bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Accumulation of Molecular Aberrations Distinctive to Hepatocellular Carcinoma Progression
Authors: Midorikawa Yutaka , Yamamoto Shogo , Tatsuno Kenji , Renard-Guillet Claire , Tsuji Shingo , Hayashi Akimasa , Ueda Hiroki , Fukuda Shiro , Fujita Takanori , Katoh Hiroto , Ishikawa Shumpei , Covington Kyle R. , Creighton Chad J. , Sugitani Masahiko , Wheeler David A. , Shibata Tatsuhiro , Nagae Genta , Takayama Tadatoshi , Aburatani Hiroyuki
Published: March 2023

Abstract
Cancer develops through the accumulation of genetic and epigenetic aberrations. To identify sequential molecular alterations that occur during the development of hepatocellular carcinoma (HCC), we compared 52 early and 108 overt HCC samples by genome sequencing. Gene mutations in the p53/RB1 pathway, WNT pathway, MLL protein family, SWI/SNF complexes, and AKT/PI3K pathway were common in HCC. In the early phase of all entities, TERT was the most frequently upregulated gene owing to diverse mechanisms. Despite frequent somatic mutations in driver genes, including CTNNB1 and TP53, early HCC was a separate molecular entity from overt HCC, as each had a distinct expression profile. Notably, WNT target genes were not activated in early HCC regardless of CTNNB1 mutation status because β-catenin did not translocate into the nucleus due to the E-cadherin/β-catenin complex at the membrane. Conversely, WNT targets were definitively upregulated in overt HCC, with CTNNB1 mutation associated with downregulation of CDH1 and hypomethylation of CpG islands in target genes. Similarly, cell-cycle genes downstream of the p53/RB pathway were upregulated only in overt HCC, with TP53 or RB1 gene mutations associated with chromosomal deletion of 4q or 16q. HCC was epigenetically distinguished into four subclasses: normal-like methylation, global-hypomethylation (favorable prognosis), stem-like methylation (poor prognosis), and CpG island methylation. These methylation statuses were globally maintained through HCC progression. Collectively, these data show that as HCC progresses, additional molecular events exclusive of driver gene mutations cooperatively contribute to transcriptional activation of downstream targets according to methylation status.
Significance:
In addition to driver gene mutations in the WNT and p53 pathways, further molecular events are required for aberrant transcriptional activation of these pathways as HCC progresses.
Data from Upregulation of E3 Ubiquitin Ligase CBLC Enhances EGFR Dysregulation and Signaling in Lung Adenocarcinoma
Authors: Hong Shiao-Ya , Kao Yu-Rung , Lee Te-Chang , Wu Cheng-Wen
Published: March 2023

Abstract
CBLC (CBL proto-oncogene c) belongs to the CBL protein family, which has E3 ubiquitin ligase activity toward activated receptor tyrosine kinases. CBLC is frequently upregulated in non–small cell lung cancer (NSCLC), yet very little is known about the functions of CBLC in tumorigenesis. Here we show that CBLC is an epigenetically demethylated target and its expression can be upregulated in NSCLC after treatment with the DNA methylation inhibitor 5′-azacytidine. Depletion of CBLC significantly inhibited cell viability and clonogenicity in vitro and reduced tumor growth in a xenograft model. CBLC silencing further sensitized EGFR-mutated NSCLC cells to treatment with tyrosine kinase inhibitors. Conversely, ectopic expression of CBLC enhanced the activation of EGFR and downstream ERK1/2 signaling after ligand stimulation by competing with CBL for EGFR binding. Analysis of ubiquitin linkages on activated EGFR (aEGFR) revealed that CBLC ubiquitinated and positively regulated aEGFR stability through the conjugation of polyubiquitin by K6 and K11 linkages. This CBLC-mediated polyubiquitination promoted either preferential recycling of aEGFR back to the plasma membrane or trafficking to the cell nucleus. IHC analyses revealed a positive correlation between phospho-EGFR and CBLC in lung adenocarcinoma. In summary, we demonstrate a novel mechanism by which aEGFR escapes lysosomal degradation in a CBLC/ubiquitin-dependent manner to sustain its activation. Our work identifies CBLC as a potential diagnostic biomarker and also points to its utilization as a novel therapeutic target for NSCLC therapy.

Significance: This work demonstrates the role of CBLC expression as a diagnostic biomarker and potential therapeutic target in lung adenocarcinoma. Cancer Res; 78(17); 4984–96. ©2018 AACR.
Data from The Cytochrome P450 Slow Metabolizers CYP2C9*2 and CYP2C9*3 Directly Regulate Tumorigenesis via Reduced Epoxyeicosatrienoic Acid Production
Authors: Sausville Lindsay N. , Gangadhariah Mahesha H. , Chiusa Manuel , Mei Shaojun , Wei Shouzuo , Zent Roy , Luther James M. , Shuey Megan M. , Capdevila Jorge H. , Falck John R. , Guengerich F. Peter , Williams Scott M. , Pozzi Ambra
Published: March 2023

Abstract
Increased expression of cytochrome P450 CYP2C9, together with elevated levels of its products epoxyeicosatrienoic acids (EET), is associated with aggressiveness in cancer. Cytochrome P450 variants CYP2C9*2 and CYP2C9*3 encode proteins with reduced enzymatic activity, and individuals carrying these variants metabolize drugs more slowly than individuals with wild-type CYP2C9*1, potentially affecting their response to drugs and altering their risk of disease. Although genetic differences in CYP2C9-dependent oxidation of arachidonic acid (AA) have been reported, the roles of CYP2C9*2 and CYP2C9*3 in EET biosynthesis and their relevance to disease are unknown. Here, we report that CYP2C9*2 and CYP2C9*3 metabolize AA less efficiently than CYP2C9*1 and that they play a role in the progression of non–small cell lung cancer (NSCLC) via impaired EET biosynthesis. When injected into mice, NSCLC cells expressing CYP2C9*2 and CYP2C9*3 produced lower levels of EETs and developed fewer, smaller, and less vascularized tumors than cells expressing CYP2C9*1. Moreover, endothelial cells expressing these two variants proliferated and migrated less than cells expressing CYP2C*1. Purified CYP2C9*2 and CYP2C9*3 exhibited attenuated catalytic efficiency in producing EETs, primarily due to impaired reduction of these two variants by NADPH-P450 reductase. Loss-of-function SNPs within CYP2C9*2 and CYP2C9*3 were associated with improved survival in female cases of NSCLC. Thus, decreased EET biosynthesis represents a novel mechanism whereby CYPC29*2 and CYP2C9*3 exert a direct protective role in NSCLC development.

Significance: These findings report single nucleotide polymorphisms in the human CYP2C9 genes, CYP2C9*2 and CYP2C9*3, exert a direct protective role in tumorigenesis by impairing EET biosynthesis. Cancer Res; 78(17); 4865–77. ©2018 AACR.
Data from Circulating Proteoglycan Endocan Mediates EGFR-Driven Progression of Non–Small Cell Lung Cancer
Authors: Yang Yi-Chieh , Pan Ke-Fan , Lee Wei-Jiunn , Chang Jer-Hwa , Tan Peng , Gu Chia-Chi , Chang Wei-Min , Yang Shun-Fa , Hsiao Michael , Hua Kuo-Tai , Chien Ming-Hsien
Published: March 2023

Abstract
Although new generations of EGFR-tyrosine kinase inhibitors (EGFR-TKI) have been developed for the treatment of patients with non–small cell lung cancer (NSCLC) with EGFR-mutant tumors, TKI resistance often returns as a result of additional EGFR mutations. In addition to seeking for next-generation EGFR-TKI, developing novel EGFR-targeting strategies may hold the key to overcome the vicious cycle of TKI resistance. Endocan is known as a receptor tyrosine kinase ligand enhancer in tumorigenesis, but the impact of endocan on EGFR-driven NSCLC progression remains unknown. In this study, higher endocan levels were found in lung tumors compared with cancer-free tissues and correlated with poor prognosis in patients with NSCLC harboring mutant EGFR; circulating endocan levels were also significantly higher in patients with mutant EGFR. Endocan facilitated EGFR signaling via direct binding and enhancing of the EGF–EGFR interaction and supported the growth of tumors driven by mutated EGFR. Activated EGFR in turn upregulated expression of endocan via JAK/STAT3 and ERK/ELK cascades, thus forming a positive regulatory loop of endocan-EGFR signaling. On the basis of the binding region between endocan and EGFR, we designed therapeutic peptides and demonstrated promising therapeutic effects in xenografts harboring EGFR mutations including TKI-resistant T790M. Together, our findings highlight the novel interaction between endocan and EGFR and new opportunities to effectively target endocan-EGFR regulatory axis in patients with TKI-resistant NSCLC.
Significance:
Endocan is a novel and critical regulator of EGF/EGFR signaling and serves as an alternative target of EGFR-TKI resistance in NSCLC.
Data from Genomic Predictors of Good Outcome, Recurrence, or Progression in High-Grade T1 Non–Muscle-Invasive Bladder Cancer
Authors: Bellmunt Joaquim , Kim Jaegil , Reardon Brendan , Perera-Bel Júlia , Orsola Anna , Rodriguez-Vida Alejo , Wankowicz Stephanie A. , Bowden Michaela , Barletta Justine A. , Morote Juan , de Torres Inés , Juanpere Nuria , Lloreta-Trull Josep , Hernandez Silvia , Mouw Kent W. , Taplin Mary-Ellen , Cejas Paloma , Long Henry W. , Van Allen Eliezer M. , Getz Gad , Kwiatkowski David J.
Published: March 2023

Abstract
High-grade T1 (HGT1) bladder cancer is the highest risk subtype of non–muscle-invasive bladder cancer with unpredictable outcome and poorly understood risk factors. Here, we examined the association of somatic mutation profiles with nonrecurrent disease (GO, good outcome), recurrence (R), or progression (PD) in a cohort of HGT1 patients. Exome sequencing was performed on 62 HGT1 and 15 matched normal tissue samples. Both tumor only (TO) and paired analyses were performed, focusing on 95 genes known to be mutated in bladder cancer. Somatic mutations, copy-number alterations, mutation load, and mutation signatures were studied. Thirty-three GO, 10 R, 18 PD, and 1 unknown outcome patients were analyzed. Tumor mutational burden (TMB) was similar to muscle-invasive disease and was highest in GO, intermediate in PD, and lowest in R patients (P = 0.017). DNA damage response gene mutations were associated with higher TMB (P < 0.0001) and GO (P = 0.003). ERCC2 and BRCA2 mutations were associated with GO. TP53, ATM, ARID1A, AHR, and SMARCB1 mutations were more frequent in PD. Focal copy-number gain in CCNE1 and CDKN2A deletion was enriched in PD or R (P = 0.047; P = 0.06). APOBEC (46%) and COSMIC5 (34%) signatures were most frequent. APOBEC-A and ERCC2 mutant tumors (COSMIC5) were associated with GO (P = 0.047; P = 0.0002). pT1b microstaging was associated with a genomic cluster (P = 0.05) with focal amplifications of E2F3/SOX4, PVRL4, CCNE1, and TP53 mutations. Findings were validated using external public datasets. These findings require confirmation but suggest that management of HGT1 bladder cancer may be improved via molecular characterization to predict outcome.
Significance:
Detailed genetic analyses of HGT1 bladder tumors identify features that correlate with outcome, e.g., high mutational burden, ERCC2 mutations, and high APOBEC-A/ERCC2 mutation signatures were associated with good outcome.
Data from Gain of HIF1 Activity and Loss of miRNA let-7d Promote Breast Cancer Metastasis to the Brain via the PDGF/PDGFR Axis
Authors: Wyss Christof B. , Duffey Nathalie , Peyvandi Sanam , Barras David , Martinez Usatorre Amaïa , Coquoz Oriana , Romero Pedro , Delorenzi Mauro , Lorusso Girieca , Rüegg Curzio
Published: March 2023

Abstract
Early detection and adjuvant therapies have significantly improved survival of patients with breast cancer over the past three decades. In contrast, management of metastatic disease remains unresolved. Brain metastasis is a late complication frequently observed among patients with metastatic breast cancer, whose poor prognosis calls for novel and more effective therapies. Here, we report that active hypoxia inducible factor-1 (HIF1) signaling and loss of the miRNA let-7d concur to promote brain metastasis in a recently established model of spontaneous breast cancer metastasis from the primary site to the brain (4T1-BM2), and additionally in murine and human experimental models of breast cancer brain metastasis (D2A1-BM2 and MDA231-BrM2). Active HIF1 and let-7d loss upregulated expression of platelet-derived growth factor (PDGF) B/A in murine and human brain metastatic cells, respectively, while either individual silencing of HIF1α and PDGF-A/B or let-7d overexpression suppressed brain metastasis formation in the tested models. Let-7d silencing upregulated HIF1α expression and HIF1 activity, indicating a regulatory hierarchy of the system. The clinical relevance of the identified targets was supported by human gene expression data analyses. Treatment of mice with nilotinib, a kinase inhibitor impinging on PDGF receptor (PDGFR) signaling, prevented formation of spontaneous brain metastases in the 4T1-BM2 model and reduced growth of established brain metastases in mouse and human models. These results identify active HIF1 signaling and let-7d loss as coordinated events promoting breast cancer brain metastasis through increased expression of PDGF-A/B. Moreover, they identify PDGFR inhibition as a potentially actionable therapeutic strategy for patients with brain metastatis.
Significance:
These findings show that loss of miRNA let-7d and active HIF1 signaling promotes breast cancer brain metastasis via PDGF and that pharmacologic inhibition of PDGFR suppresses brain metastasis, suggesting novel therapeutic opportunities.
Data from Plasma Gelsolin Inhibits CD8+ T-cell Function and Regulates Glutathione Production to Confer Chemoresistance in Ovarian Cancer
Authors: Asare-Werehene Meshach , Communal Laudine , Carmona Euridice , Han Youngjin , Song Yong Sang , Burger Dylan , Mes-Masson Anne-Marie , Tsang Benjamin K.
Published: March 2023

Abstract
Although initial treatment of ovarian cancer is successful, tumors typically relapse and become resistant to treatment. Because of poor infiltration of effector T cells, patients are mostly unresponsive to immunotherapy. Plasma gelsolin (pGSN) is transported by exosomes (small extracellular vesicle, sEV) and plays a key role in ovarian cancer chemoresistance, yet little is known about its role in immunosurveillance. Here, we report the immunomodulatory roles of sEV-pGSN in ovarian cancer chemoresistance. In chemosensitive conditions, secretion of sEV-pGSN was low, allowing for optimal CD8+ T-cell function. This resulted in increased T-cell secretion of IFNγ, which reduced intracellular glutathione (GSH) production and sensitized chemosensitive cells to cis-diaminedichloroplatinum (CDDP)-induced apoptosis. In chemoresistant conditions, increased secretion of sEV-pGSN by ovarian cancer cells induced apoptosis in CD8+ T cells. IFNγ secretion was therefore reduced, resulting in high GSH production and resistance to CDDP-induced death in ovarian cancer cells. These findings support our hypothesis that sEV-pGSN attenuates immunosurveillance and regulates GSH biosynthesis, a phenomenon that contributes to chemoresistance in ovarian cancer.
Significance:
These findings provide new insight into pGSN-mediated immune cell dysfunction in ovarian cancer chemoresistance and demonstrate how this dysfunction can be exploited to enhance immunotherapy.
Data from Prospective Identification of Elevated Circulating CDCP1 in Patients Years before Onset of Lung Cancer
Authors: Dagnino Sonia , Bodinier Barbara , Guida Florence , Smith-Byrne Karl , Petrovic Dusan , Whitaker Matthew D. , Haugdahl Nøst Therese , Agnoli Claudia , Palli Domenico , Sacerdote Carlotta , Panico Salvatore , Tumino Rosario , Schulze Matthias B. , Johansson Mikael , Keski-Rahkonen Pekka , Scalbert Augustin , Vineis Paolo , Johansson Mattias , Sandanger Torkjel M. , Vermeulen Roel C.H. , Chadeau-Hyam Marc
Published: March 2023

Abstract
Increasing evidence points to a role for inflammation in lung carcinogenesis. A small number of circulating inflammatory proteins have been identified as showing elevated levels prior to lung cancer diagnosis, indicating the potential for prospective circulating protein concentration as a marker of early carcinogenesis. To identify novel markers of lung cancer risk, we measured a panel of 92 circulating inflammatory proteins in 648 prediagnostic blood samples from two prospective cohorts in Italy and Norway (women only). To preserve the comparability of results and protect against confounding factors, the main statistical analyses were conducted in women from both studies, with replication sought in men (Italian participants). Univariate and penalized regression models revealed for the first time higher blood levels of CDCP1 protein in cases that went on to develop lung cancer compared with controls, irrespective of time to diagnosis, smoking habits, and gender. This association was validated in an additional 450 samples. Associations were stronger for future cases of adenocarcinoma where CDCP1 showed better explanatory performance. Integrative analyses combining gene expression and protein levels of CDCP1 measured in the same individuals suggested a link between CDCP1 and the expression of transcripts of LRRN3 and SEM1. Enrichment analyses indicated a potential role for CDCP1 in pathways related to cell adhesion and mobility, such as the WNT/β-catenin pathway. Overall, this study identifies lung cancer–related dysregulation of CDCP1 expression years before diagnosis.
Significance:
Prospective proteomics analyses reveal an association between increased levels of circulating CDCP1 and lung carcinogenesis irrespective of smoking and years before diagnosis, and integrating gene expression indicates potential underlying mechanisms.

See related commentary by Itzstein et al., p. 3441.
Data from Disabling the Fanconi Anemia Pathway in Stem Cells Leads to Radioresistance and Genomic Instability
Authors: Deng Xinzhu , Tchieu Jason , Higginson Daniel S. , Hsu Kuo-Shun , Feldman Regina , Studer Lorenz , Shaham Shai , Powell Simon N. , Fuks Zvi , Kolesnick Richard
Published: March 2023

Abstract
Fanconi anemia is an inherited genome instability syndrome characterized by interstrand cross-link hypersensitivity, congenital defects, bone marrow failure, and cancer predisposition. Although DNA repair mediated by Fanconi anemia genes has been extensively studied, how inactivation of these genes leads to specific cellular phenotypic consequences associated with Fanconi anemia is not well understood. Here we report that Fanconi anemia stem cells in the C. elegans germline and in murine embryos display marked nonhomologous end joining (NHEJ)–dependent radiation resistance, leading to survival of progeny cells carrying genetic lesions. In contrast, DNA cross-linking does not induce generational genomic instability in Fanconi anemia stem cells, as widely accepted, but rather drives NHEJ-dependent apoptosis in both species. These findings suggest that Fanconi anemia is a stem cell disease reflecting inappropriate NHEJ, which is mutagenic and carcinogenic as a result of DNA misrepair, while marrow failure represents hematopoietic stem cell apoptosis.
Significance:
This study finds that Fanconi anemia stem cells preferentially activate error-prone NHEJ-dependent DNA repair to survive irradiation, thereby conferring generational genomic instability that is instrumental in carcinogenesis.
Data from Neuroblastoma Formation Requires Unconventional CD4 T Cells and Arginase-1–Dependent Myeloid Cells
Authors: Van de Velde Lee-Ann , Allen E. Kaitlynn , Crawford Jeremy Chase , Wilson Taylor L. , Guy Clifford S. , Russier Marion , Zeitler Leonie , Bahrami Armita , Finkelstein David , Pelletier Stephane , Schultz-Cherry Stacey , Thomas Paul G. , Murray Peter J.
Published: March 2023

Abstract
Immune cells regulate tumor growth by mirroring their function as tissue repair organizers in normal tissues. To understand the different facets of immune–tumor collaboration through genetics, spatial transcriptomics, and immunologic manipulation with noninvasive, longitudinal imaging, we generated a penetrant double oncogene–driven autochthonous model of neuroblastoma. Spatial transcriptomic analysis showed that CD4+ and myeloid populations colocalized within the tumor parenchyma, while CD8+ T cells and B cells were peripherally dispersed. Depletion of CD4+ T cells or CCR2+ macrophages, but not B cells, CD8+ T cells, or natural killer (NK) cells, prevented tumor formation. Tumor CD4+ T cells displayed unconventional phenotypes and were clonotypically diverse and antigen independent. Within the myeloid fraction, tumor growth required myeloid cells expressing arginase-1. Overall, these results demonstrate how arginine-metabolizing myeloid cells conspire with pathogenic CD4+ T cells to create permissive conditions for tumor formation, suggesting that these protumorigenic pathways could be disabled by targeting myeloid arginine metabolism.
Significance:
A new model of human neuroblastoma provides ways to track tumor formation and expansion in living animals, allowing identification of CD4+ T-cell and macrophage functions required for oncogenesis.
Data from BRD4 Regulates Transcription Factor ΔNp63α to Drive a Cancer Stem Cell Phenotype in Squamous Cell Carcinomas
Authors: Fisher Matthew L. , Balinth Seamus , Hwangbo Yon , Wu Caizhi , Ballon Carlos , Wilkinson John E. , Goldberg Gary L. , Mills Alea A.
Published: March 2023

Abstract
Bromodomain containing protein 4 (BRD4) plays a critical role in controlling the expression of genes involved in development and cancer. Inactivation of BRD4 inhibits cancer growth, making it a promising anticancer drug target. The cancer stem cell (CSC) population is a key driver of recurrence and metastasis in patients with cancer. Here we show that cancer stem-like cells can be enriched from squamous cell carcinomas (SCC), and that these cells display an aggressive phenotype with enhanced stem cell marker expression, migration, invasion, and tumor growth. BRD4 is highly elevated in this aggressive subpopulation of cells, and its function is critical for these CSC-like properties. Moreover, BRD4 regulates ΔNp63α, a key transcription factor that is essential for epithelial stem cell function that is often overexpressed in cancers. BRD4 regulates an EZH2/STAT3 complex that leads to increased ΔNp63α-mediated transcription. Targeting BRD4 in human SCC reduces ΔNp63α, leading to inhibition of spheroid formation, migration, invasion, and tumor growth. These studies identify a novel BRD4-regulated signaling network in a subpopulation of cancer stem-like cells, elucidating a possible avenue for effective therapeutic intervention.
Significance:
This study identifies a signaling cascade driven by BRD4 that upregulates ΔNp63α to promote cancer stem-like properties, which has potential therapeutic implications for the treatment of squamous cell carcinomas.
Data from Tie2 Receptor in Tumor-Infiltrating Macrophages Is Dispensable for Tumor Angiogenesis and Tumor Relapse after Chemotherapy
Authors: Jakab Moritz , Rostalski Till , Lee Ki Hong , Mogler Carolin , Augustin Hellmut G.
Published: March 2023

Abstract
Tumor relapse after chemotherapy relies on the reconstruction of damaged tumor vasculature. In this context, proangiogenic Tie2-expressing macrophages have been suggested to serve as crucial instructors of tumor revascularization by secreting angiogenic factors while being closely associated with the vessel wall. Although the proangiogenic nature of Tie2+ macrophages is well described, the functional contribution of macrophage Tie2 expression remains elusive. Here, we employed a Cre-loxP system to specifically delete Tie2 in macrophages. In multiple syngeneic solid tumor models and two distinct chemotherapeutic treatment regimens, macrophage-expressed Tie2 did not contribute to primary tumor growth, tumor revascularization after chemotherapy, tumor recurrence, or metastasis. Exposing cultured murine macrophage cell lines and bone marrow–derived macrophages to hypoxia or stimulating them with Ang2 did not induce expression of Tie2 at the RNA or protein level. Furthermore, a comprehensive meta-analysis of publicly available single cell RNA sequencing datasets of human and murine tumor-infiltrating CD11b+ myeloid cells did not reveal a transcriptionally distinct macrophage population marked by the expression of Tie2. Collectively, these data question the previously reported critical role of Tie2-expressing macrophages for tumor angiogenesis and tumor relapse after chemotherapy. Moreover, lack of Tie2 inducibility and absence of Tie2-positive macrophages in multiple recently published tumor studies refute a possible prognostic value of macrophage-expressed Tie2.
Significance:
Multiple preclinical tumor models, cell stimulation experiments, and meta-analysis of published tumor single cell RNA sequencing data challenge the reported role of Tie2-positive macrophages for tumor angiogenesis, metastasis, and relapse after chemotherapy.

See related commentary by Zhang and Brekken, p. 1172
Data from Novel Mouse Models of Bladder Cancer Identify a Prognostic Signature Associated with Risk of Disease Progression
Authors: Park Soonbum , Rong Lijie , Owczarek Tomasz B. , Bernardo Matteo Di , Shoulson Rivka L. , Chua Chee-Wai , Kim Jaime Y. , Lankarani Amir , Chakrapani Prithi , Syed Talal , McKiernan James M. , Solit David B. , Shen Michael M. , Al-Ahmadie Hikmat A. , Abate-Shen Cory
Published: March 2023

Abstract
To study the progression of bladder cancer from non–muscle-invasive to muscle-invasive disease, we have developed a novel toolkit that uses complementary approaches to achieve gene recombination in specific cell populations in the bladder urothelium in vivo, thereby allowing us to generate a new series of genetically engineered mouse models (GEMM) of bladder cancer. One method is based on the delivery of adenoviruses that express Cre recombinase in selected cell types in the urothelium, and a second uses transgenic drivers in which activation of inducible Cre alleles can be limited to the bladder urothelium by intravesicular delivery of tamoxifen. Using both approaches, targeted deletion of the Pten and p53 tumor suppressor genes specifically in basal urothelial cells gave rise to muscle-invasive bladder tumors. Furthermore, preinvasive lesions arising in basal cells displayed upregulation of molecular pathways related to bladder tumorigenesis, including proinflammatory pathways. Cross-species analyses comparing a mouse gene signature of early bladder cancer with a human signature of bladder cancer progression identified a conserved 28-gene signature of early bladder cancer that is associated with poor prognosis for human bladder cancer and that outperforms comparable gene signatures. These findings demonstrate the relevance of these GEMMs for studying the biology of human bladder cancer and introduce a prognostic gene signature that may help to stratify patients at risk for progression to potentially lethal muscle-invasive disease.
Significance:
Analyses of bladder cancer progression in a new series of genetically engineered mouse models has identified a gene signature of poor prognosis in human bladder cancer.
Data from Transient Inhibition of the JAK/STAT Pathway Prevents B-ALL Development in Genetically Predisposed Mice
Authors: Casado-García Ana , Isidro-Hernández Marta , Oak Ninad , Mayado Andrea , Mann-Ran Christine , Raboso-Gallego Javier , Alemán-Arteaga Silvia , Buhles Alexandra , Sterker Dario , Sánchez Elena G. , Martínez-Cano Jorge , Blanco Oscar , Orfao Alberto , Alonso-López Diego , De Las Rivas Javier , Riesco Susana , Prieto-Matos Pablo , González-Murillo África , García Criado Francisco Javier , García Cenador María Begoña , Radimerski Thomas , Ramírez-Orellana Manuel , Cobaleda César , Yang Jun J. , Vicente-Dueñas Carolina , Weiss Andreas , Nichols Kim E. , Sánchez-García Isidro
Published: March 2023

Abstract
Preventing development of childhood B-cell acute lymphoblastic leukemia (B-ALL), a disease with devastating effects, is a longstanding and unsolved challenge. Heterozygous germline alterations in the PAX5 gene can lead to B-ALL upon accumulation of secondary mutations affecting the JAK/STAT signaling pathway. Preclinical studies have shown that this malignant transformation occurs only under immune stress such as exposure to infectious pathogens. Here we show in Pax5+/− mice that transient, early-life administration of clinically relevant doses of ruxolitinib, a JAK1/2 inhibitor, significantly mitigates the risk of B-ALL following exposure to infection; 1 of 29 animals treated with ruxolitinib developed B-ALL versus 8 of 34 untreated mice. Ruxolitinib treatment preferentially targeted Pax5+/− versus wild-type B-cell progenitors and exerted unique effects on the Pax5+/− B-cell progenitor transcriptional program. These findings provide the first in vivo evidence for a potential strategy to prevent B-ALL development.
Significance:
JAK/STAT inhibition suppresses tumorigenesis in a B-ALL–susceptible mouse model, presenting a novel approach to prevent B-ALL onset.
Data from Genomic and Transcriptomic Analysis of Relapsed and Refractory Childhood Solid Tumors Reveals a Diverse Molecular Landscape and Mechanisms of Immune Evasion
Authors: Byron Sara A. , Hendricks William P.D. , Nagulapally Abhinav B. , Kraveka Jacqueline M. , Ferguson William S. , Brown Valerie I. , Eslin Don E. , Mitchell Deanna , Cornelius Albert , Roberts William , Isakoff Michael S. , Oesterheld Javier E. , Wada Randal K. , Rawwas Jawhar , Neville Kathleen , Zage Peter E. , Harrod Virginia L. , Bergendahl Genevieve , VanSickle Elizabeth , Dykema Karl , Bond Jeffrey , Chou Hsien-Chao , Wei Jun S. , Wen Xinyu , Reardon Hue V. , Roos Alison , Nasser Sara , Izatt Tyler , Enriquez Daniel , Hegde Apurva M. , Cisneros Faith , Christofferson Austin , Turner Bryce , Szelinger Szabolcs , Keats Jonathan J. , Halperin Rebecca F. , Khan Javed , Saulnier Sholler Giselle L. , Trent Jeffrey M.
Published: March 2023

Abstract
Children with treatment-refractory or relapsed (R/R) tumors face poor prognoses. As the genomic underpinnings driving R/R disease are not well defined, we describe here the genomic and transcriptomic landscapes of R/R solid tumors from 202 patients enrolled in Beat Childhood Cancer Consortium clinical trials. Tumor mutational burden (TMB) was elevated relative to untreated tumors at diagnosis, with one-third of tumors classified as having a pediatric high TMB. Prior chemotherapy exposure influenced the mutational landscape of these R/R tumors, with more than 40% of tumors demonstrating mutational signatures associated with platinum or temozolomide chemotherapy and two tumors showing treatment-associated hypermutation. Immunogenomic profiling found a heterogenous pattern of neoantigen and MHC class I expression and a general absence of immune infiltration. Transcriptional analysis and functional gene set enrichment analysis identified cross-pathology clusters associated with development, immune signaling, and cellular signaling pathways. While the landscapes of these R/R tumors reflected those of their corresponding untreated tumors at diagnosis, important exceptions were observed, suggestive of tumor evolution, treatment resistance mechanisms, and mutagenic etiologies of treatment.
Significance:
Tumor heterogeneity, chemotherapy exposure, and tumor evolution contribute to the molecular profiles and increased mutational burden that occur in treatment-refractory and relapsed childhood solid tumors.
Data from Preclinical In Vivo Validation of the RAD51 Test for Identification of Homologous Recombination-Deficient Tumors and Patient Stratification
Authors: Pellegrino Benedetta , Herencia-Ropero Andrea , Llop-Guevara Alba , Pedretti Flaminia , Moles-Fernández Alejandro , Viaplana Cristina , Villacampa Guillermo , Guzmán Marta , Rodríguez Olga , Grueso Judit , Jiménez Jose , Arenas Enrique J. , Degasperi Andrea , Dias João M.L. , Forment Josep V. , O'Connor Mark J. , Déas Olivier , Cairo Stefano , Zhou Yinghui , Musolino Antonino , Caldas Carlos , Nik-Zainal Serena , Clarke Robert B. , Nuciforo Paolo , Díez Orland , Serres-Créixams Xavier , Peg Vicente , Espinosa-Bravo Martín , Macarulla Teresa , Oaknin Ana , Mateo Joaquin , Arribas Joaquín , Dienstmann Rodrigo , Bellet Meritxell , Oliveira Mafalda , Saura Cristina , Gutiérrez-Enríquez Sara , Balmaña Judith , Serra Violeta
Published: March 2023

Abstract
PARP inhibitors (PARPi) are approved drugs for platinum-sensitive, high-grade serous ovarian cancer (HGSOC) and for breast, prostate, and pancreatic cancers (PaC) harboring genetic alterations impairing homologous recombination repair (HRR). Detection of nuclear RAD51 foci in tumor cells is a marker of HRR functionality, and we previously established a test to detect RAD51 nuclear foci. Here, we aimed to validate the RAD51 score cut off and compare the performance of this test to other HRR deficiency (HRD) detection methods. Laboratory models from BRCA1/BRCA2-associated breast cancer, HGSOC, and PaC were developed and evaluated for their response to PARPi and cisplatin. HRD in these models and patient samples was evaluated by DNA sequencing of HRR genes, genomic HRD tests, and RAD51 foci detection. We established patient-derived xenograft models from breast cancer (n = 103), HGSOC (n = 4), and PaC (n = 2) that recapitulated patient HRD status and treatment response. The RAD51 test showed higher accuracy than HRR gene mutations and genomic HRD analysis for predicting PARPi response (95%, 67%, and 71%, respectively). RAD51 detection captured dynamic changes in HRR status upon acquisition of PARPi resistance. The accuracy of the RAD51 test was similar to HRR gene mutations for predicting platinum response. The predefined RAD51 score cut off was validated, and the high predictive value of the RAD51 test in preclinical models was confirmed. These results collectively support pursuing clinical assessment of the RAD51 test in patient samples from randomized trials testing PARPi or platinum-based therapies.
Significance:
This work demonstrates the high accuracy of a histopathology-based test based on the detection of RAD51 nuclear foci in predicting response to PARPi and cisplatin.
Data from Elimination of Radiation-Induced Senescence in the Brain Tumor Microenvironment Attenuates Glioblastoma Recurrence
Authors: Fletcher-Sananikone Eliot , Kanji Suman , Tomimatsu Nozomi , Di Cristofaro Luis Fernando Macedo , Kollipara Rahul K. , Saha Debabrata , Floyd John R. , Sung Patrick , Hromas Robert , Burns Terry C. , Kittler Ralf , Habib Amyn A. , Mukherjee Bipasha , Burma Sandeep
Published: March 2023

Abstract
Glioblastomas (GBM) are routinely treated with ionizing radiation (IR) but inevitably recur and develop therapy resistance. During treatment, the tissue surrounding tumors is also irradiated. IR potently induces senescence, and senescent stromal cells can promote the growth of neighboring tumor cells by secreting factors that create a senescence-associated secretory phenotype (SASP). Here, we carried out transcriptomic and tumorigenicity analyses in irradiated mouse brains to elucidate how radiotherapy-induced senescence of non-neoplastic brain cells promotes tumor growth. Following cranial irradiation, widespread senescence in the brain occurred, with the astrocytic population being particularly susceptible. Irradiated brains showed an altered transcriptomic profile characterized by upregulation of CDKN1A (p21), a key enforcer of senescence, and several SASP factors, including HGF, the ligand of the receptor tyrosine kinase (RTK) Met. Preirradiation of mouse brains increased Met-driven growth and invasiveness of orthotopically implanted glioma cells. Importantly, irradiated p21−/− mouse brains did not exhibit senescence and consequently failed to promote tumor growth. Senescent astrocytes secreted HGF to activate Met in glioma cells and to promote their migration and invasion in vitro, which could be blocked by HGF-neutralizing antibodies or the Met inhibitor crizotinib. Crizotinib also slowed the growth of glioma cells implanted in preirradiated brains. Treatment with the senolytic drug ABT-263 (navitoclax) selectively killed senescent astrocytes in vivo, significantly attenuating growth of glioma cells implanted in preirradiated brains. These results indicate that SASP factors in the irradiated tumor microenvironment drive GBM growth via RTK activation, underscoring the potential utility of adjuvant senolytic therapy for preventing GBM recurrence after radiotherapy.
Significance:
This study uncovers mechanisms by which radiotherapy can promote GBM recurrence by inducing senescence in non-neoplastic brain cells, suggesting that senolytic therapy can blunt recurrent GBM growth and aggressiveness.
Data from Aberrant Transcript Usage Is Associated with Homologous Recombination Deficiency and Predicts Therapeutic Response
Authors: Kang Hyeon Gu , Hwangbo Haeun , Kim Myung Ji , Kim Sinae , Lee Eun Ji , Park Min Ji , Kim Jae-Weon , Kim Byoung-Gie , Cho Eun-Hae , Chang Suhwan , Lee Jung-Yun , Choi Jung Kyoon
Published: March 2023

Abstract
BRCA1/2 mutations account for only a small fraction of homologous recombination (HR) deficiency (HRD) cases. Recently developed genomic HRD (gHRD) tests suffer confounding factors that cause low precision in predicting samples that will respond to PARP inhibitors and DNA damaging agents. Here we present molecular and clinical evidence of transcriptional HRD (tHRD) that is based on aberrant transcript usage (aTU) of minor isoforms. Specifically, increased TU of nonfunctional isoforms of DNA repair genes was prevalent in breast and ovarian cancer with gHRD. Functional assays validated the association of aTU with impaired HR activity. Machine learning–based tHRD detection by the transcript usage (TU) pattern of key genes was superior to directly screening for gHRD or BRCA1/2 mutations in accurately predicting responses of cell lines and patients with cancer to PARP inhibitors and genotoxic drugs. This approach demonstrated the capability of tHRD status to reflect functional HR status, including in a cohort of olaparib-treated ovarian cancer with acquired platinum resistance. Diagnostic tests based on tHRD are expected to broaden the clinical utility of PARP inhibitors.
Significance:
A novel but widespread transcriptional mechanism by which homologous recombination deficiency arises independently of BRCA1/2 mutations can be utilized as a companion diagnostic for PARP inhibitors.
Data from Functional Genomic Analysis of CDK4 and CDK6 Gene Dependency across Human Cancer Cell Lines
Authors: Zhang Zhouwei , Golomb Lior , Meyerson Matthew
Published: March 2023

Abstract
Cyclin-dependent kinase 4 (CDK4) and CDK6 are key cell-cycle regulators that are frequently dysregulated in human malignancies. CDK4/6 inhibitors are clinically approved for the treatment of hormone receptor–positive, HER2–negative (HR+/HER2−) breast cancer, but improved specificity and reduced toxicity might expand their use to other indications. Through analysis of publicly available genome-wide loss-of-function data combined with single and dual-targeting CRISPR assays, we found differential cell proliferation vulnerability of cell lines to either CDK4 deletion alone, CDK6 deletion alone, combined CDK4/CDK6 deletion, or neither. CDK6 expression was the best single predictor of CDK4 (negatively correlated) and CDK6 (positively correlated) dependencies in the cancer cell lines, with adenocarcinoma cell lines being more sensitive to CDK4 deletion and hematologic and squamous cancer cell lines being more sensitive to CDK6 deletion. RB–E2F signaling was confirmed as a main downstream node of CDK4/6 in these experiments as shown by the survival effects of RB1 deletion. Finally, we show in a subset of cancer cell lines not dependent on CDK4/6 that CDK2–CCNE1 is an important alternative dependency for cell proliferation. Together, our comprehensive data exploration and functional experiments delineate the landscape of pan-cancer CDK4/6 gene dependencies and define unique cancer cell populations that might be sensitive to CDK4-selective or CDK6-selective inhibitors.
Significance:
This study provides functional genomic insight toward understanding the scenarios in which cancer cells are differentially sensitive to CDK4 or CDK6 inhibition and their implications in current treatment strategies.
Data from Quantitative Micro-Elastography Enables In Vivo Detection of Residual Cancer in the Surgical Cavity during Breast-Conserving Surgery
Authors: Gong Peijun , Chin Synn Lynn , Allen Wes M. , Ballal Helen , Anstie James D. , Chin Lixin , Ismail Hina M. , Zilkens Renate , Lakhiani Devina D. , McCarthy Matthew , Fang Qi , Firth Daniel , Newman Kyle , Thomas Caleb , Li Jiayue , Sanderson Rowan W. , Foo Ken Y. , Yeomans Chris , Dessauvagie Benjamin F. , Latham Bruce , Saunders Christobel M. , Kennedy Brendan F.
Published: March 2023

Abstract
Breast-conserving surgery (BCS) is commonly used for the treatment of early-stage breast cancer. Following BCS, approximately 20% to 30% of patients require reexcision because postoperative histopathology identifies cancer in the surgical margins of the excised specimen. Quantitative micro-elastography (QME) is an imaging technique that maps microscale tissue stiffness and has demonstrated a high diagnostic accuracy (96%) in detecting cancer in specimens excised during surgery. However, current QME methods, in common with most proposed intraoperative solutions, cannot image cancer directly in the patient, making their translation to clinical use challenging. In this proof-of-concept study, we aimed to determine whether a handheld QME probe, designed to interrogate the surgical cavity, can detect residual cancer directly in the breast cavity in vivo during BCS. In a first-in-human study, 21 BCS patients were scanned in vivo with the QME probe by five surgeons. For validation, protocols were developed to coregister in vivo QME with postoperative histopathology of the resected tissue to assess the capability of QME to identify residual cancer. In four cavity aspects presenting cancer and 21 cavity aspects presenting benign tissue, QME detected elevated stiffness in all four cancer cases, in contrast to low stiffness observed in 19 of the 21 benign cases. The results indicate that in vivo QME can identify residual cancer by directly imaging the surgical cavity, potentially providing a reliable intraoperative solution that can enable more complete cancer excision during BCS.
Significance:
Optical imaging of microscale tissue stiffness enables the detection of residual breast cancer directly in the surgical cavity during breast-conserving surgery, which could potentially contribute to more complete cancer excision.
Data from The X-Linked Helicase DDX3X Is Required for Lymphoid Differentiation and MYC-Driven Lymphomagenesis
Authors: Lacroix Marion , Beauchemin Hugues , Fraszczak Jennifer , Ross Julie , Shooshtarizadeh Peiman , Chen Riyan , Möröy Tarik
Published: March 2023

Abstract
The X-linked gene DDX3X encodes an RNA helicase that is mutated at high frequencies in several types of human B-cell lymphoma. Females have two active DDX3X alleles and males carry a DDX3Y homolog on the Y chromosome. We show here that pan-hematopoietic, homozygous deletion of Ddx3x in female mice perturbs erythropoiesis, causing early developmental arrest. However, both hemizygous male and heterozygous female embryos develop normally, suggesting that one Ddx3x allele is sufficient for fetal hematopoietic development in females and that the Ddx3y allele can compensate for the loss of Ddx3x in males. In adult mice, DDX3X deficiency altered hematopoietic progenitors, early lymphoid development, marginal zone and germinal center B cells, and lymphomagenesis in a sex-dependent manner. Loss of both Ddx3x alleles abrogated MYC-driven lymphomagenesis in females, whereas Ddx3x deletion in males did not affect the formation of B-cell lymphoma in both mouse models. Moreover, tumors that appeared in male mice lacking DDX3X showed upregulated expression of DDX3Y, indicating a critical requirement for DDX3 activity for lymphomagenesis. These data reveal sex-specific roles of DDX3X in erythro- and lymphopoiesis as well as in MYC-driven lymphomagenesis.
Significance:
The sex-dependent effects of DDX3X deficiency in malignant transformation of B cells and the compensatory role of DDX3Y support inhibition of DDX3 as a treatment strategy for MYC-driven B-cell lymphoma.
Data from Retinoic Acid Receptor Activation Reduces Metastatic Prostate Cancer Bone Lesions by Blocking the Endothelial-to-Osteoblast Transition
Authors: Yu Guoyu , Corn Paul G. , Shen Pengfei , Song Jian H. , Lee Yu-Chen , Lin Song-Chang , Pan Jing , Agarwal Sandeep K. , Panaretakis Theocharis , Pacifici Maurizio , Logothetis Christopher J. , Yu-Lee Li-Yuan , Lin Sue-Hwa
Published: March 2023

Abstract
Metastatic prostate cancer in the bone induces bone-forming lesions that contribute to progression and therapy resistance. Prostate cancer–induced bone formation originates from endothelial cells (EC) that have undergone endothelial-to-osteoblast (EC-to-OSB) transition in response to tumor-secreted BMP4. Current strategies targeting prostate cancer–induced bone formation are lacking. Here, we show that activation of retinoic acid receptor (RAR) inhibits EC-to-OSB transition and reduces prostate cancer–induced bone formation. Treatment with palovarotene, an RARγ agonist being tested for heterotopic ossification in fibrodysplasia ossificans progressiva, inhibited EC-to-OSB transition and osteoblast mineralization in vitro and decreased tumor-induced bone formation and tumor growth in several osteogenic prostate cancer models, and similar effects were observed with the pan-RAR agonist all-trans-retinoic acid (ATRA). Knockdown of RARα, β, or γ isoforms in ECs blocked BMP4-induced EC-to-OSB transition and osteoblast mineralization, indicating a role for all three isoforms in prostate cancer–induced bone formation. Furthermore, treatment with palovarotene or ATRA reduced plasma Tenascin C, a factor secreted from EC-OSB cells, which may be used to monitor treatment response. Mechanistically, BMP4-activated pSmad1 formed a complex with RAR in the nucleus of ECs to activate EC-to-OSB transition. RAR activation by palovarotene or ATRA caused pSmad1 degradation by recruiting the E3-ubiquitin ligase Smad ubiquitination regulatory factor1 (Smurf1) to the nuclear pSmad1/RARγ complex, thus blocking EC-to-OSB transition. Collectively, these findings suggest that palovarotene can be repurposed to target prostate cancer–induced bone formation to improve clinical outcomes for patients with bone metastasis.
Significance:
This study provides mechanistic insights into how RAR agonists suppress prostate cancer–induced bone formation and offers a rationale for developing RAR agonists for prostate cancer bone metastasis therapy.

See related commentary by Bhowmick and Bhowmick, p. 2975