assessed in genome-wide CRISPR and RNAi loss-of-function screens: 17% were critical for cell proliferation, indicating their potential as therapeutic targets. Lists of prioritized novel metabolic targets were developed for 14 cancer types; the most broadly shared and functionally validated target was acetyl-CoA carboxylase 1 (ACC1). Small molecule inhibition of ACC reduced breast cancer viability in vitro and suppressed tumor growth in cell line– and patient-derived xenografts in vivo. Evaluation of the effects of drug treatment revealed significant metabolic and transcriptional perturbations. Overall, this systematic analysis of isozyme expression patterns elucidates an important aspect of cancer metabolic plasticity and reveals putative metabolic vulnerabilities.
Significance:
This study exploits the loss of metabolic isozyme diversity common in cancer and reveals a rich pool of potential therapeutic targets that will allow the repurposing of existing inhibitors for anticancer therapy.

See related commentary by Kehinde and Parker, p. 1695
Data from Patient-Derived Triple-Negative Breast Cancer Organoids Provide Robust Model Systems That Recapitulate Tumor Intrinsic Characteristics
Authors: Bhatia Sonam , Kramer Melissa , Russo Suzanne , Naik Payal , Arun Gayatri , Brophy Kyle , Andrews Peter , Fan Cheng , Perou Charles M. , Preall Jonathan , Ha Taehoon , Plenker Dennis , Tuveson David A. , Rishi Arvind , Wilkinson John E. , McCombie W. Richard , Kostroff Karen , Spector David L.
Published: March 2023

Abstract
Triple-negative breast cancer (TNBC) is an aggressive form of breast cancer with poor patient outcomes, highlighting the unmet clinical need for targeted therapies and better model systems. Here, we developed and comprehensively characterized a diverse biobank of normal and breast cancer patient-derived organoids (PDO) with a focus on TNBCs. PDOs recapitulated patient tumor intrinsic properties and a subset of PDOs can be propagated for long-term culture (LT-TNBC). Single cell profiling of PDOs identified cell types and gene candidates affiliated with different aspects of cancer progression. The LT-TNBC organoids exhibit signatures of aggressive MYC-driven, basal-like breast cancers and are largely comprised of luminal progenitor (LP)-like cells. The TNBC LP-like cells are distinct from normal LPs and exhibit hyperactivation of NOTCH and MYC signaling. Overall, this study validates TNBC PDOs as robust models for understanding breast cancer biology and progression, paving the way for personalized medicine and tailored treatment options.
Significance:
A comprehensive analysis of patient-derived organoids of TNBC provides insights into cellular heterogeneity and mechanisms of tumorigenesis at the single-cell level.
Data from CRISPR/Cas9-Induced DNA Damage Enriches for Mutations in a p53-Linked Interactome: Implications for CRISPR-Based Therapies
Authors: Jiang Long , Ingelshed Katrine , Shen Yunbing , Boddul Sanjaykumar V. , Iyer Vaishnavi Srinivasan , Kasza Zsolt , Sedimbi Saikiran , Lane David P. , Wermeling Fredrik
Published: March 2023

Abstract
Inactivating p53 mutations are the most abundant genetic alterations found in cancer. Here we show that CRISPR/Cas9-induced double-stranded DNA breaks enrich for cells deficient in p53 and in genes of a core CRISPR–p53 tumor suppressor interactome. Such enrichment could predispose to cancer development and thus pose a challenge for clinical CRISPR use. Transient p53 inhibition could suppress the enrichment of cells with these mutations. The level of DNA damage response induced by an sgRNA influenced the enrichment of p53-deficient cells and could be a relevant parameter in sgRNA design to limit cellular enrichment. Furthermore, a dataset of >800 human cancer cell lines identified additional factors influencing the enrichment of p53-mutated cells, including strong baseline CDKN1A expression as a predictor for an active CRISPR–p53 axis. Taken together, these data provide details about p53 biology in the context of CRISPR-induced DNA damage and identify strategies to enable safer CRISPR use.
Significance:
CRISPR-mediated DNA damage enriches for cells with escape mutations in a core CRISPR–p53 interactome, which can be suppressed by transient inhibition of p53.
Data from Structure, Dynamics, and Impact of Replication Stress–Induced Structural Variants in Hepatocellular Carcinoma
Authors: Bayard Quentin , Cordier Pierre , Péneau Camille , Imbeaud Sandrine , Hirsch Theo Z. , Renault Victor , Nault Jean-Charles , Blanc Jean-Frédéric , Calderaro Julien , Desdouets Chantal , Zucman-Rossi Jessica , Letouzé Eric
Published: March 2023

Abstract
Oncogene activation leads to replication stress and promotes genomic instability. Here we combine optical mapping and whole-genome sequencing (WGS) to explore in depth the nature of structural variants (SV) induced by replication stress in cyclin-activated hepatocellular carcinomas (CCN-HCC). In addition to classical tandem duplications, CCN-HCC displayed frequent intra-chromosomal and interchromosomal templated insertion cycles (TIC), likely resulting from template switching events. Template switching preferentially involves active topologically associated domains that are proximal to one another within the 3D genome. Template sizes depend on the type of cyclin activation and are coordinated within each TIC. Replication stress induced continuous accumulation of SVs during CCN-HCC progression, fostering the acquisition of new driver alterations and large-scale copy-number changes at TIC borders. Together, this analysis sheds light on the mechanisms, dynamics, and consequences of SV accumulation in tumors with oncogene-induced replication stress.
Significance:
Optical mapping and whole-genome sequencing integration unravels a unique signature of replication stress–induced structural variants that drive genomic evolution and the acquisition of driver events in CCN-HCC.
Data from Aberrant RNA Splicing Is a Primary Link between Genetic Variation and Pancreatic Cancer Risk
Authors: Tian Jianbo , Chen Can , Rao Meilin , Zhang Ming , Lu Zequn , Cai Yimin , Ying Pingting , Li Bin , Wang Haoxue , Wang Lu , Li Yao , Huang Jinyu , Fan Linyun , Cai Xiaomin , Ning Caibo , Li Yanmin , Zhang Fuwei , Wang Wenzhuo , Jiang Yuan , Liu Yizhuo , Wang Min , Li Heng , Huang Chaoqun , Yang Zhiyong , Chang Jiang , Zhu Ying , Yang Xiaojun , Miao Xiaoping
Published: March 2023

Abstract
Understanding the genetic variation underlying transcript splicing is essential for fully dissecting the molecular mechanisms of common diseases. The available evidence from splicing quantitative trait locus (sQTL) studies using pancreatic ductal adenocarcinoma (PDAC) tissues have been limited to small sample sizes. Here we present a genome-wide sQTL analysis to identify SNP that control mRNA splicing in 176 PDAC samples from TCGA. From this analysis, 16,175 sQTLs were found to be significantly enriched in RNA-binding protein (RBP) binding sites and chromatin regulatory elements and overlapped with known loci from PDAC genome-wide association studies (GWAS). sQTLs and expression quantitative trait loci (eQTL) showed mostly nonoverlapping patterns, suggesting sQTLs provide additional insights into the etiology of disease. Target genes affected by sQTLs were closely related to cancer signaling pathways, high mutational burden, immune infiltration, and pharmaceutical targets, which will be helpful for clinical applications. Integration of a large-scale population consisting of 2,782 patients with PDAC and 7,983 healthy controls identified an sQTL variant rs1785932-T allele that promotes alternative splicing of ELP2 exon 6 and leads to a lower level of the ELP2 full-length isoform (ELP2_V1) and a higher level of a truncated ELP2 isoform (ELP2_V2), resulting in decreased risk of PDAC [OR = 0.83; 95% confidence interval (CI), 0.77–0.89; P = 1.16 × 10−6]. The ELP2_V2 isoform functioned as a potential tumor suppressor gene, inhibiting PDAC cell proliferation by exhibiting stronger binding affinity to JAK1/STAT3 than ELP2_V1 and subsequently blocking the pathologic activation of the phosphorylated STAT3 (pSTAT3) pathway. Collectively, these findings provide an informative sQTL resource and insights into the regulatory mechanisms linking splicing variants to PDAC risk.
Significance:
In pancreatic cancer, splicing quantitative trait loci analysis identifies a rs1785932 variant that contributes to decreased risk of disease by influencing ELP2 mRNA splicing and blocking the STAT3 oncogenic pathway.
Data from A Pan-Cancer Immunogenomic Atlas for Immune Checkpoint Blockade Immunotherapy
Authors: Yang Jing , Zhao Shilin , Wang Jing , Sheng Quanhu , Liu Qi , Shyr Yu
Published: March 2023

Abstract
The ability to identify robust genomic signatures that predict response to immune checkpoint blockade is restricted by limited sample sizes and ungeneralizable performance across cohorts. To address these challenges, we established Cancer-Immu (http://bioinfo.vanderbilt.edu/database/Cancer-Immu/), a comprehensive platform that integrates large-scale multidimensional omics data, including genetic, bulk, and single-cell transcriptomic, proteomic, and dynamic genomic profiles, with clinical phenotypes to explore consistent and rare immunogenomic connections. Currently Cancer-Immu has incorporated data for 3,652 samples for 16 cancer types. It provides easy access to immunogenomic data and empowers researchers to translate omics datasets into biological insights and clinical applications.
Significance:
Cancer-Immu is a comprehensive functional portal for unraveling immune-genomic connections to improve immune checkpoint blockade–based cancer immunotherapy.
Data from In Vivo Modeling of Patient Genetic Heterogeneity Identifies New Ways to Target Cholangiocarcinoma
Authors: Younger Nicholas T. , Wilson Mollie L. , Martinez Lyons Anabel , Jarman Edward J. , Meynert Alison M. , Grimes Graeme R. , Gournopanos Konstantinos , Waddell Scott H. , Tennant Peter A. , Wilson David H. , Guest Rachel V. , Wigmore Stephen J. , Acosta Juan Carlos , Kendall Timothy J. , Taylor Martin S. , Sproul Duncan , Mill Pleasantine , Boulter Luke
Published: March 2023

Abstract
Intrahepatic cholangiocarcinoma (ICC) is an aggressive malignancy of the bile ducts within the liver characterized by high levels of genetic heterogeneity. In the context of such genetic variability, determining which oncogenic mutations drive ICC growth has been difficult, and developing modes of patient stratification and targeted therapies remains challenging. Here we model the interactions between rare mutations with more common driver genes and combine in silico analysis of patient data with highly multiplexed in vivo CRISPR-spCas9 screens to perform a functional in vivo study into the role genetic heterogeneity plays in driving ICC. Novel tumor suppressors were uncovered, which, when lost, cooperate with the RAS oncoprotein to drive ICC growth. Focusing on a set of driver mutations that interact with KRAS to initiate aggressive, sarcomatoid-type ICC revealed that tumor growth relies on Wnt and PI3K signaling. Pharmacologic coinhibition of Wnt and PI3K in vivo impeded ICC growth regardless of mutational profile. Therefore, Wnt and PI3K activity should be considered as a signature by which patients can be stratified for treatment independent of tumor genotype, and inhibitors of these pathways should be levied to treat ICC.
Significance:
This work shows that, despite significant genetic heterogeneity, intrahepatic cholangiocarcinoma relies on a limited number of signaling pathways to grow, suggesting common therapeutic vulnerabilities across patients.
Data from Polypharmacologic Reprogramming of Tumor-Associated Macrophages toward an Inflammatory Phenotype
Authors: Nishida-Aoki Nao , Gujral Taranjit S.
Published: March 2023

Abstract
Tumor-associated macrophages (TAM) are an important component of the tumor microenvironment (TME) that can promote tumor progression, metastasis, and resistance to therapies. Although TAMs represent a promising target for therapeutic intervention, the complexity of the TME has made the study of TAMs challenging. Here, we established a physiologically relevant in vitro TAM polarization system that recapitulates TAM protumoral activities. This system was used to characterize dynamic changes in gene expression and protein phosphorylation during TAM polarization and to screen phenotypic kinase inhibitors that impact TAM programming. BMS-794833, a multitargeted compound, was identified as a potent inhibitor of TAM polarization. BMS-794833 decreased protumoral properties of TAMs in vitro and suppressed tumor growth in mouse triple-negative breast cancer models. The effect of BMS-794833 was independent of its primary targets (MET and VEGFR2) but was dependent on its effect on multiple signaling pathways, including focal adhesion kinases, SRC family kinases, STAT3, and p38 MAPKs. Collectively, these findings underline the efficacy of polypharmacologic strategies in reprogramming complex signaling cascades activated during TAM polarization.
Significance:
A physiologically relevant in vitro system of TAM polarization uncovers signaling pathways that regulate polarization and identifies strategies to target macrophage reprogramming to suppress cancer growth.
Data from Combined Inhibition of G9a and EZH2 Suppresses Tumor Growth via Synergistic Induction of IL24-Mediated Apoptosis
Authors: Casciello Francesco , Kelly Gregory M. , Ramarao-Milne Priya , Kamal Nabilah , Stewart Teneale A. , Mukhopadhyay Pamela , Kazakoff Stephen H. , Miranda Mariska , Kim Dorim , Davis Felicity M. , Hayward Nicholas K. , Vertino Paula M. , Waddell Nicola , Gannon Frank , Lee Jason S.
Published: March 2023

Abstract
G9a and EZH2 are two histone methyltransferases commonly upregulated in several cancer types, yet the precise roles that these enzymes play cooperatively in cancer is unclear. We demonstrate here that frequent concurrent upregulation of both G9a and EZH2 occurs in several human tumors. These methyltransferases cooperatively repressed molecular pathways responsible for tumor cell death. In genetically distinct tumor subtypes, concomitant inhibition of G9a and EZH2 potently induced tumor cell death, highlighting the existence of tumor cell survival dependency at the epigenetic level. G9a and EZH2 synergistically repressed expression of genes involved in the induction of endoplasmic reticulum (ER) stress and the production of reactive oxygen species. IL24 was essential for the induction of tumor cell death and was identified as a common target of G9a and EZH2. Loss of function of G9a and EZH2 activated the IL24-ER stress axis and increased apoptosis in cancer cells while not affecting normal cells. These results indicate that G9a and EZH2 promotes the evasion of ER stress–mediated apoptosis by repressing IL24 transcription, therefore suggesting that their inhibition may represent a potential therapeutic strategy for solid cancers.
Significance:
These findings demonstrate a novel role for G9a and EZH2 histone methyltransferases in suppressing apoptosis, which can be targeted with small molecule inhibitors as a potential approach to improve solid cancer treatment.
Data from Gain of Chromosome 1q Perturbs a Competitive Endogenous RNA Network to Promote Melanoma Metastasis
Authors: Xu Xiaonan , Wang Kaizhen , Vera Olga , Verma Akanksha , Jasani Neel , Bok Ilah , Elemento Olivier , Du Dongliang , Yu Xiaoqing , Karreth Florian A.
Published: March 2023

Abstract
Somatic copy-number alterations (CNA) promote cancer, but the underlying driver genes may not be comprehensively identified if only the functions of the encoded proteins are considered. mRNAs can act as competitive endogenous RNAs (ceRNA), which sponge miRNAs to posttranscriptionally regulate gene expression in a protein coding–independent manner. We investigated the contribution of ceRNAs to the oncogenic effects of CNAs. Chromosome 1q gains promoted melanoma progression and metastasis at least in part through overexpression of three mRNAs with ceRNA activity: CEP170, NUCKS1, and ZC3H11A. These ceRNAs enhanced melanoma metastasis by sequestering tumor suppressor miRNAs. Orthogonal genetic assays with miRNA inhibitors and target site blockers, along with rescue experiments, demonstrated that miRNA sequestration is critical for the oncogenic effects of CEP170, NUCKS1, and ZC3H11A mRNAs. Furthermore, chromosome 1q ceRNA-mediated miRNA sequestration alleviated the repression of several prometastatic target genes. This regulatory RNA network was evident in other cancer types, suggesting chromosome 1q ceRNA deregulation as a common driver of cancer progression. Taken together, this work demonstrates that ceRNAs mediate the oncogenicity of somatic CNAs.
Significance:
The function of CEP170, NUCKS1, and ZC3H11A mRNAs as competitive endogenous RNAs that sequester tumor suppressor microRNAs underlies the oncogenic activity of chromosome 1q gains.
Data from Targeting RAS Mutant Colorectal Cancer with Dual Inhibition of MEK and CDK4/6
Authors: Sorokin Alexey V. , Kanikarla Marie Preeti , Bitner Lea , Syed Muddassir , Woods Melanie , Manyam Ganiraju , Kwong Lawrence N. , Johnson Benny , Morris Van K. , Jones Philip , Menter David G. , Lee Michael S. , Kopetz Scott
Published: March 2023

Abstract
KRAS and NRAS mutations occur in 45% of colorectal cancers, with combined MAPK pathway and CDK4/6 inhibition identified as a potential therapeutic strategy. In the current study, this combinatorial treatment approach was evaluated in a co-clinical trial in patient-derived xenografts (PDX), and safety was established in a clinical trial of binimetinib and palbociclib in patients with metastatic colorectal cancer with RAS mutations. Across 18 PDX models undergoing dual inhibition of MEK and CDK4/6, 60% of tumors regressed, meeting the co-clinical trial primary endpoint. Prolonged duration of response occurred predominantly in TP53 wild-type models. Clinical evaluation of binimetinib and palbociclib in a safety lead-in confirmed safety and provided preliminary evidence of activity. Prolonged treatment in PDX models resulted in feedback activation of receptor tyrosine kinases and acquired resistance, which was reversed with a SHP2 inhibitor. These results highlight the clinical potential of this combination in colorectal cancer, along with the utility of PDX-based co-clinical trial platforms for drug development.
Significance:
This co-clinical trial of combined MEK-CDK4/6 inhibition in RAS mutant colorectal cancer demonstrates therapeutic efficacy in patient-derived xenografts and safety in patients, identifies biomarkers of response, and uncovers targetable mechanisms of resistance.
Data from Delineating the RAS Conformational Landscape
Authors: Parker Mitchell I. , Meyer Joshua E. , Golemis Erica A. , Dunbrack, Roland L.
Published: March 2023

Abstract
Mutations in RAS isoforms (KRAS, NRAS, and HRAS) are among the most frequent oncogenic alterations in many cancers, making these proteins high priority therapeutic targets. Effectively targeting RAS isoforms requires an exact understanding of their active, inactive, and druggable conformations. However, there is no structural catalog of RAS conformations to guide therapeutic targeting or examining the structural impact of RAS mutations. Here we present an expanded classification of RAS conformations based on analyses of the catalytic switch 1 (SW1) and switch 2 (SW2) loops. From 721 human KRAS, NRAS, and HRAS structures available in the Protein Data Bank (206 RAS–protein cocomplexes, 190 inhibitor-bound, and 325 unbound, including 204 WT and 517 mutated structures), we created a broad conformational classification based on the spatial positions of Y32 in SW1 and Y71 in SW2. Clustering all well-modeled SW1 and SW2 loops using a density-based machine learning algorithm defined additional conformational subsets, some previously undescribed. Three SW1 conformations and nine SW2 conformations were identified, each associated with different nucleotide states (GTP-bound, nucleotide-free, and GDP-bound) and specific bound proteins or inhibitor sites. The GTP-bound SW1 conformation could be further subdivided on the basis of the hydrogen bond type made between Y32 and the GTP γ-phosphate. Further analysis clarified the catalytic impact of G12D and G12V mutations and the inhibitor chemistries that bind to each druggable RAS conformation. Overall, this study has expanded our understanding of RAS structural biology, which could facilitate future RAS drug discovery.
Significance:
Analysis of >700 RAS structures helps define an expanded landscape of active, inactive, and druggable RAS conformations, the structural impact of common RAS mutations, and previously uncharacterized RAS inhibitor–binding modes.
Data from Pharmacologic Targeting of TFIIH Suppresses KRAS-Mutant Pancreatic Ductal Adenocarcinoma and Synergizes with TRAIL
Authors: Moser Russell , Annis James , Nikolova Olga , Whatcott Cliff , Gurley Kay , Mendez Eduardo , Moran-Jones Kim , Dorrell Craig , Sears Rosalie C. , Kuo Calvin , Han Haiyong , Biankin Andrew , Grandori Carla , Von Hoff Daniel D. , Kemp Christopher J.
Published: March 2023

Abstract
Pancreatic ductal adenocarcinoma (PDAC) typically presents as metastatic disease at diagnosis and remains refractory to treatment. Next-generation sequencing efforts have described the genomic landscape, classified molecular subtypes, and confirmed frequent alterations in major driver genes, with coexistent alterations in KRAS and TP53 correlating with the highest metastatic burden and poorest outcomes. However, translating this information to guide therapy remains a challenge. By integrating genomic analysis with an arrayed RNAi druggable genome screen and drug profiling of a KRAS/TP53 mutant PDAC cell line derived from a patient-derived xenograft (PDCL), we identified numerous targetable vulnerabilities that reveal both known and novel functional aspects of pancreatic cancer biology. A dependence on the general transcription and DNA repair factor TFIIH complex, particularly the XPB subunit and the CAK complex (CDK7/CyclinH/MAT1), was identified and further validated utilizing a panel of genomically subtyped KRAS mutant PDCLs. TFIIH function was inhibited with a covalent inhibitor of CDK7/12/13 (THZ1), a CDK7/CDK9 kinase inhibitor (SNS-032), and a covalent inhibitor of XPB (triptolide), which led to disruption of the protein stability of the RNA polymerase II subunit RPB1. Loss of RPB1 following TFIIH inhibition led to downregulation of key transcriptional effectors of KRAS-mutant signaling and negative regulators of apoptosis, including MCL1, XIAP, and CFLAR, initiating caspase-8 dependent apoptosis. All three drugs exhibited synergy in combination with a multivalent TRAIL, effectively reinforcing mitochondrial-mediated apoptosis. These findings present a novel combination therapy, with direct translational implications for current clinical trials on metastatic pancreatic cancer patients.
Significance:
This study utilizes functional genetic and pharmacological profiling of KRAS-mutant pancreatic adenocarcinoma to identify therapeutic strategies and finds that TFIIH inhibition synergizes with TRAIL to induce apoptosis in KRAS-driven pancreatic cancer.
Data from An Active Learning Framework Improves Tumor Variant Interpretation
Authors: Blee Alexandra M. , Li Bian , Pecen Turner , Meiler Jens , Nagel Zachary D. , Capra John A. , Chazin Walter J.
Published: March 2023

Abstract
For precision medicine to reach its full potential for treatment of cancer and other diseases, protein variant effect prediction tools are needed to characterize variants of unknown significance (VUS) in a patient's genome with respect to their likelihood to influence treatment response and outcomes. However, the performance of most variant prediction tools is limited by the difficulty of acquiring sufficient training and validation data. To overcome these limitations, we applied an iterative active learning approach starting from available biochemical, evolutionary, and functional annotations. With active learning, VUS that are most challenging to classify by an initial machine learning model are functionally evaluated and then reincorporated with the phenotype information in subsequent iterations of algorithm training. The potential of active learning to improve variant interpretation was first demonstrated by applying it to synthetic and deep mutational scanning datasets for four cancer-relevant proteins. The utility of the approach to guide interpretation and functional validation of tumor VUS was then probed on the nucleotide excision repair (NER) protein xeroderma pigmentosum complementation group A (XPA), a potential biomarker for cancer therapy sensitivity. A quantitative high-throughput cell-based NER activity assay was used to validate XPA VUS selected by the active learning strategy. In all cases, active learning yielded a significant improvement in variant effect predictions over traditional learning. These analyses suggest that active learning is well suited to significantly improve interpretation of VUS and cancer patient genomes.
Significance:
A novel machine learning approach predicts the impact of tumor mutations on cellular phenotypes, overcomes limited training data, minimizes costly functional validation, and advances efforts to implement cancer precision medicine.
Data from Developmental Pathways Are Epigenetically Reprogrammed during Lung Cancer Brain Metastasis
Authors: Karlow Jennifer A. , Devarakonda Siddhartha , Xing Xiaoyun , Jang Hyo Sik , Govindan Ramaswamy , Watson Mark , Wang Ting
Published: March 2023

Abstract
Non–small cell lung cancer (NSCLC) is one of the most commonly diagnosed and deadliest cancers worldwide, with roughly half of all patients initially presenting with both primary and metastatic disease. While the major events in the metastatic cascade have been identified, a mechanistic understanding of how NSCLC routinely and successfully colonizes the brain is largely unknown. Recent studies have begun demonstrating the role of epigenetic misregulation during tumorigenesis and metastasis, including widespread changes in DNA methylation and histone modifications. To better understand the role of altered DNA methylation in NSCLC metastasis to the brain, we measured DNA methylation during disease progression for 12 patients, globally profiling the methylation status of normal lung, primary lung tumor, and brain metastasis samples. The variation in methylation was similar during metastatic spread and primary tumorigenesis but less coordinated across genomic features during metastasis. The greatest recurrent changes during metastatic progression were methylation gains in DNA methylation valleys (DMV) harboring the constitutive heterochromatin mark H3K9me3 as well as bivalent marks H3K27me3 and H3K4me1. In a lymph node–derived cancer cell line, EZH2 binding within DMVs was lost, accompanied by an increase in DNA methylation, exemplifying epigenetic switching. The vast majority of the differentially methylated region–associated DMVs harbored developmental genes, suggesting that altered epigenetic regulation of developmentally important genes may confer a selective advantage during metastatic progression. The characterization of epigenetic changes during NSCLC brain metastasis identified recurrent methylation patterns that may be prognostic biomarkers and contributors to disease progression.
Significance:
Altered DNA methylation in lung cancer brain metastases corresponds with loss of EZH2 occupancy at developmental genes, which could promote stem-like phenotypes permissive of dissemination and survival in different microenvironments.