bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Targeting AXL and mTOR Pathway Overcomes Primary and Acquired Resistance to WEE1 Inhibition in Small-Cell Lung Cancer
Authors: Sen Triparna , Tong Pan , Diao Lixia , Li Lerong , Fan Youhong , Hoff Jennifer , Heymach John V. , Wang Jing , Byers Lauren Averett
Published: April 2023

Abstract
Purpose: Drugs targeting DNA repair and cell-cycle checkpoints have emerged as promising therapies for small-cell lung cancer (SCLC). Among these, the WEE1 inhibitor AZD1775 has shown clinical activity in a subset of SCLC patients, but resistance is common. Understanding primary and acquired resistance mechanisms will be critical for developing effective WEE1 inhibitor combinations.

Experimental Design: AZD1775 sensitivity in SCLC cell lines was correlated with baseline expression level of 200 total or phosphorylated proteins measured by reverse-phase protein array (RPPA) to identify predictive markers of primary resistance. We further established AZD1775 acquired resistance models to identify mechanism of acquired resistance. Combination regimens were tested to overcome primary and acquired resistance to AZD1775 in in vitro and in vivo SCLC models.

Results: High-throughput proteomic profiling demonstrate that SCLC models with primary resistance to AZD1775 express high levels of AXL and phosphorylated S6 and that WEE1/AXL or WEE1/mTOR inhibitor combinations overcome resistance in vitro and in vivo. Furthermore, AXL, independently and via mTOR, activates the ERK pathway, leading to recruitment and activation of another G2-checkpoint protein, CHK1. AZD1775 acquired resistance models demonstrated upregulation of AXL, pS6, and MET, and resistance was overcome with the addition of AXL (TP0903), dual-AXL/MET (cabozantinib), or mTOR (RAD001) inhibitors.

Conclusions: AXL promotes resistance to WEE1 inhibition via downstream mTOR signaling and resulting activation of a parallel DNA damage repair pathway, CHK1. These findings suggest rational combinations to enhance the clinical efficacy of AZD1775, which is currently in clinical trials for SCLC and other malignancies. Clin Cancer Res; 23(20); 6239–53. ©2017 AACR.
Data from Depletion of FOXM1 via MET Targeting Underlies Establishment of a DNA Damage–Induced Senescence Program in Gastric Cancer
Authors: Francica Paola , Nisa Lluís , Aebersold Daniel M. , Langer Rupert , Bladt Friedhelm , Blaukat Andree , Stroka Deborah , Martínez María Rodríguez , Zimmer Yitzhak , Medová Michaela
Published: April 2023

Abstract
Purpose: Deregulated signaling via the MET receptor tyrosine kinase is abundant in gastric tumors, with up to 80% of cases displaying aberrant MET expression. A growing body of evidence suggests MET as a potential target for tumor radiosensitization.

Experimental Design: Cellular proliferation and DNA damage-induced senescence were studied in a panel of MET-overexpressing human gastric cancer cell lines as well as in xenograft models after MET inhibition and/or ionizing radiation. Pathways activation and protein expression were assessed by immunoblotting and immunohistochemistry. Tumor tissue microarrays (91 gastric cancer patients) were generated and copy number alteration (178 patients) and gene expression (373 patients) data available at The Cancer Genome Atlas were analyzed to assess the coalterations of MET and FOXM1.

Results: MET targeting administered before ionizing radiation instigates DNA damage–induced senescence (∼80%, P < 0.001) rather than cell death. MET inhibition–associated senescence is linked to the blockade of MAPK pathway, correlates with downregulation of FOXM1, and can be abrogated (11.8% vs. 95.3%, P < 0.001) by ectopic expression of FOXM1 in the corresponding gastric tumor cells. Cells with ectopic FOXM1 expression demonstrate considerable (∼20%, P < 0.001) growth advantage despite MET targeting, suggesting a novel clinically relevant resistance mechanism to MET inhibition as the copresence of both MET and FOXM1 protein (33%) and mRNA (30%) overexpression as well as gene amplification (24,7%) are common in patients with gastric cancer.

Conclusions: FOXM1, a negative regulator of senescence, has been identified as a key downstream effector and potential clinical biomarker that mediates MET signaling following infliction of DNA damage in gastric tumors. Clin Cancer Res; 22(21); 5322–36. ©2016 AACR.
Data from Gene Copy Number Estimation from Targeted Next-Generation Sequencing of Prostate Cancer Biopsies: Analytic Validation and Clinical Qualification
Authors: Seed George , Yuan Wei , Mateo Joaquin , Carreira Suzanne , Bertan Claudia , Lambros Maryou , Boysen Gunther , Ferraldeschi Roberta , Miranda Susana , Figueiredo Ines , Riisnaes Ruth , Crespo Mateus , Rodrigues Daniel Nava , Talevich Eric , Robinson Dan R. , Kunju Lakshmi P. , Wu Yi-Mi , Lonigro Robert , Sandhu Shahneen , Chinnaiyan Arul M. , de Bono Johann S.
Published: April 2023

Abstract
Purpose: Precise detection of copy number aberrations (CNA) from tumor biopsies is critically important to the treatment of metastatic prostate cancer. The use of targeted panel next-generation sequencing (NGS) is inexpensive, high throughput, and easily feasible, allowing single-nucleotide variant calls, but CNA estimation from this remains challenging.

Experimental Design: We evaluated CNVkit for CNA identification from amplicon-based targeted NGS in a cohort of 110 fresh castration-resistant prostate cancer biopsies and used capture-based whole-exome sequencing (WES), array comparative genomic hybridization (aCGH), and FISH to explore the viability of this approach.

Results: We showed that this method produced highly reproducible CNA results (r = 0.92), with the use of pooled germline DNA as a coverage reference supporting precise CNA estimation. CNA estimates from targeted NGS were comparable with WES (r = 0.86) and aCGH (r = 0.7); for key selected genes (BRCA2, MYC, PIK3CA, PTEN, and RB1), CNA estimation correlated well with WES (r = 0.91) and aCGH (r = 0.84) results. The frequency of CNAs in our population was comparable with that previously described (i.e., deep deletions: BRCA2 4.5%; RB1 8.2%; PTEN 15.5%; amplification: AR 45.5%; gain: MYC 31.8%). We also showed, utilizing FISH, that CNA estimation can be impacted by intratumor heterogeneity and demonstrated that tumor microdissection allows NGS to provide more precise CNA estimates.

Conclusions: Targeted NGS and CNVkit-based analyses provide a robust, precise, high-throughput, and cost-effective method for CNA estimation for the delivery of more precise patient care. Clin Cancer Res; 23(20); 6070–7. ©2017 AACR.
Data from Macrophages Facilitate Resistance to Anti-VEGF Therapy by Altered VEGFR Expression
Authors: Dalton Heather J. , Pradeep Sunila , McGuire Michael , Hailemichael Yared , MA Shaolin , Lyons Yasmin , Armaiz-Pena Guillermo N. , Previs Rebecca A. , Hansen Jean Marie , Rupaimoole Rajesha , Gonzalez-Villasana Vianey , Cho Min Soon , Wu Sherry Y. , Mangala Lingegowda S. , Jennings Nicholas B. , Hu Wei , Langley Robert , Mu Hong , Andreeff Michael , Bar-Eli Menashe , Overwijk Willem , Ram Prahlad , Lopez-Berestein Gabriel , Coleman Robert L. , Sood Anil K.
Published: April 2023

Abstract
Purpose: VEGF-targeted therapies have modest efficacy in cancer patients, but acquired resistance is common. The mechanisms underlying such resistance are poorly understood.

Experimental Design: To evaluate the potential role of immune cells in the development of resistance to VEGF blockade, we first established a preclinical model of adaptive resistance to anti-VEGF therapy. Additional in vitro and in vivo studies were carried out to characterize the role of macrophages in such resistance.

Results: Using murine cancer models of adaptive resistance to anti-VEGF antibody (AVA), we found a previously unrecognized role of macrophages in such resistance. Macrophages were actively recruited to the tumor microenvironment and were responsible for the emergence of AVA resistance. Depletion of macrophages following emergence of resistance halted tumor growth and prolonged survival of tumor-bearing mice. In a macrophage-deficient mouse model, resistance to AVA failed to develop, but could be induced by injection of macrophages. Downregulation of macrophage VEGFR-1 and VEGFR-3 expression accompanied upregulation of alternative angiogenic pathways, facilitating escape from anti-VEGF therapy.

Conclusions: These findings provide a new understanding of the mechanisms underlying the modest efficacy of current antiangiogenesis therapies and identify new opportunities for combination approaches for ovarian and other cancers. Clin Cancer Res; 23(22); 7034–46. ©2017 AACR.
Data from Myeloma Cell Dynamics in Response to Treatment Supports a Model of Hierarchical Differentiation and Clonal Evolution
Authors: Tang Min , Zhao Rui , van de Velde Helgi , Tross Jennifer G. , Mitsiades Constantine , Viselli Suzanne , Neuwirth Rachel , Esseltine Dixie-Lee , Anderson Kenneth , Ghobrial Irene M. , San Miguel Jesús F. , Richardson Paul G. , Tomasson Michael H. , Michor Franziska
Published: April 2023

Abstract
Purpose: Since the pioneering work of Salmon and Durie, quantitative measures of tumor burden in multiple myeloma have been used to make clinical predictions and model tumor growth. However, such quantitative analyses have not yet been performed on large datasets from trials using modern chemotherapy regimens.

Experimental Design: We analyzed a large set of tumor response data from three randomized controlled trials of bortezomib-based chemotherapy regimens (total sample size n = 1,469 patients) to establish and validate a novel mathematical model of multiple myeloma cell dynamics.

Results: Treatment dynamics in newly diagnosed patients were most consistent with a model postulating two tumor cell subpopulations, "progenitor cells" and "differentiated cells." Differential treatment responses were observed with significant tumoricidal effects on differentiated cells and less clear effects on progenitor cells. We validated this model using a second trial of newly diagnosed patients and a third trial of refractory patients. When applying our model to data of relapsed patients, we found that a hybrid model incorporating both a differentiation hierarchy and clonal evolution best explains the response patterns.

Conclusions: The clinical data, together with mathematical modeling, suggest that bortezomib-based therapy exerts a selection pressure on myeloma cells that can shape the disease phenotype, thereby generating further inter-patient variability. This model may be a useful tool for improving our understanding of disease biology and the response to chemotherapy regimens. Clin Cancer Res; 22(16); 4206–14. ©2016 AACR.
Data from Histone Deacetylase 3 Inhibition Overcomes BIM Deletion Polymorphism–Mediated Osimertinib Resistance in EGFR-Mutant Lung Cancer
Authors: Tanimoto Azusa , Takeuchi Shinji , Arai Sachiko , Fukuda Koji , Yamada Tadaaki , Roca Xavier , Ong S. Tiong , Yano Seiji
Published: April 2023

Abstract
Purpose: The BIM deletion polymorphism is associated with apoptosis resistance to EGFR tyrosine kinase inhibitors (EGFR-TKI), such as gefitinib and erlotinib, in non–small cell lung cancer (NSCLC) harboring EGFR mutations. Here, we investigated whether the BIM deletion polymorphism contributes to resistance against osimertinib, a third-generation EGFR-TKI. In addition, we determined the efficacy of a histone deacetylase (HDAC) inhibitor, vorinostat, against this form of resistance and elucidated the underlying mechanism.

Experimental Design: We used EGFR-mutated NSCLC cell lines, which were either heterozygous or homozygous for the BIM deletion polymorphism, to evaluate the effect of osimertinib in vitro and in vivo. Protein expression was examined by Western blotting. Alternative splicing of BIM mRNA was analyzed by RT-PCR.

Results: EGFR-mutated NSCLC cell lines with the BIM deletion polymorphism exhibited apoptosis resistance to osimertinib in a polymorphism dosage–dependent manner, and this resistance was overcome by combined use with vorinostat. Experiments with homozygous BIM deletion–positive cells revealed that vorinostat affected the alternative splicing of BIM mRNA in the deletion allele, increased the expression of active BIM protein, and thereby induced apoptosis in osimertinib-treated cells. These effects were mediated predominantly by HDAC3 inhibition. In xenograft models, combined use of vorinostat with osimertinib could regress tumors in EGFR-mutated NSCLC cells homozygous for the BIM deletion polymorphism. Moreover, this combination could induce apoptosis even when tumor cells acquired EGFR-T790M mutations.

Conclusions: These findings indicate the importance of developing HDAC3-selective inhibitors, and their combined use with osimertinib, for treating EGFR-mutated lung cancers carrying the BIM deletion polymorphism. Clin Cancer Res; 23(12); 3139–49. ©2016 AACR.
Data from Immune Cytolytic Activity Stratifies Molecular Subsets of Human Pancreatic Cancer
Authors: Balli David , Rech Andrew J. , Stanger Ben Z. , Vonderheide Robert H.
Published: April 2023

Abstract
Purpose: Immunotherapy has the potential to improve the dismal prognosis in pancreatic ductal adenocarcinoma (PDA), but clinical trials, including those with single-agent PD-1 or PD-L1 inhibition, have been disappointing. Our aim was to examine the immune landscape of PDA as it relates to aspects of tumor biology, including neoepitope burden.

Experimental Design: We used publicly available expression data from 134 primary resection PDA samples from The Cancer Genome Atlas to stratify patients according to a cytolytic T-cell activity expression index. We correlated cytolytic immune activity with mutational, structural, and neoepitope features of the tumor.

Results: Human PDA displays a range of intratumoral cytolytic T-cell activity. PDA tumors with low cytolytic activity exhibited significantly increased copy number alterations, including recurrent amplifications of MYC and NOTCH2 and recurrent deletions and mutations of CDKN2A/B. In sharp contrast to other tumor types, high cytolytic activity in PDA did not correlate with increased mutational burden or neoepitope load (MHC class I and class II). Cytolytic-high tumors exhibited increased expression of multiple immune checkpoint genes compared to cytolytic-low tumors, except for PD-L1 expression, which was uniformly low.

Conclusions: These data identify a subset of human PDA with high cytolytic T-cell activity. Rather than being linked to mutation burden or neoepitope load, immune activation indices in PDA were inversely linked to genomic alterations, suggesting that intrinsic oncogenic processes drive immune inactivity in human PDA. Furthermore, these data highlight the potential importance of immune checkpoints other than PD-L1/PD-1 as therapeutic targets in this lethal disease. Clin Cancer Res; 23(12); 3129–38. ©2016 AACR.
Data from Genomic Heterogeneity and Exceptional Response to Dual Pathway Inhibition in Anaplastic Thyroid Cancer
Authors: Gibson William J. , Ruan Daniel T. , Paulson Vera A. , Barletta Justine A. , Hanna Glenn J. , Kraft Stefan , Calles Antonio , Nehs Matthew A. , Moore Francis D. , Taylor-Weiner Amaro , Wala Jeremiah A. , Zack Travis I. , Lee Thomas C. , Fennessy Fiona M. , Alexander Erik K. , Thomas Tom , Janne Pasi A. , Garraway Levi A. , Carter Scott L. , Beroukhim Rameen , Lorch Jochen H. , Van Allen Eliezer M.
Published: April 2023

Abstract
Purpose: Cancers may resist single-agent targeted therapies when the flux of cellular growth signals is shifted from one pathway to another. Blockade of multiple pathways may be necessary for effective inhibition of tumor growth. We document a case in which a patient with anaplastic thyroid carcinoma (ATC) failed to respond to either mTOR/PI3K or combined RAF/MEK inhibition but experienced a dramatic response when both drug regimens were combined.

Experimental Design: Multi-region whole-exome sequencing of five diagnostic and four autopsy tumor biopsies was performed. Meta-analysis of DNA and RNA sequencing studies of ATC was performed.

Results: Sequencing revealed truncal BRAF and PIK3CA mutations, which are known to activate the MAPK and PI3K/AKT pathways, respectively. Meta-analysis demonstrated 10.3% cooccurrence of MAPK and PI3K pathway alterations in ATC. These tumors display a separate transcriptional profile from other ATCs, consistent with a novel subgroup of ATC.

Conclusions: BRAF and PIK3CA mutations define a distinct subset of ATC. Blockade of the MAPK and PI3K pathways appears necessary for tumor response in this subset of ATC. This identification of synergistic activity between targeted agents may inform clinical trial design in ATC. Clin Cancer Res; 23(9); 2367–73. ©2016 AACR.
Data from Cell-Cycle and DNA-Damage Response Pathway Is Involved in Leptomeningeal Metastasis of Non–Small Cell Lung Cancer
Authors: Fan Yun , Zhu Xuehua , Xu Yan , Lu Xuesong , Xu Yanjun , Wang Mengzhao , Xu Haiyan , Ding Jingyan , Ye Xin , Fang Luo , Huang Zhiyu , Gong Lei , Lu Hongyang , Mao Weimin , Hu Min
Published: April 2023

Abstract
Purpose: Leptomeningeal metastasis (LM) is a detrimental complication of non–small cell lung cancer (NSCLC) and associated with poor prognosis. However, the underlying mechanisms of the metastasis process are still poorly understood.

Experimental Design: We performed next-generation panel sequencing of primary tumor tissue, cerebrospinal fluid (CSF), and matched normal controls from epidermal growth factor receptor (EGFR) mutation-positive NSCLC patients with LM.

Results: The status of EGFR-activating mutations was highly concordant between primary tumor and CSF. PIK3CA aberrations were high in these patients, implicating an association with LM risk. Intriguingly, low overlapping of somatic protein-changing variants was observed between paired CSF and primary lesions, exhibiting tumor heterogeneity and genetic divergence. Moreover, genes with CSF-recurrent genomic alterations were predominantly involved in cell-cycle regulation and DNA-damage response (DDR), suggesting a role of the pathway in LM development.

Conclusions: Our study has shed light on the genomic variations of NSCLC-LM, demonstrated genetic heterogeneity and divergence, uncovered involvement of cell-cycle and DDR pathway, and paved the way for potential therapeutic approaches to this unmet medical need. Clin Cancer Res; 24(1); 209–16. ©2017 AACR.
Data from A PAM50-Based Chemoendocrine Score for Hormone Receptor–Positive Breast Cancer with an Intermediate Risk of Relapse
Authors: Prat Aleix , Lluch Ana , Turnbull Arran K. , Dunbier Anita K. , Calvo Lourdes , Albanell Joan , de la Haba-Rodríguez Juan , Arcusa Angels , Chacón José Ignacio , Sánchez-Rovira Pedro , Plazaola Arrate , Muñoz Montserrat , Paré Laia , Parker Joel S. , Ribelles Nuria , Jimenez Begoña , Bin Aiderus Abdul Aziz , Caballero Rosalía , Adamo Barbara , Dowsett Mitch , Carrasco Eva , Martín Miguel , Dixon J. Michael , Perou Charles M. , Alba Emilio
Published: April 2023

Abstract
Purpose: Hormone receptor–positive (HR+) breast cancer is clinically and biologically heterogeneous, and subgroups with different prognostic and treatment sensitivities need to be identified.

Experimental Design: Research-based PAM50 subtyping and expression of additional genes was performed on 63 patients with HR+/HER2− disease randomly assigned to neoadjuvant multiagent chemotherapy versus endocrine therapy in a phase II trial. The biology associated with treatment response was used to derive a PAM50-based chemoendocrine score (CES). CES's predictive ability was evaluated in 4 independent neoadjuvant data sets (n = 675) and 4 adjuvant data sets (n = 1,505). The association of CES, intrinsic biology, and PAM50 risk of relapse (ROR) was explored across 6,007 tumors.

Results: Most genes associated with endocrine sensitivity were also found associated with chemotherapy resistance. In the chemotherapy test/validation data sets, CES was independently associated with pathologic complete response (pCR), even after adjusting for intrinsic subtype. pCR rates of the CES endocrine–sensitive (CES-E), uncertain (CES-U), and chemotherapy-sensitive (CES-C) groups in both data sets combined were 25%, 11%, and 2%, respectively. In the endocrine test/validation data sets, CES was independently associated with response. Compared with ROR, >90% of ROR-low and ROR-high tumors were identified as CES-E and CES-C, respectively; however, each CES group represented >25% of ROR-intermediate disease. In terms of survival outcome, CES-C was associated with poor relapse-free survival in patients with ROR-intermediate disease treated with either adjuvant endocrine therapy only or no adjuvant systemic therapy, but not in patients treated with (neo)adjuvant chemotherapy.

Conclusions: CES is a genomic signature capable of estimating chemoendocrine sensitivity in HR+ breast cancer beyond intrinsic subtype and risk of relapse. Clin Cancer Res; 23(12); 3035–44. ©2016 AACR.
Data from A Prospective Evaluation of Early Detection Biomarkers for Ovarian Cancer in the European EPIC Cohort
Authors: Terry Kathryn L. , Schock Helena , Fortner Renée T. , Hüsing Anika , Fichorova Raina N. , Yamamoto Hidemi S. , Vitonis Allison F. , Johnson Theron , Overvad Kim , Tjønneland Anne , Boutron-Ruault Marie-Christine , Mesrine Sylvie , Severi Gianluca , Dossus Laure , Rinaldi Sabina , Boeing Heiner , Benetou Vassiliki , Lagiou Pagona , Trichopoulou Antonia , Krogh Vittorio , Kuhn Elisabetta , Panico Salvatore , Bueno-de-Mesquita H. Bas , Onland-Moret N. Charlotte , Peeters Petra H. , Gram Inger Torhild , Weiderpass Elisabete , Duell Eric J. , Sanchez Maria-Jose , Ardanaz Eva , Etxezarreta Nerea , Navarro Carmen , Idahl Annika , Lundin Eva , Jirström Karin , Manjer Jonas , Wareham Nicholas J. , Khaw Kay-Tee , Byrne Karl Smith , Travis Ruth C. , Gunter Marc J. , Merritt Melissa A. , Riboli Elio , Cramer Daniel W. , Kaaks Rudolf
Published: April 2023

Abstract
Purpose: About 60% of ovarian cancers are diagnosed at late stage, when 5-year survival is less than 30% in contrast to 90% for local disease. This has prompted search for early detection biomarkers. For initial testing, specimens taken months or years before ovarian cancer diagnosis are the best source of information to evaluate early detection biomarkers. Here we evaluate the most promising ovarian cancer screening biomarkers in prospectively collected samples from the European Prospective Investigation into Cancer and Nutrition study.

Experimental Design: We measured CA125, HE4, CA72.4, and CA15.3 in 810 invasive epithelial ovarian cancer cases and 1,939 controls. We calculated the sensitivity at 95% and 98% specificity as well as area under the receiver operator curve (C-statistic) for each marker individually and in combination. In addition, we evaluated marker performance by stage at diagnosis and time between blood draw and diagnosis.

Results: We observed the best discrimination between cases and controls within 6 months of diagnosis for CA125 (C-statistic = 0.92), then HE4 (0.84), CA72.4 (0.77), and CA15.3 (0.73). Marker performance declined with longer time between blood draw and diagnosis and for earlier staged disease. However, assessment of discriminatory ability at early stage was limited by small numbers. Combinations of markers performed modestly, but significantly better than any single marker.

Conclusions: CA125 remains the single best marker for the early detection of invasive epithelial ovarian cancer, but can be slightly improved by combining with other markers. Identifying novel markers for ovarian cancer will require studies including larger numbers of early-stage cases. Clin Cancer Res; 22(18); 4664–75. ©2016 AACR.

See related commentary by Skates, p. 4542
Data from 18F-fluoromisonidazole PET and Activity of Neoadjuvant Nintedanib in Early HER2-Negative Breast Cancer: A Window-of-Opportunity Randomized Trial
Authors: Quintela-Fandino Miguel , Lluch Ana , Manso Luis , Calvo Isabel , Cortes Javier , García-Saenz José Angel , Gil-Gil Miguel , Martinez-Jánez Noelia , Gonzalez-Martin Antonio , Adrover Encarna , de Andres Raquel , Viñas Gemma , Llombart-Cussac Antonio , Alba Emilio , Guerra Juan , Bermejo Begoña , Zamora Esther , Moreno-Anton Fernando , Pernas Simon Sonia , Carrato Alfredo , Lopez-Alonso Antonio , Escudero María José , Campo Ruth , Carrasco Eva , Palacios José , Mulero Francisca , Colomer Ramon
Published: April 2023

Abstract
Purpose: We previously detected promising efficacy of neoadjuvant nintedanib (a multityrosine kinase inhibitor, TKI) in early HER2-negative breast cancer. In a preclinical study, we monitored stromal hypoxia with 18F-fluoromisonidazole-positron emission tomography (18F-FMISO-PET); we found that reoxygenation of tumors (or lack of it) during a window-of-opportunity (WoO) treatment with TKIs correlated with the benefit (or lack of it) from TKI-plus-chemotherapy combinations. We studied the predictive role of 18F-FMISO-PET for the TKI nintedanib in the neoadjuvant setting in a phase II WoO randomized trial.

Experimental Design: Patients were randomized to a 14-day WoO of nintedanib preceded and followed by an 18F-FMISO-PET, followed by nintedanib plus weekly paclitaxel (Arm A) or an 18F-FMISO-PET followed by weekly paclitaxel (Arm B) before surgery. The endpoint was residual cancer burden (RCB). The objective was to detect the patients with no response (RCB-III) on the basis of the baseline or evolutive 18F-FMISO-PET values/changes.

Results: One-hundred and thirty HER2-negative patients were randomized. Seventeen (27.9%), 34 (55.7%), and 8 (13.1%) patients had an RCB of III, II, and I/0, respectively, in Arm A. In this arm, baseline hypoxic tumors had a 4.4-fold higher chance of experiencing RCB = 3 (P = 0.036) compared with baseline normoxic tumors. Nintedanib WoO induced tumor reoxygenation in 24.5% of the patients; those not reoxygenating showed a trend toward higher chance of experiencing RCB-III (6.4-fold; P = 0.09). In Arm B, 18F-FMISO-PET lacked predictive/prognostic value.

Conclusions: Baseline hypoxic tumors (measured with 18F-FMISO-PET) do not benefit from neoadjuvant nintedanib. Clin Cancer Res; 23(6); 1432–41. ©2016 AACR.
Data from Detection of Activating Estrogen Receptor Gene (ESR1) Mutations in Single Circulating Tumor Cells
Authors: Paolillo Carmela , Mu Zhaomei , Rossi Giovanna , Schiewer Matthew J. , Nguyen Thomas , Austin Laura , Capoluongo Ettore , Knudsen Karen , Cristofanilli Massimo , Fortina Paolo
Published: April 2023

Abstract
Purpose: Early detection is essential for treatment plans before onset of metastatic disease. Our purpose was to demonstrate feasibility to detect and monitor estrogen receptor 1 (ESR1) gene mutations at the single circulating tumor cell (CTC) level in metastatic breast cancer (MBC).

Experimental Design: We used a CTC molecular characterization approach to investigate heterogeneity of 14 hotspot mutations in ESR1 and their correlation with endocrine resistance. Combining the CellSearch and DEPArray technologies allowed recovery of 71 single CTCs and 12 WBC from 3 ER-positive MBC patients. Forty CTCs and 12 WBC were subjected to whole genome amplification by MALBAC and Sanger sequencing.

Results: Among 3 selected patients, 2 had an ESR1 mutation (Y537). One showed two different ESR1 variants in a single CTC and another showed loss of heterozygosity. All mutations were detected in matched cell-free DNA (cfDNA). Furthermore, one had 2 serial blood samples analyzed and showed changes in both cfDNA and CTCs with emergence of mutations in ESR1 (Y537S and T570I), which has not been reported previously.

Conclusions: CTCs are easily accessible biomarkers to monitor and better personalize management of patients with previously demonstrated ER-MBC who are progressing on endocrine therapy. We showed that single CTC analysis can yield important information on clonal heterogeneity and can be a source of discovery of novel and potential driver mutations. Finally, we also validate a workflow for liquid biopsy that will facilitate early detection of ESR1 mutations, the emergence of endocrine resistance and the choice of further target therapy. Clin Cancer Res; 23(20); 6086–93. ©2017 AACR.
Data from PIK3CA Mutations Contribute to Acquired Cetuximab Resistance in Patients with Metastatic Colorectal Cancer
Authors: Xu Jian-Ming , Wang Yan , Wang You-Liang , Liu Tao , Ni Ming , Li Man-Sheng , Lin Li , Ge Fei-Jiao , Gong Chun , Gu Jun-Yan , Jia Ru , Wang He-Fei , Chen Yu-Ling , Liu Rong-Rui , Zhao Chuan-Hua , Tan Zhao-Li , Jin Yang , Zhu Yun-Ping , Ogino Shuji , Qian Zhi-Rong
Published: April 2023

Abstract
Purpose: Mutations in KRAS are considered to be the main drivers of acquired resistance to epidermal growth factor receptor (EGFR) blockade in patients with metastatic colorectal cancer (mCRC). However, the potential role of other genes downstream of the EGFR signaling pathway in conferring acquired resistance has not been extensively investigated.

Experimental Design: Using circulating tumor DNA (ctDNA) from patients with mCRC and with acquired cetuximab resistance, we developed a targeted amplicon ultra-deep sequencing method to screen for low-abundance somatic mutations in a panel of genes that encode components of the EGFR signaling pathway. Mutations with significantly increased variant frequencies upon disease progression were selected by using quartile analysis. The functional consequences of the identified mutations were validated in cultured cells.

Results: We analyzed 32 patients with acquired cetuximab resistance in a development cohort. Of them, seven (22%) carried five novel PIK3CA mutations, whereas eight (25%) carried previously reported KRAS mutations. Functional studies showed that novel PIK3CA mutations (all in exon 19; p.K944N, p.F930S, p.V955G, p.V955I, and p.K966E) promote cell viability in the presence of cetuximab. Only one novel PIK3CA mutation (p.K944N) was verified in one of the 27 patients with acquired resistance in a validation cohort, simultaneous KRAS and PIK3CA hotspot mutations were detected in two patients. Among the above 59 acquired resistance patients, those with PIK3CA or RAS mutations detected in ctDNA showed a pronounced decrease in progression-free survival than patients with no mutation.

Conclusions: The PIK3CA mutations may potentially contribute to acquired cetuximab resistance in patients with mCRC. Clin Cancer Res; 23(16); 4602–16. ©2017 AACR.
Data from Hypoxia-Inducible PIM Kinase Expression Promotes Resistance to Antiangiogenic Agents
Authors: Casillas Andrea L. , Toth Rachel K. , Sainz Alva G. , Singh Neha , Desai Ankit A. , Kraft Andrew S. , Warfel Noel A.
Published: April 2023

Abstract
Purpose: Patients develop resistance to antiangiogenic drugs, secondary to changes in the tumor microenvironment, including hypoxia. PIM kinases are prosurvival kinases and their expression increases in hypoxia. The goal of this study was to determine whether targeting hypoxia-induced PIM kinase expression is effective in combination with VEGF-targeting agents. The rationale for this therapeutic approach is based on the fact that antiangiogenic drugs can make tumors hypoxic, and thus more sensitive to PIM inhibitors.

Experimental Design: Xenograft and orthotopic models of prostate and colon cancer were used to assess the effect of PIM activation on the efficacy of VEGF-targeting agents. IHC and in vivo imaging were used to analyze angiogenesis, apoptosis, proliferation, and metastasis. Biochemical studies were performed to characterize the novel signaling pathway linking PIM and HIF1.

Results: PIM was upregulated following treatment with anti-VEGF therapies, and PIM1 overexpression reduced the ability of these drugs to disrupt vasculature and block tumor growth. PIM inhibitors reduced HIF1 activity, opposing the shift to a pro-angiogenic gene signature associated with hypoxia. Combined inhibition of PIM and VEGF produced a synergistic antitumor response characterized by decreased proliferation, reduced tumor vasculature, and decreased metastasis.

Conclusions: This study describes PIM kinase expression as a novel mechanism of resistance to antiangiogenic agents. Our data provide justification for combining PIM and VEGF inhibitors to treat solid tumors. The unique ability of PIM inhibitors to concomitantly target HIF1 and selectively kill hypoxic tumor cells addresses two major components of tumor progression and therapeutic resistance. Clin Cancer Res; 24(1); 169–80. ©2017 AACR.
Data from Association of Tissue Abiraterone Levels and SLCO Genotype with Intraprostatic Steroids and Pathologic Response in Men with High-Risk Localized Prostate Cancer
Authors: Mostaghel Elahe A. , Cho Eunpi , Zhang Ailin , Alyamani Mohammad , Kaipainen Arja , Green Sean , Marck Brett T. , Sharifi Nima , Wright Jonathan L. , Gulati Roman , True Lawrence D. , Loda Massimo , Matsumoto Alvin M. , Tamae Daniel , Penning Trevor N. , Balk Steven P. , Kantoff Phillip W. , Nelson Peter S. , Taplin Mary-Ellen , Montgomery R. Bruce
Published: April 2023

Abstract
Purpose: Germline variation in solute carrier organic anion (SLCO) genes influences cellular steroid uptake and is associated with prostate cancer outcomes. We hypothesized that, due to its steroidal structure, the CYP17A inhibitor abiraterone may undergo transport by SLCO-encoded transporters and that SLCO gene variation may influence intracellular abiraterone levels and outcomes.

Experimental Design: Steroid and abiraterone levels were measured in serum and tissue from 58 men with localized prostate cancer in a clinical trial of LHRH agonist plus abiraterone acetate plus prednisone for 24 weeks prior to prostatectomy. Germline DNA was genotyped for 13 SNPs in six SLCO genes.

Results: Abiraterone levels spanned a broad range (serum median 28 ng/mL, 108 nmol/L; tissue median 77 ng/mL, 271 nmol/L) and were correlated (r = 0.355, P = 0.001). Levels correlated positively with steroids upstream of CYP17A (pregnenolone, progesterone), and inversely with steroids downstream of CYP17A (DHEA, AED, testosterone). Serum PSA and tumor volumes were higher in men with undetectable versus detectable tissue abiraterone at prostatectomy (median 0.10 vs. 0.03 ng/dL, P = 0.02; 1.28 vs. 0.44 cc, P = 0.09, respectively). SNPs in SLCO2B1 associated with significant differences in tissue abiraterone (rs1789693, P = 0.0008; rs12422149, P = 0.03) and higher rates of minimal residual disease (tumor volume < 0.5 cc; rs1789693, 67% vs. 27%, P = 0.009; rs1077858, 46% vs. 0%, P = 0.03). LNCaP cells expressing SLCO2B1 showed two- to fourfold higher abiraterone levels compared with vector controls (P < 0.05).

Conclusions: Intraprostatic abiraterone levels and genetic variation in SLCO genes are associated with pathologic responses in high-risk localized prostate cancer. Variation in SLCO genes may serve as predictors of response to abiraterone treatment. Clin Cancer Res; 23(16); 4592–601. ©2017 AACR.
Data from Anti-EGFR Targeted Monoclonal Antibody Isotype Influences Antitumor Cellular Immunity in Head and Neck Cancer Patients
Authors: Trivedi Sumita , Srivastava Raghvendra M. , Concha-Benavente Fernando , Ferrone Soldano , Garcia-Bates Tatiana M. , Li Jing , Ferris Robert L.
Published: April 2023

Abstract
Purpose: EGF receptor (EGFR) is highly overexpressed on several cancers and two targeted anti-EGFR antibodies which differ by isotype are FDA-approved for clinical use. Cetuximab (IgG1 isotype) inhibits downstream signaling of EGFR and activates antitumor, cellular immune mechanisms. As panitumumab (IgG2 isotype) may inhibit downstream EGFR signaling similar to cetuximab, it might also induce adaptive immunity.

Experimental Design: We measured in vitro activation of cellular components of the innate and adaptive immune systems. We also studied the in vivo activation of components of the adaptive immune system in patient specimens from two recent clinical trials using cetuximab or panitumumab.

Results: Both monoclonal antibodies (mAb) primarily activate natural killer (NK) cells, although cetuximab is significantly more potent than panitumumab. Cetuximab-activated neutrophils mediate antibody-dependent cellular cytotoxicity (ADCC) against head and neck squamous cell carcinomas (HNSCC) tumor cells, and interestingly, this effect was FcγRIIa- and FcγRIIIa genotype–dependent. Panitumumab may activate monocytes through CD32 (FcγRIIa); however, monocytes activated by either mAb are not able to mediate ADCC. Cetuximab enhanced dendritic cell (DC) maturation to a greater extent than panitumumab, which was associated with improved tumor antigen cross-presentation by cetuximab compared with panitumumab. This correlated with increased EGFR-specific cytotoxic CD8+ T cells in patients treated with cetuximab compared with those treated with panitumumab.

Conclusions: Although panitumumab effectively inhibits EGFR signaling to a similar extent as cetuximab, it is less effective at triggering antitumor, cellular immune mechanisms which may be crucial for effective therapy of HNSCC. Clin Cancer Res; 22(21); 5229–37. ©2016 AACR.
Data from T2–FLAIR Mismatch, an Imaging Biomarker for IDH and 1p/19q Status in Lower-grade Gliomas: A TCGA/TCIA Project
Authors: Patel Sohil H. , Poisson Laila M. , Brat Daniel J. , Zhou Yueren , Cooper Lee , Snuderl Matija , Thomas Cheddhi , Franceschi Ana M. , Griffith Brent , Flanders Adam E. , Golfinos John G. , Chi Andrew S. , Jain Rajan
Published: April 2023

Abstract
Purpose: Lower-grade gliomas (WHO grade II/III) have been classified into clinically relevant molecular subtypes based on IDH and 1p/19q mutation status. The purpose was to investigate whether T2/FLAIR MRI features could distinguish between lower-grade glioma molecular subtypes.

Experimental Design: MRI scans from the TCGA/TCIA lower grade glioma database (n = 125) were evaluated by two independent neuroradiologists to assess (i) presence/absence of homogenous signal on T2WI; (ii) presence/absence of “T2–FLAIR mismatch” sign; (iii) sharp or indistinct lesion margins; and (iv) presence/absence of peritumoral edema. Metrics with moderate–substantial agreement underwent consensus review and were correlated with glioma molecular subtypes. Somatic mutation, DNA copy number, DNA methylation, gene expression, and protein array data from the TCGA lower-grade glioma database were analyzed for molecular–radiographic associations. A separate institutional cohort (n = 82) was analyzed to validate the T2–FLAIR mismatch sign.

Results: Among TCGA/TCIA cases, interreader agreement was calculated for lesion homogeneity [κ = 0.234 (0.111–0.358)], T2–FLAIR mismatch sign [κ = 0.728 (0.538–0.918)], lesion margins [κ = 0.292 (0.135–0.449)], and peritumoral edema [κ = 0.173 (0.096–0.250)]. All 15 cases that were positive for the T2–FLAIR mismatch sign were IDH-mutant, 1p/19q non-codeleted tumors (P < 0.0001; PPV = 100%, NPV = 54%). Analysis of the validation cohort demonstrated substantial interreader agreement for the T2–FLAIR mismatch sign [κ = 0.747 (0.536–0.958)]; all 10 cases positive for the T2–FLAIR mismatch sign were IDH-mutant, 1p/19q non-codeleted tumors (P < 0.00001; PPV = 100%, NPV = 76%).

Conclusions: Among lower-grade gliomas, T2–FLAIR mismatch sign represents a highly specific imaging biomarker for the IDH-mutant, 1p/19q non-codeleted molecular subtype. Clin Cancer Res; 23(20); 6078–85. ©2017 AACR.
Data from Downregulation of SAFB Sustains the NF-κB Pathway by Targeting TAK1 during the Progression of Colorectal Cancer
Authors: Jiao Hong-Li , Ye Ya-Ping , Yang Run-Wei , Sun Hui-Ying , Wang Shu-Yang , Wang Yong-Xia , Xiao Zhi-Yuan , He Liu-Qing , Cai Juan-Juan , Wei Wen-Ting , Chen Yan-Ru , Gu Chun-Cai , Cai Yue-Long , Hu Yun-Teng , Lai Qiu-Hua , Qiu Jun-Feng , Liang Li , Cao Guang-Wen , Liao Wen-Ting , Ding Yan-Qing
Published: April 2023

Abstract
Purpose: To investigate the role and the underlying mechanism of scaffold attachment factor B (SAFB) in the progression of colorectal cancer (CRC).

Experimental Design: SAFB expression was analyzed in the Cancer Outlier Profile Analysis of Oncomine and in 175 paraffin-embedded archived CRC tissues. Gene Ontology analyses were performed to explore the mechanism of SAFB in CRC progression. Western blot, RT-PCR, luciferase assay, and chromatin immunoprecipitation (ChIP) were used to detect the regulation of transforming growth factor-β–activated kinase 1 (TAK1) and NF-κB signaling by SAFB. The role of SAFB in invasion, metastasis, and angiogenesis was investigated using in vitro and in vivo assays. The relationship between SAFB and TAK1 was analyzed in CRC tissues.

Results: SAFB was downregulated in CRC tissues, and low expression of SAFB was significantly associated with an aggressive phenotype and poorer survival of CRC patients. The downregulation of SAFB activated NF-κB signaling by targeting the TAK1 promoter. Ectopic expression of SAFB inhibited the development of aggressive features and metastasis of CRC cells both in vitro and in vivo. The overexpression of TAK1 could rescue the aggressive features in SAFB-overexpressed cells. Furthermore, the expression of SAFB in CRC tissues was negatively correlated with the expression of TAK1- and NF-κB–related genes.

Conclusions: Our results show that SAFB regulated the activity of NF-κB signaling in CRC by targeting TAK1. This novel mechanism provides a comprehensive understanding of both SAFB and the NF-κB signaling pathway in the progression of CRC and indicates that the SAFB–TAK1–NF-κB axis is a potential target for early therapeutic intervention in CRC progression. Clin Cancer Res; 23(22); 7108–18. ©2017 AACR.
Data from Intra- and Interobserver Reproducibility Assessment of PD-L1 Biomarker in Non–Small Cell Lung Cancer
Authors: Cooper Wendy A. , Russell Prudence A. , Cherian Maya , Duhig Edwina E. , Godbolt David , Jessup Peter J. , Khoo Christine , Leslie Connull , Mahar Annabelle , Moffat David F. , Sivasubramaniam Vanathi , Faure Celine , Reznichenko Alena , Grattan Amanda , Fox Stephen B.
Published: April 2023

Abstract
Purpose: Reliable and reproducible methods for identifying PD-L1 expression on tumor cells are necessary to identify responders to anti–PD-1 therapy. We tested the reproducibility of the assessment of PD-L1 expression in non–small cell lung cancer (NSCLC) tissue samples by pathologists.

Experimental Design: NSCLC samples were stained with PD-L1 22C3 pharmDx kit using the Dako Autostainer Link 48 Platform. Two sample sets of 60 samples each were designed to assess inter- and intraobserver reproducibility considering two cut points for positivity: 1% or 50% of PD-L1 stained tumor cells. A randomization process was used to obtain equal distribution of PD-L1 positive and negative samples within each sample set. Ten pathologists were randomly assigned to two subgroups. Subgroup 1 analyzed all samples on two consecutive days. Subgroup 2 performed the same assessments, except they received a 1-hour training session prior to the second assessment.

Results: For intraobserver reproducibility, the overall percent agreement (OPA) was 89.7% [95% confidence interval (CI), 85.7–92.6] for the 1% cut point and 91.3% (95% CI, 87.6–94.0) for the 50% cut point. For interobserver reproducibility, OPA was 84.2% (95% CI, 82.8–85.5) for the 1% cut point and 81.9% (95% CI, 80.4–83.3) for the 50% cut point, and Cohen's κ coefficients were 0.68 (95% CI, 0.65–0.71) and 0.58 (95% CI, 0.55–0.62), respectively. The training was found to have no or very little impact on intra- or interobserver reproducibility.

Conclusions: Pathologists reported good reproducibility at both 1% and 50% cut points. More adapted training could potentially increase reliability, in particular for samples with PD-L1 proportion, scores around 50%. Clin Cancer Res; 23(16); 4569–77. ©2017 AACR.
Data from Modulation of Navitoclax Sensitivity by Dihydroartemisinin-Mediated MCL-1 Repression in BCR-ABL+ B-Lineage Acute Lymphoblastic Leukemia
Authors: Budhraja Amit , Turnis Meghan E. , Churchman Michelle L. , Kothari Anisha , Yang Xue , Xu Haiyan , Kaminska Ewa , Panetta John C. , Finkelstein David , Mullighan Charles G. , Opferman Joseph T.
Published: April 2023

Abstract
Purpose: BCR-ABL+ B-ALL leukemic cells are highly dependent on the expression of endogenous antiapoptotic MCL-1 to promote viability and are resistant to BH3-mimetic agents such as navitoclax (ABT-263) that target BCL-2, BCL-XL, and BCL-W. However, the survival of most normal blood cells and other cell types is also dependent on Mcl-1. Despite the requirement for MCL-1 in these cell types, initial reports of MCL-1–specific BH3-mimetics have not described any overt toxicities associated with single-agent use, but these agents are still early in clinical development. Therefore, we sought to identify approved drugs that could sensitize leukemic cells to ABT-263.

Experimental Design: A screen identified dihydroartemisinin (DHA), a water-soluble metabolite of the antimalarial artemisinin. Using mouse and human leukemic cell lines, and primary patient-derived xenografts, the effect of DHA on survival was tested, and mechanistic studies were carried out to discover how DHA functions. We further tested in vitro and in vivo whether combining DHA with ABT-263 could enhance the response of leukemic cells to combination therapy.

Results: DHA causes the downmodulation of MCL-1 expression by triggering a cellular stress response that represses translation. The repression of MCL-1 renders leukemic cells highly sensitive to synergistic cell death induced by ABT-263 in a mouse model of BCR-ABL+ B-ALL both in vitro and in vivo. Furthermore, DHA synergizes with ABT-263 in human Ph+ ALL cell lines, and primary patient-derived xenografts of Ph+ ALL in culture.

Conclusions: Our findings suggest that combining DHA with ABT-263 can improve therapeutic response in BCR-ABL+ B-ALL. Clin Cancer Res; 23(24); 7558–68. ©2017 AACR.
Data from Histone Deacetylase Inhibitor Enhances the Efficacy of MEK Inhibitor through NOXA-Mediated MCL1 Degradation in Triple-Negative and Inflammatory Breast Cancer
Authors: Torres-Adorno Angie M. , Lee Jangsoon , Kogawa Takahiro , Ordentlich Peter , Tripathy Debu , Lim Bora , Ueno Naoto T.
Published: April 2023

Abstract
Purpose: Inflammatory breast cancer (IBC), diagnosed clinically, and triple-negative breast cancer (TNBC), diagnosed by molecular receptor status, are the two most aggressive forms of breast cancer, and both lack effective targeted therapies. We previously demonstrated involvement of histone deacetylase (HDAC) inhibitor entinostat in regulating apoptosis in IBC and TNBC cells; here, we aimed to identify novel combination therapy candidates.

Experimental Design: Potential therapeutic targets were identified by mRNA expression profiling of TNBC and IBC cells treated with entinostat. Drug action and synergism were assessed by in vitro proliferation assays, tumor growth in vivo, and proteomic analyses. Gain/loss-of-expression studies were utilized to functionally validate the role of identified targets in sensitivity of TNBC and IBC cells to combination therapy.

Results: Entinostat induced activity of the oncogenic ERK pathway and expression of proapoptotic NOXA. These are known to stabilize and degrade, respectively, MCL1, an antiapoptotic Bcl-2 protein. In breast cancer patients, high-MCL1/low-NOXA tumor expression correlated significantly with poor survival outcomes. Combination treatment of entinostat with MEK inhibitor pimasertib reduced the growth of TNBC and IBC cells in vitro and inhibited tumor growth in vivo. The synergistic action of combination therapy was observed in TNBC and IBC cell lines in which NOXA expression was induced following entinostat treatment. The therapeutic activity depended on induction of mitochondrial cell death pathways initiated by NOXA-mediated MCL1 degradation.

Conclusions: Our preclinical findings provide a rationale for the clinical testing of combination HDAC and MEK pathway inhibition for TNBC and IBC that exhibit elevated baseline tumor MCL1 expression. Clin Cancer Res; 23(16); 4780–92. ©2017 AACR.