bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from SLC43A3 Is a Biomarker of Sensitivity to the Telomeric DNA Damage Mediator 6-Thio-2′-Deoxyguanosine
Authors: Mender Ilgen , Batten Kimberly , Peyton Michael , Vemula Aishwarya , Cornelius Crystal , Girard Luc , Gao Boning , Minna John D. , Shay Jerry W.
Published: March 2023

Abstract
Cell membrane transporters facilitate the passage of nucleobases and nucleosides for nucleotide synthesis and metabolism, and are important for the delivery of nucleoside analogues used in anticancer drug therapy. Here, we investigated if cell membrane transporters are involved in the cellular uptake of the nucleoside analogue DNA damage mediator 6-thio-2′-deoxyguanosine (6-thio-dG). A large panel of non–small cell lung cancer (NSCLC) cell lines (73 of 77) were sensitive to 6-thio-dG; only four NSCLC lines were resistant to 6-thio-dG. When analyzed by microarray and RNA sequencing, the resistant NSCLC cell lines clustered together, providing a molecular signature for patients that may not respond to 6-thio-dG. Significant downregulation of solute carrier family 43 A3 (SLC43A3), an equilibrative nucleobase transporter, was identified as a candidate in this molecular resistance signature. High levels of SLC43A3 mRNA predicted sensitivity to 6-thio-dG and therefore SLC43A3 could serve as a promising biomarker for 6-thio-dG sensitivity in patients with NSCLC.
Significance:
These findings identify a biomarker of resistance to the telomeric DNA damage mediator 6-thio-2′-deoxyguanosine.
Data from Chromosomal Instability and mTORC1 Activation through PTEN Loss Contribute to Proteotoxic Stress in Ovarian Carcinoma
Authors: Chui M. Herman , Doodnauth Sasha A. , Erdmann Natalie , Tiedemann Rodger E. , Sircoulomb Fabrice , Drapkin Ronny , Shaw Patricia , Rottapel Robert
Published: March 2023

Abstract
High-grade serous ovarian carcinoma commonly arises from fallopian tube secretory epithelium and is characterized by a high level of chromosomal instability. To model the acquisition of aneuploidy during early carcinogenesis, chromosome missegregation was induced in immortalized tubal epithelial cells, which proved acutely detrimental to cellular fitness. The phenotype was characterized by accumulation of misfolded proteins, activation of the unfolded protein response (UPR), decreased protein synthesis, and enhanced vulnerability to proteasome inhibition. However, chromosome missegregation also resulted in heightened transformation potential, assessed by colony formation in soft agar. Ovarian cancer cells retained intrinsic sensitivity to proteasome inhibitors under adherent culture conditions, but acquired resistance as spheroids (recapitulating their native configuration in ascites) by downregulating protein synthesis via mTORC1 suppression. Loss of PTEN drove constitutive mTORC1 activity, enhanced proteotoxic stress, as evidenced by UPR induction, and resensitized tumor spheroids to proteasome inhibition both in vitro and in vivo. In cohorts of primary ovarian carcinomas, mTORC1 and UPR signaling pathways were closely associated. These results implicate attenuation of protein synthesis as a protective mechanism in tumor spheroids, which may explain the overall poor response to bortezomib in clinical trials of patients with advanced ovarian cancer. However, patients with PTEN-deficient tumors may represent a subpopulation potentially amenable to treatment with proteasome inhibitors or other therapeutic agents that disrupt protein homeostasis.
Significance:
Chromosome instability and protein synthesis are important factors that determine the efficacy of proteotoxic stress-inducing agents, such as proteasome inhibitors, in the treatment of ovarian cancer.
Data from Chromothripsis in Human Breast Cancer
Authors: Bolkestein Michiel , Wong John K.L. , Thewes Verena , Körber Verena , Hlevnjak Mario , Elgaafary Shaymaa , Schulze Markus , Kommoss Felix K.F. , Sinn Hans-Peter , Anzeneder Tobias , Hirsch Steffen , Devens Frauke , Schröter Petra , Höfer Thomas , Schneeweiss Andreas , Lichter Peter , Zapatka Marc , Ernst Aurélie
Published: March 2023

Abstract
Chromothripsis is a form of genome instability by which a presumably single catastrophic event generates extensive genomic rearrangements of one or a few chromosomes. Widely assumed to be an early event in tumor development, this phenomenon plays a prominent role in tumor onset. In this study, an analysis of chromothripsis in 252 human breast cancers from two patient cohorts (149 metastatic breast cancers, 63 untreated primary tumors, 29 local relapses, and 11 longitudinal pairs) using whole-genome and whole-exome sequencing reveals that chromothripsis affects a substantial proportion of human breast cancers, with a prevalence over 60% in a cohort of metastatic cases and 25% in a cohort comprising predominantly luminal breast cancers. In the vast majority of cases, multiple chromosomes per tumor were affected, with most chromothriptic events on chromosomes 11 and 17 including, among other significantly altered drivers, CCND1, ERBB2, CDK12, and BRCA1. Importantly, chromothripsis generated recurrent fusions that drove tumor development. Chromothripsis-related rearrangements were linked with univocal mutational signatures, with clusters of point mutations due to kataegis in close proximity to the genomic breakpoints and with the activation of specific signaling pathways. Analyzing the temporal order of events in tumors with and without chromothripsis as well as longitudinal analysis of chromothriptic patterns in tumor pairs offered important insights into the role of chromothriptic chromosomes in tumor evolution.
Significance:
These findings identify chromothripsis as a major driving event in human breast cancer.
Data from HDAC6 Inhibition Synergizes with Anti-PD-L1 Therapy in ARID1A-Inactivated Ovarian Cancer
Authors: Fukumoto Takeshi , Fatkhutdinov Nail , Zundell Joseph A. , Tcyganov Evgenii N. , Nacarelli Timothy , Karakashev Sergey , Wu Shuai , Liu Qin , Gabrilovich Dmitry I. , Zhang Rugang
Published: March 2023

Abstract
ARID1A, encoding a subunit of the SWI/SNF complex, is the most frequently mutated epigenetic regulator in human cancers and is mutated in more than 50% of ovarian clear cell carcinomas (OCCC), a disease that currently has no effective therapy. Inhibition of histone deacetylase 6 (HDAC6) suppresses the growth of ARID1A-mutated tumors and modulates tumor immune microenvironment. Here, we show that inhibition of HDAC6 synergizes with anti-PD-L1 immune checkpoint blockade in ARID1A-inactivated ovarian cancer. ARID1A directly repressed transcription of CD274, the gene encoding PD-L1. Reduced tumor burden and improved survival were observed in ARID1Aflox/flox/PIK3CAH1047R OCCC mice treated with the HDAC6 inhibitor ACY1215 and anti-PD-L1 immune checkpoint blockade as a result of activation and increased presence of IFNγ-positive CD8 T cells. We confirmed that the combined treatment limited tumor progression in a cytotoxic T-cell–dependent manner, as depletion of CD8+ T cells abrogated these antitumor effects. Together, these findings indicate that combined HDAC6 inhibition and immune checkpoint blockade represents a potential treatment strategy for ARID1A-mutated cancers.
Significance:
These findings offer a mechanistic rationale for combining epigenetic modulators and existing immunotherapeutic interventions against a disease that has been so far resistant to checkpoint blockade as a monotherapy.

See related commentary by Prokunina-Olsson, p. 5476
Data from Pharmacokinetic Profiles Determine Optimal Combination Treatment Schedules in Computational Models of Drug Resistance
Authors: Irurzun-Arana Itziar , McDonald Thomas O. , Trocóniz Iñaki F. , Michor Franziska
Published: March 2023

Abstract
Identification of optimal schedules for combination drug administration relies on accurately estimating the correct pharmacokinetics, pharmacodynamics, and drug interaction effects. Misspecification of pharmacokinetics can lead to wrongly predicted timing or order of treatments, leading to schedules recommended on the basis of incorrect assumptions about absorption and elimination of a drug and its effect on tumor growth. Here, we developed a computational modeling platform and software package for combination treatment strategies with flexible pharmacokinetic profiles and multidrug interaction curves that are estimated from data. The software can be used to compare prespecified schedules on the basis of the number of resistant cells where drug interactions and pharmacokinetic curves can be estimated from user-provided data or models. We applied our approach to publicly available in vitro data of treatment with different tyrosine kinase inhibitors of BT-20 triple-negative breast cancer cells and of treatment with erlotinib of PC-9 non–small cell lung cancer cells. Our approach is publicly available in the form of an R package called ACESO (https://github.com/Michorlab/aceso) and can be used to investigate optimum dosing for any combination treatment.
Significance:
These findings introduce a computational modeling platform and software package for combination treatment strategies with flexible pharmacokinetic profiles and multidrug interaction curves that are estimated from data.
Data from The Alternative Splicing Factor, MBNL1, Inhibits Glioblastoma Tumor Initiation and Progression by Reducing Hypoxia-Induced Stemness
Authors: Voss Dillon M. , Sloan Anthony , Spina Raffaella , Ames Heather M. , Bar Eli E.
Published: March 2023

Abstract
Muscleblind-like proteins (MBNL) belong to a family of tissue-specific regulators of RNA metabolism that control premessenger RNA splicing. Inactivation of MBNL causes an adult-to-fetal alternative splicing transition, resulting in the development of myotonic dystrophy. We have previously shown that the aggressive brain cancer, glioblastoma (GBM), maintains stem-like features (glioma stem cell, GSC) through hypoxia-induced responses. Accordingly, we hypothesize here that hypoxia-induced responses in GBM might also include MBNL-based alternative splicing to promote tumor progression. When cultured in hypoxia condition, GSCs rapidly exported muscleblind-like-1 (MBNL1) out of the nucleus, resulting in significant inhibition of MBNL1 activity. Notably, hypoxia-regulated inhibition of MBNL1 also resulted in evidence of adult-to-fetal alternative splicing transitions. Forced expression of a constitutively active isoform of MBNL1 inhibited GSC self-renewal and tumor initiation in orthotopic transplantation models. Induced expression of MBNL1 in established orthotopic tumors dramatically inhibited tumor progression, resulting in significantly prolonged survival. This study reveals that MBNL1 plays an essential role in GBM stemness and tumor progression, where hypoxic responses within the tumor inhibit MBNL1 activity, promoting stem-like phenotypes and tumor growth. Reversing these effects on MBNL1 may therefore, yield potent tumor suppressor activities, uncovering new therapeutic opportunities to counter this disease.
Significance:
This study describes an unexpected mechanism by which RNA-binding protein, MBNL1, activity is inhibited in hypoxia by a simple isoform switch to regulate glioma stem cell self-renewal, tumorigenicity, and progression.
Data from Intratumoral Copper Modulates PD-L1 Expression and Influences Tumor Immune Evasion
Authors: Voli Florida , Valli Emanuele , Lerra Luigi , Kimpton Kathleen , Saletta Federica , Giorgi Federico M. , Mercatelli Daniele , Rouaen Jourdin R.C. , Shen Sylvie , Murray Jayne E. , Ahmed-Cox Aria , Cirillo Giuseppe , Mayoh Chelsea , Beavis Paul A. , Haber Michelle , Trapani Joseph A. , Kavallaris Maria , Vittorio Orazio
Published: March 2023

Abstract
Therapeutic checkpoint antibodies blocking programmed death receptor 1/programmed death ligand 1 (PD-L1) signaling have radically improved clinical outcomes in cancer. However, the regulation of PD-L1 expression on tumor cells is still poorly understood. Here we show that intratumoral copper levels influence PD-L1 expression in cancer cells. Deep analysis of the The Cancer Genome Atlas database and tissue microarrays showed strong correlation between the major copper influx transporter copper transporter 1 (CTR-1) and PD-L1 expression across many cancers but not in corresponding normal tissues. Copper supplementation enhanced PD-L1 expression at mRNA and protein levels in cancer cells and RNA sequencing revealed that copper regulates key signaling pathways mediating PD-L1–driven cancer immune evasion. Conversely, copper chelators inhibited phosphorylation of STAT3 and EGFR and promoted ubiquitin-mediated degradation of PD-L1. Copper-chelating drugs also significantly increased the number of tumor-infiltrating CD8+ T and natural killer cells, slowed tumor growth, and improved mouse survival. Overall, this study reveals an important role for copper in regulating PD-L1 and suggests that anticancer immunotherapy might be enhanced by pharmacologically reducing intratumor copper levels.
Significance:
These findings characterize the role of copper in modulating PD-L1 expression and contributing to cancer immune evasion, highlighting the potential for repurposing copper chelators as enhancers of antitumor immunity.
Data from CRISPR/Cas9-Mediated Point Mutation in Nkx3.1 Prolongs Protein Half-Life and Reverses Effects Nkx3.1 Allelic Loss
Authors: Bowen Cai , Shibata Maho , Zhang Hailan , Bergren Sarah K. , Shen Michael M. , Gelmann Edward P.
Published: March 2023

Abstract
NKX3.1 is the most commonly deleted gene in prostate cancer and is a gatekeeper suppressor. NKX3.1 is haploinsufficient, and pathogenic reduction in protein levels may result from genetic loss, decreased transcription, and increased protein degradation caused by inflammation or PTEN loss. NKX3.1 acts by retarding proliferation, activating antioxidants, and enhancing DNA repair. DYRK1B-mediated phosphorylation at serine 185 of NKX3.1 leads to its polyubiquitination and proteasomal degradation. Because NKX3.1 protein levels are reduced, but never entirely lost, in prostate adenocarcinoma, enhancement of NKX3.1 protein levels represents a potential therapeutic strategy. As a proof of principle, we used CRISPR/Cas9-mediated editing to engineer in vivo a point mutation in murine Nkx3.1 to code for a serine to alanine missense at amino acid 186, the target for Dyrk1b phosphorylation. Nkx3.1S186A/−, Nkx3.1+/−, and Nkx3.1+/+ mice were analyzed over one year to determine the levels of Nkx3.1 expression and effects of the mutant protein on the prostate. Allelic loss of Nkx3.1 caused reduced levels of Nkx3.1 protein, increased proliferation, and prostate hyperplasia and dysplasia, whereas Nkx3.1S186A/− mouse prostates had increased levels of Nkx3.1 protein, reduced prostate size, normal histology, reduced proliferation, and increased DNA end labeling. At 2 months of age, when all mice had normal prostate histology, Nkx3.1+/− mice demonstrated indices of metabolic activation, DNA damage response, and stress response. These data suggest that modulation of Nkx3.1 levels alone can exert long-term control over premalignant changes and susceptibility to DNA damage in the prostate.
Significance:
These findings show that prolonging the half-life of Nkx3.1 reduces proliferation, enhances DNA end-labeling, and protects from DNA damage, ultimately blocking the proneoplastic effects of Nkx3.1 allelic loss.
Data from Slow-Cycling Cancer Stem Cells Regulate Progression and Chemoresistance in Colon Cancer
Authors: Shiokawa Daisuke , Sakai Hiroaki , Ohata Hirokazu , Miyazaki Toshiaki , Kanda Yusuke , Sekine Shigeki , Narushima Daichi , Hosokawa Masahito , Kato Mamoru , Suzuki Yutaka , Takeyama Haruko , Kambara Hideki , Nakagama Hitoshi , Okamoto Koji
Published: March 2023

Abstract
Cancer chemoresistance is often attributed to the presence of cancer stem cell (CSC)-like cells, but whether they are homogeneously chemoresistant remains unclear. We previously showed that in colon tumors, a subpopulation of LGR5+ CSC-like cells driven by TCF1 (TCF7), a Wnt-responsive transcription factor, were responsible for tumorigenicity. Here we demonstrate that the tumorigenic subpopulation of mouse LGR5+ cells exists in a slow-cycling state and identify a unique 22-gene signature that characterizes these slow-cycling CSC. Seven of the signature genes are specifically expressed in slow-cycling LGR5+ cells from xenografted human colon tumors and are upregulated in colon cancer clinical specimens. Among these seven, four genes (APCDD1, NOTUM, PROX1, and SP5) are known to be direct Wnt target genes, and PROX1 was expressed in the invasive fronts of colon tumors. PROX1 was activated by TCF1 to induce CDKN1C and maintain a slow-cycling state in colon cancer organoids. Strikingly, PROX1 was required for recurrent growth after chemotherapeutic treatment, suggesting that inhibition of slow-cycling CSC by targeting the TCF1–PROX1–CDKN1C pathway is an effective strategy to combat refractory colon cancer in combination with conventional chemotherapy.
Significance:
These findings illustrate the importance of a slow-cycling CSC subpopulation in colon cancer development and chemoresistance, with potential implications for the identified slow-cycling CSC signatures and the TCF1–PROX1–CDKN1C pathway as therapeutic targets.
Data from Hes1 Is Essential in Proliferating Ductal Cell–Mediated Development of Intrahepatic Cholangiocarcinoma
Authors: Matsumori Tomoaki , Kodama Yuzo , Takai Atsushi , Shiokawa Masahiro , Nishikawa Yoshihiro , Matsumoto Tomonori , Takeda Haruhiko , Marui Saiko , Okada Hirokazu , Hirano Tomonori , Kuwada Takeshi , Sogabe Yuko , Kakiuchi Nobuyuki , Tomono Teruko , Mima Atsushi , Morita Toshihiro , Ueda Tatsuki , Tsuda Motoyuki , Yamauchi Yuki , Kuriyama Katsutoshi , Sakuma Yojiro , Ota Yuji , Maruno Takahisa , Uza Norimitsu , Marusawa Hiroyuki , Kageyama Ryoichiro , Chiba Tsutomu , Seno Hiroshi
Published: March 2023

Abstract
Intrahepatic cholangiocarcinoma (ICC) is frequently driven by aberrant KRAS activation and develops in the liver with chronic inflammation. Although the Notch signaling pathway is critically involved in ICC development, detailed mechanisms of Notch-driven ICC development are still unknown. Here, we use mice whose Notch signaling is genetically engineered to show that the Notch signaling pathway, specifically the Notch/Hes1 axis, plays an essential role in expanding ductular cells in the liver with chronic inflammation or oncogenic Kras activation. Activation of Notch1 enhanced the development of proliferating ductal cells (PDC) in injured livers, while depletion of Hes1 led to suppression. In correlation with PDC expansion, ICC development was also regulated by the Notch/Hes1 axis and suppressed by Hes1 depletion. Lineage-tracing experiments using EpcamcreERT2 mice further confirmed that Hes1 plays a critical role in the induction of PDC and that ICC could originate from PDC. Analysis of human ICC specimens showed PDC in nonneoplastic background tissues, confirming HES1 expression in both PDC and ICC tumor cells. Our findings provide novel direct experimental evidence that Hes1 plays an essential role in the development of ICC via PDC.
Significance:
This study contributes to the identification of the cells of origin that initiate ICC and suggests that HES1 may represent a therapeutic target in ICC.
Data from Obesity/Type 2 Diabetes-Associated Liver Tumors Are Sensitive to Cyclin D1 Deficiency
Authors: Luo Chi , Liang Jiaxin , Sharabi Kfir , Hatting Maximilian , Perry Elizabeth A. , Tavares Clint D.J. , Goyal Lipika , Srivastava Amitabh , Bilodeau Marc , Zhu Andrew X. , Sicinski Piotr , Puigserver Pere
Published: March 2023

Abstract
Type 2 diabetes, which is mainly linked to obesity, is associated with increased incidence of liver cancer. We have previously found that in various models of obesity/diabetes, hyperinsulinemia maintains heightened hepatic expression of cyclin D1, suggesting a plausible mechanism linking diabetes and liver cancer progression. Here we show that cyclin D1 is greatly elevated in human livers with diabetes and is among the most significantly upregulated genes in obese/diabetic liver tumors. Liver-specific cyclin D1 deficiency protected obese/diabetic mice against hepatic tumorigenesis, whereas lean/nondiabetic mice developed tumors irrespective of cyclin D1 status. Cyclin D1 dependency positively correlated with liver cancer sensitivity to palbociclib, an FDA-approved CDK4 inhibitor, which was effective in treating orthotopic liver tumors under obese/diabetic conditions. The antidiabetic drug metformin suppressed insulin-induced hepatic cyclin D1 expression and protected against obese/diabetic hepatocarcinogenesis. These results indicate that the cyclin D1–CDK4 complex represents a potential selective therapeutic vulnerability for liver tumors in obese/diabetic patients.
Significance:
Obesity/diabetes-associated liver tumors are specifically vulnerable to cyclin D1 deficiency and CDK4 inhibition, suggesting that the obese/diabetic environment confers cancer-selective dependencies that can be therapeutically exploited.
Data from The D2 and D3 Sublineages of Human Papilloma Virus 16–Positive Cervical Cancer in Guatemala Differ in Integration Rate and Age of Diagnosis
Authors: Lou Hong , Boland Joseph F. , Torres-Gonzalez Edmundo , Albanez Anaseidy , Zhou Weiyin , Steinberg Mia K. , Diaw Lena , Mitchell Jason , Roberson David , Cullen Michael , Garland Lisa , Bass Sara , Burk Robert D. , Yeager Meredith , Wentzensen Nicolas , Schiffman Mark , Freites Enrique Alvirez , Gharzouzi Eduardo , Mirabello Lisa , Dean Michael
Published: March 2023

Abstract
Human papillomavirus (HPV) 16 displays substantial sequence variation; four HPV16 lineages (A, B, C, and D) have been described as well as multiple sublineages. To identify molecular events associated with HPV16 carcinogenesis, we evaluated viral variation, the integration of HPV16, and somatic mutation in 96 cervical cancer samples from Guatemala. A total of 65% (62/96) of the samples had integrated HPV16 sequences and integration was associated with an earlier age of diagnosis and premenopausal disease. HPV16 integration sites were broadly distributed in the genome, but in one tumor, HPV16 integrated into the promoter of the IFN regulatory factor 4 (IRF4) gene, which plays an important role in the regulation of the IFN response to viral infection. The HPV16 D2 and D3 sublineages were found in 23% and 30% of the tumors, respectively, and were significantly associated with adenocarcinoma. D2-positive tumors had a higher rate of integration, earlier age of diagnosis, and a lower rate of somatic mutation, whereas D3-positive tumors were less likely to integrate, had later age of diagnosis, and exhibited a higher rate of somatic mutation. In conclusion, Guatemalan cervical tumors have a high frequency of very high-risk HPV16 D2 and D3 sublineages harboring distinct histology, which may help guide future therapeutic strategies to target the tumor and reduce recurrence.
Significance:
This study details the biological and molecular properties of the most pathogenic forms of HPV16, the cause of the majority of cervical cancers.
Data from EZH2-Mediated Downregulation of the Tumor Suppressor DAB2IP Maintains Ovarian Cancer Stem Cells
Authors: Zong Xingyue , Wang Weini , Ozes Ali , Fang Fang , Sandusky George E. , Nephew Kenneth P.
Published: March 2023

Abstract
The majority of women diagnosed with epithelial ovarian cancer eventually develop recurrence, which rapidly evolves into chemoresistant disease. Persistence of ovarian cancer stem cells (OCSC) at the end of therapy may be responsible for emergence of resistant tumors. In this study, we demonstrate that in OCSC, the tumor suppressor disabled homolog 2–interacting protein (DAB2IP) is silenced by EZH2-mediated H3K27 trimethylation of the DAB2IP promoter. CRISPR/Cas9-mediated deletion of DAB2IP in epithelial ovarian cancer cell lines upregulated expression of stemness-related genes and induced conversion of non-CSC to CSC, while enforced expression of DAB2IP suppressed CSC properties. Transcriptomic analysis showed that overexpression of DAB2IP in ovarian cancer significantly altered stemness-associated genes and bioinformatic analysis revealed WNT signaling as a dominant pathway mediating the CSC inhibitory effect of DAB2IP. Specifically, DAB2IP inhibited WNT signaling via downregulation of WNT5B, an important stemness inducer. Reverse phase protein array further demonstrated activation of noncanonical WNT signaling via C-JUN as a downstream target of WNT5B, which was blocked by inhibiting RAC1, a prominent regulator of C-JUN activation. Coadministration of EZH2 inhibitor GSK126 and RAC1 inhibitor NSC23766 suppressed OCSC survival in vitro and inhibited tumor growth and increased platinum sensitivity in vivo. Overall, these data establish that DAB2IP suppresses the cancer stem cell phenotype via inhibition of WNT5B-induced activation of C-JUN and can be epigenetically silenced by EZH2 in OCSC. Targeting the EZH2/DAB2IP/C-JUN axis therefore presents a promising strategy to prevent ovarian cancer recurrence and has potential for clinical translation.
Significance:
These findings show that combining an epigenetic therapy with a noncanonical WNT signaling pathway inhibitor has the potential to eradicate ovarian cancer stem cells and to prevent ovarian cancer recurrence.
Data from Widespread Repression of Gene Expression in Cancer by a Wnt/β-Catenin/MAPK Pathway
Authors: Harmston Nathan , Lim Jun Yi Stanley , Arqués Oriol , Palmer Héctor G. , Petretto Enrico , Virshup David M. , Madan Babita
Published: March 2023

Abstract
Aberrant Wnt signaling drives a number of cancers through regulation of diverse downstream pathways. Wnt/β-catenin signaling achieves this in part by increasing the expression of proto-oncogenes such as MYC and cyclins. However, global assessment of the Wnt-regulated transcriptome in vivo in genetically distinct cancers demonstrates that Wnt signaling suppresses the expression of as many genes as it activates. In this study, we examined the set of genes that are upregulated upon inhibition of Wnt signaling in Wnt-addicted pancreatic and colorectal cancer models. Decreasing Wnt signaling led to a marked increase in gene expression by activating ERK and JNK; these changes in gene expression could be mitigated in part by concurrent inhibition of MEK. These findings demonstrate that increased Wnt signaling in cancer represses MAPK activity, preventing RAS-mediated senescence while allowing cancer cells to proliferate. These results shift the paradigm from Wnt/β-catenin primarily as an activator of transcription to a more nuanced view where Wnt/β-catenin signaling drives both widespread gene repression and activation.
Significance:
These findings show that Wnt/β-catenin signaling causes widespread gene repression via inhibition of MAPK signaling, thus fine tuning the RAS-MAPK pathway to optimize proliferation in cancer.
Data from Cyclophosphamide and Vinorelbine Activate Stem-Like CD8+ T Cells and Improve Anti-PD-1 Efficacy in Triple-Negative Breast Cancer
Authors: Falvo Paolo , Orecchioni Stefania , Hillje Roman , Raveane Alessandro , Mancuso Patrizia , Camisaschi Chiara , Luzi Lucilla , Pelicci PierGiuseppe , Bertolini Francesco
Published: March 2023

Abstract
Checkpoint inhibitors (CI) instigate anticancer immunity in many neoplastic diseases, albeit only in a fraction of patients. The clinical success of cyclophosphamide (C)-based haploidentical stem-cell transplants indicates that this drug may re-orchestrate the immune system. Using models of triple-negative breast cancer (TNBC) with different intratumoral immune contexture, we demonstrate that a combinatorial therapy of intermittent C, CI, and vinorelbine activates antigen-presenting cells (APC), and abrogates local and metastatic tumor growth by a T-cell–related effect. Single-cell transcriptome analysis of >50,000 intratumoral immune cells after therapy treatment showed a gene signature suggestive of a change resulting from exposure to a mitogen, ligand, or antigen for which it is specific, as well as APC-to-T-cell adhesion. This transcriptional program also increased intratumoral Tcf1+ stem-like CD8+ T cells and altered the balance between terminally and progenitor-exhausted T cells favoring the latter. Overall, our data support the clinical investigation of this therapy in TNBC.
Significance:
A combinatorial therapy in mouse models of breast cancer increases checkpoint inhibition by activating antigen-presenting cells, enhancing intratumoral Tcf1+ stem-like CD8+ T cells, and increasing progenitor exhausted CD8+ T cells.
Data from Unconventional Secretion of PKCδ Exerts Tumorigenic Function via Stimulation of ERK1/2 Signaling in Liver Cancer
Authors: Yamada Kohji , Oikawa Tsunekazu , Kizawa Ryusuke , Motohashi Saya , Yoshida Saishu , Kumamoto Tomotaka , Saeki Chisato , Nakagawa Chika , Shimoyama Yuya , Aoki Katsuhiko , Tachibana Toshiaki , Saruta Masayuki , Ono Masaya , Yoshida Kiyotsugu
Published: March 2023

Abstract
Expression of human protein kinase C delta (PKCδ) protein has been linked to many types of cancers. PKCδ is known to be a multifunctional PKC family member and has been rigorously studied as an intracellular signaling molecule. Here we show that PKCδ is a secretory protein that regulates cell growth of liver cancer. Full-length PKCδ was secreted to the extracellular space in living liver cancer cells under normal cell culture conditions and in xenograft mouse models. Patients with liver cancer showed higher levels of serum PKCδ than patients with chronic hepatitis or liver cirrhosis or healthy individuals. In liver cancer cells, PKCδ secretion was executed in an endoplasmic reticulum (ER)-Golgi–independent manner, and the inactivation status of cytosolic PKCδ was required for its secretion. Furthermore, colocalization studies showed that extracellular PKCδ was anchored on the cell surface of liver cancer cells via association with glypican 3, a liver cancer–related heparan sulfate proteoglycan. Addition of exogenous PKCδ activated IGF-1 receptor (IGF1R) activation and subsequently enhanced activation of ERK1/2, which led to accelerated cell growth in liver cancer cells. Conversely, treatment with anti-PKCδ antibody attenuated activation of both IGF1R and ERK1/2 and reduced cell proliferation and spheroid formation of liver cancer cells and tumor growth in xenograft mouse models. This study demonstrates the presence of PKCδ at the extracellular space and the function of PKCδ as a growth factor and provides a rationale for the extracellular PKCδ-targeting therapy of liver cancer.
Significance:
PKCδ secretion from liver cancer cells behaves as a humoral growth factor that contributes to cell growth via activation of proliferative signaling molecules, which may be potential diagnostic or therapeutic targets.
Data from Glia-Selective Deletion of Complement C1q Prevents Radiation-Induced Cognitive Deficits and Neuroinflammation
Authors: Markarian Mineh , Krattli Robert P. , Baddour Jabra D. , Alikhani Leila , Giedzinski Erich , Usmani Manal T. , Agrawal Anshu , Baulch Janet E. , Tenner Andrea J. , Acharya Munjal M.
Published: March 2023

Abstract
The adverse neurocognitive sequelae following clinical radiotherapy (RT) for central nervous system (CNS) malignancies are often long-lasting without any clinical recourse. Despite recent progress, the cellular mechanisms mediating RT-induced cognitive deficits (RICD) are poorly understood. The complement system is an immediate sensor of a disturbed inflammatory environment and a potent mediator of gliosis with a range of nonimmune functions in the CNS, including synaptic pruning, which is detrimental if dysregulated. We hypothesize that complement-mediated changes in glial cell function significantly contribute to RICD. The underlying alterations in CNS complement cascade proteins (C1q, C3), TLR4, and colabeling with glia (IBA1, GFAP) were examined using gene expression, immunofluorescence, and in silico modeling approaches in the adult mouse brain following 9 Gy cranial RT. Three-dimensional volumetric quantification showed elevated molecular signatures of gliosis at short- and long-term post-RT times. We found significant elevations in complement C1q, C3, and TLR4 post-RT accompanied by increased colabeling of astrocytes and microglia. To address the mechanism of RT-induced complement cascade activation, neuroinflammation, and cognitive dysfunction, we used a genetic approach—conditional, microglia-selective C1q (Flox) knockdown mice—to determine whether a glia-specific, upstream complement cascade contributes to RICD. C1q-Flox mice exposed to cranial RT showed no cognitive deficits compared with irradiated WT mice. Further, irradiated C1q-Flox mice were protected from RT-induced microglial activation and synaptic loss, elevation of anaphylatoxin C5a receptor, astrocytic-C3, and microglial-TLR4 expression in the brain. Our findings demonstrate for the first time a microglia-specific mechanism of RICD involving an upstream complement cascade component, C1q.
Significance:
Clinically-relevant radiotherapy induces aberrant complement activation, leading to brain injury. Microglia-selective genetic deletion of CNS complement C1q ameliorates radiation-induced cognitive impairments, synaptic loss, and neuroinflammation, highlighting the potential for C1q as a novel therapeutic target.

See related commentary by Korimerla and Wahl, p. 1635
Data from The miR–181a–SFRP4 Axis Regulates Wnt Activation to Drive Stemness and Platinum Resistance in Ovarian Cancer
Authors: Belur Nagaraj Anil , Knarr Matthew , Sekhar Sreeja , Connor R. Shae , Joseph Peronne , Kovalenko Olga , Fleming Alexis , Surti Arshia , Nurmemmedov Elmar , Beltrame Luca , Marchini Sergio , Kahn Michael , DiFeo Analisa
Published: March 2023

Abstract
Wnt signaling is a major driver of stemness and chemoresistance in ovarian cancer, yet the genetic drivers that stimulate its expression remain largely unknown. Unlike other cancers, mutations in the Wnt pathway are not reported in high-grade serous ovarian cancer (HGSOC). Hence, a key challenge that must be addressed to develop effective targeted therapies is to identify nonmutational drivers of Wnt activation. Using an miRNA sensor-based approach, we have identified miR-181a as a novel driver of Wnt/β-catenin signaling. miR-181ahigh primary HGSOC cells exhibited increased Wnt/β-catenin signaling, which was associated with increased stem-cell frequency and platinum resistance. Consistent with these findings, inhibition of β-catenin decreased stem-like properties in miR-181ahigh cell populations and downregulated miR-181a. The Wnt inhibitor SFRP4 was identified as a novel target of miR-181a. Overall, our results demonstrate that miR-181a is a nonmutational activator of Wnt signaling that drives stemness and chemoresistance in HGSOC, suggesting that the miR–181a–SFRP4 axis can be evaluated as a novel biomarker for β-catenin–targeted therapy in this disease.
Significance:
These results demonstrate that miR-181a is an activator of Wnt signaling that drives stemness and chemoresistance in HGSOC and may be targeted therapeutically in recurrent disease.
Data from Lymphatic PD-L1 Expression Restricts Tumor-Specific CD8+ T-cell Responses
Authors: Cousin Nikola , Cap Stefan , Dihr Manuel , Tacconi Carlotta , Detmar Michael , Dieterich Lothar C.
Published: March 2023

Abstract
Lymph node (LN)–resident lymphatic endothelial cells (LEC) mediate peripheral tolerance by self-antigen presentation on MHC-I and constitutive expression of T-cell inhibitory molecules, including PD-L1 (CD274). Tumor-associated LECs also upregulate PD-L1, but the specific role of lymphatic PD-L1 in tumor immunity is not well understood. In this study, we generated a mouse model lacking lymphatic PD-L1 expression and challenged these mice with two orthotopic tumor models, B16F10 melanoma and MC38 colorectal carcinoma. Lymphatic PD-L1 deficiency resulted in consistent expansion of tumor-specific CD8+ T cells in tumor-draining LNs in both tumor models, reduced primary tumor growth in the MC38 model, and increased efficacy of adoptive T-cell therapy in the B16F10 model. Strikingly, lymphatic PD-L1 acted primarily by inducing apoptosis in tumor-specific CD8+ central memory T cells. Overall, these findings demonstrate that LECs restrain tumor-specific immunity via PD-L1, which may explain why some patients with cancer without PD-L1 expression in the tumor microenvironment still respond to PD-L1/PD-1–targeted immunotherapy.
Significance:
A new lymphatic-specific PD-L1 knockout mouse model reveals that lymphatic endothelial PD-L1 expression reduces tumor immunity, inducing apoptosis in tumor-specific CD8+ central memory cells in tumor-draining lymph nodes.
Data from PLEKHA4 Promotes Wnt/β-Catenin Signaling–Mediated G1–S Transition and Proliferation in Melanoma
Authors: Shami Shah Adnan , Cao Xiaofu , White Andrew C. , Baskin Jeremy M.
Published: March 2023

Abstract
Despite recent promising advances in targeted therapies and immunotherapies, patients with melanoma incur substantial mortality. In particular, inhibitors targeting BRAF-mutant melanoma can lead to resistance, and no targeted therapies exist for NRAS-mutant melanoma, motivating the search for additional therapeutic targets and vulnerable pathways. Here we identify a regulator of Wnt/β-catenin signaling, PLEKHA4, as a factor required for melanoma proliferation and survival. PLEKHA4 knockdown in vitro decreased Dishevelled levels, attenuated Wnt/β-catenin signaling, and blocked progression through the G1–S cell-cycle transition. In mouse xenograft and allograft models, inducible PLEKHA4 knockdown attenuated tumor growth in BRAF- and NRAS-mutant melanomas and exhibited an additive effect with the clinically used inhibitor encorafenib in a BRAF-mutant model. As an E3 ubiquitin ligase regulator with both lipid- and protein-binding partners, PLEKHA4 presents several opportunities for targeting with small molecules. Our work identifies PLEKHA4 as a promising drug target for melanoma and clarifies a controversial role for Wnt/β-catenin signaling in the control of melanoma proliferation.
Significance:
This study establishes that melanoma cell proliferation requires the protein PLEKHA4 to promote pathologic Wnt signaling for proliferation, highlighting PLEKHA4 inhibition as a new avenue for the development of targeted therapies.
Data from Targeting Pyruvate Kinase M2 Phosphorylation Reverses Aggressive Cancer Phenotypes
Authors: Apostolidi Maria , Vathiotis Ioannis A. , Muthusamy Viswanathan , Gaule Patricia , Gassaway Brandon M. , Rimm David L. , Rinehart Jesse
Published: March 2023

Abstract
Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype with low survival rate and a lack of biomarkers and targeted treatments. Here, we target pyruvate kinase M2 (PKM2), a key metabolic component of oncogenesis. In patients with TNBC, PKM2pS37 was identified as a prominent phosphoprotein corresponding to the aggressive breast cancer phenotype that showed a characteristic nuclear staining pattern and prognostic value. Phosphorylation of PKM2 at S37 was connected with a cyclin-dependent kinase (CDK) pathway in TNBC cells. In parallel, pyruvate kinase activator TEPP-46 bound PKM2pS37 and reduced its nuclear localization. In a TNBC mouse xenograft model, treatment with either TEPP-46 or the potent CDK inhibitor dinaciclib reduced tumor growth and diminished PKM2pS37. Combinations of dinaciclib with TEPP-46 reduced cell invasion, impaired redox balance, and triggered cancer cell death. Collectively, these data support an approach to identify PKM2pS37-positive TNBC and target the PKM2 regulatory axis as a potential treatment.
Significance:
PKM2 phosphorylation marks aggressive breast cancer cell phenotypes and targeting PKM2pS37 could be an effective therapeutic approach for treating triple-negative breast cancer.
Data from CKAP2L Promotes Non–Small Cell Lung Cancer Progression through Regulation of Transcription Elongation
Authors: Monteverde Tiziana , Sahoo Sudhakar , La Montagna Manuela , Magee Peter , Shi Lei , Lee Dave , Sellers Robert , Baker Alexander R. , Leong Hui Sun , Fassan Matteo , Garofalo Michela
Published: March 2023

Abstract
Chromosomal instability (CIN) is a driver of clonal diversification and intratumor heterogeneity, providing genetic diversity that contributes to tumor progression. It is estimated that approximately 80% of solid cancers, including non–small cell lung cancer (NSCLC), exhibit features of CIN, which affects tumor growth and response to therapy. However, the molecular mechanisms connecting CIN to tumor progression are still poorly understood. Through an RNAi screen performed on genes involved in CIN and overexpressed in human lung adenocarcinoma samples, we identified the cytoskeleton-associated protein 2-like (CKAP2L) as a potential oncogene that promotes lung cancer proliferation and growth in vitro and in vivo. Mechanistically, CKAP2L directly interacted with RNA Pol II and regulated transcription elongation of key genes involved in spindle assembly checkpoint, chromosome segregation, cell cycle, and E2F signaling. Furthermore, depletion of CKAP2L increased the sensitivity of NSCLC cells to alvocidib, a pan-CDK inhibitor, leading to a significant reduction of cell proliferation and an increase in cell death. Altogether, these findings shed light on the molecular mechanisms through which CKAP2L, a protein involved in CIN, promotes cancer progression and suggest that its inhibition represents a novel therapeutic strategy in NSCLC.
Significance:
These findings demonstrate the oncogenic function of CKAP2L through regulation of transcription elongation and suggest that targeting CKAP2L could enhance therapeutic response in patients with NSCLC.
Data from Survivin Expression Is Differentially Regulated by a Selective Cross-talk between RBM38 and miRNAs let-7b or miR-203a
Authors: Lucchesi Christopher A. , Zhang Jin , Ma Buyong , Nussinov Ruth , Chen Xinbin
Published: March 2023

Abstract
RNA-binding motif 38 (RBM38) is a member of a protein family with a highly conserved RNA-binding motif and has been shown to regulate mRNA processing, stability, and translation. Survivin is an essential modulator of apoptotic and nonapoptotic cell death as well as a stress responder. Survivin mRNA is the fourth most frequently overexpressed transcript in the human cancer transcriptome, and its aberrant expression is associated with chemo-/radioresistance and poor prognosis. In this study, we examined whether survivin expression is regulated by RBM38. RBM38 bound to survivin 3′-untranslated region and suppressed miRNA let-7b from binding to and degrading survivin mRNA, leading to increased survivin expression. RBM38 interacted with argonaute-2 (AGO2) and facilitated miR-203a–mediated degradation of survivin mRNA, leading to decreased survivin expression. Due to the abundance of let-7b over miR-203a, RBM38 ultimately increased survivin expression in HCT116 and MCF7 cells. In addition, Ser-195 in RBM38 interacted with Glu-73/-76 in AGO2, and Pep8, an eight-amino acid peptide spanning the region of Ser-195 in RBM38, blocked the RBM38–AGO2 interaction and inhibited miR-203a–mediated mRNA degradation, leading to enhanced survivin expression. Furthermore, Pep8 cooperated with YM155, an inhibitor of survivin, to suppress tumor spheroid growth and viability. Pep8 sensitized tumor cells to YM155-induced DNA damage in an RBM38-dependent manner. Together, our data indicate that RBM38 is a dual regulator of survivin and that Pep8/YM155 may be therapeutically explored for tumor suppression.
Significance:
These findings show that RBM38 exerts opposing effects on survivin expression via two miRNAs, and disruption of the RBM38–AGO2 complex by an eight-amino acid peptide sensitizes tumor spheroids to survivin inhibitor YM155.
Data from Multiomics Characterization of Low-Grade Serous Ovarian Carcinoma Identifies Potential Biomarkers of MEK Inhibitor Sensitivity and Therapeutic Vulnerability
Authors: Shrestha Raunak , Llaurado Fernandez Marta , Dawson Amy , Hoenisch Joshua , Volik Stanislav , Lin Yen-Yi , Anderson Shawn , Kim Hannah , Haegert Anne M. , Colborne Shane , Wong Nelson K.Y. , McConeghy Brian , Bell Robert H. , Brahmbhatt Sonal , Lee Cheng-Han , DiMattia Gabriel E. , Le Bihan Stephane , Morin Gregg B. , Collins Colin C. , Carey Mark S.
Published: March 2023

Abstract
Low-grade serous ovarian carcinoma (LGSOC) is a rare tumor subtype with high case fatality rates in patients with metastatic disease. There is a pressing need to develop effective treatments using newly available preclinical models for therapeutic discovery and drug evaluation. Here, we use multiomics integration of whole-exome sequencing, RNA sequencing, and mass spectrometry–based proteomics on 14 LGSOC cell lines to elucidate novel biomarkers and therapeutic vulnerabilities. Comparison of LGSOC cell line data with LGSOC tumor data enabled predictive biomarker identification of MEK inhibitor (MEKi) efficacy, with KRAS mutations found exclusively in MEKi-sensitive cell lines and NRAS mutations found mostly in MEKi-resistant cell lines. Distinct patterns of Catalogue of Somatic Mutations in Cancer mutational signatures were identified in MEKi-sensitive and MEKi-resistant cell lines. Deletions of CDKN2A/B and MTAP genes were more frequent in cell lines than tumor samples and possibly represent key driver events in the absence of KRAS/NRAS/BRAF mutations. These LGSOC cell lines were representative models of the molecular aberrations found in LGSOC tumors. For prediction of in vitro MEKi efficacy, proteomic data provided better discrimination than gene expression data. Condensin, minichromosome maintenance, and replication factor C protein complexes were identified as potential treatment targets in MEKi-resistant cell lines. This study suggests that CDKN2A/B or MTAP deficiency may be exploited using synthetically lethal treatment strategies, highlighting the importance of using proteomic data as a tool for molecular drug prediction. Multiomics approaches are crucial to improving our understanding of the molecular underpinnings of LGSOC and applying this information to develop new therapies.
Significance:
These findings highlight the utility of global multiomics to characterize LGSOC cell lines as research models, to determine biomarkers of MEKi resistance, and to identify potential novel therapeutic targets.
Data from Pan-Cancer Analysis of Ligand–Receptor Cross-talk in the Tumor Microenvironment
Authors: Ghoshdastider Umesh , Rohatgi Neha , Mojtabavi Naeini Marjan , Baruah Probhonjon , Revkov Egor , Guo Yu Amanda , Rizzetto Simone , Wong Angeline M.L. , Solai Sundar , Nguyen Tin T. , Yeong Joe Poh Sheng , Iqbal Jabed , Tan Puay Hoon , Chowbay Balram , Dasgupta Ramanuj , Skanderup Anders J.
Published: March 2023

Abstract
Signaling between cancer and nonmalignant (stromal) cells in the tumor microenvironment (TME) is a key to tumor progression. Here, we deconvoluted bulk tumor transcriptomes to infer cross-talk between ligands and receptors on cancer and stromal cells in the TME of 20 solid tumor types. This approach recovered known transcriptional hallmarks of cancer and stromal cells and was concordant with single-cell, in situ hybridization and IHC data. Inferred autocrine cancer cell interactions varied between tissues but often converged on Ephrin, BMP, and FGFR-signaling pathways. Analysis of immune checkpoints nominated interactions with high levels of cancer-to-immune cross-talk across distinct tumor types. Strikingly, PD-L1 was found to be highly expressed in stromal rather than cancer cells. Overall, our study presents a new resource for hypothesis generation and exploration of cross-talk in the TME.
Significance:
This study provides deconvoluted bulk tumor transcriptomes across multiple cancer types to infer cross-talk in the tumor microenvironment.