bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Evaluation of Colon Cancer–Specific Antigen 2 as a Potential Serum Marker for Colorectal Cancer
Authors: Leman Eddy S. , Schoen Robert E. , Magheli Ahmed , Sokoll Lori J. , Chan Daniel W. , Getzenberg Robert H.
Published: April 2023

Abstract
Purpose: A blood test to detect colon cancer at a preventable stage would represent a major advancement. We have previously identified colon cancer–specific markers using focused proteomics analysis of nuclear structural proteins. Two of these markers, colon cancer–specific antigen (CCSA)-3 and CCSA-4, have been developed into blood-based markers that are able to distinguish individuals with colorectal cancer from those without. CCSA-2 is a distinct novel colon cancer marker identified using focused proteomics.

Experimental Design: Using an indirect ELISA on serum samples obtained from two institutions, we evaluated CCSA-2 as a serum-based colon cancer marker. A total of 111 serum samples from individuals who underwent colonoscopy and were subsequently diagnosed as either being normal or having hyperplastic polyps, nonadvanced adenomas, advanced adenomas, and colorectal cancer were evaluated. A diverse control population that consisted of 125 serum samples was also included in this study.

Results: Receiver operating characteristic analyses were used to measure the sensitivity and specificity of CCSA-2. CCSA-2 at a cutoff of 10.8 μg/mL has overall specificity of 78.4% [95% confidence interval (95% CI), 67.3-87.1%] and sensitivity of 97.3% (95% CI, 85.8-99.5%) in separating individuals with advanced adenomas and colorectal cancer from normal, hyperplastic, and nonadvanced adenoma populations. The receiver operating characteristic curve for CCSA-2 has an area under the curve of 0.90 (95% CI, 0.83-0.95).

Conclusion: Our initial study shows that CCSA-2 is a potential serum-based marker for colon cancer detection with high sensitivity and specificity.
Data from PIK3CA Mutations May Be Discordant between Primary and Corresponding Metastatic Disease in Breast Cancer
Authors: Dupont Jensen Jeanette , Laenkholm Anne-Vibeke , Knoop Ann , Ewertz Marianne , Bandaru Raj , Liu Weihua , Hackl Wolfgang , Barrett J. Carl , Gardner Humphrey
Published: April 2023

Abstract
Purpose:PIK3CA mutations are frequent in breast cancer and activate the PI3K/Akt pathway. Unexpectedly, PIK3CA mutation appears in general to be associated with better outcome. In a cohort of patients where both primary and metastatic lesions were available, the objective was to assess changes in PIK3CA mutations. We wished to discern whether selective pressures occur and the influence of PIK3CA mutation on time to recurrence.

Experimental Design: Formalin-fixed paraffin-embedded tumor blocks were obtained from 104 patients with paired samples from primary tumors and corresponding asynchronous metastatic breast tumors. Samples were analyzed for PIK3CA mutations (exons 9 and 20) as well as immunohistochemical evaluation for PTEN, pAKT, Ki67, ER, and HER2.

Results:PIK3CA mutation was detected in 45% of the primary tumors. Overall, there was a net gain in mutation in metastatic disease, to 53%; nonetheless, there were instances where metastases were wild type in patients with PIK3CA mutant primary tumors. Laser capture microdissection on a subset of cases revealed microheterogeneity for PIK3CA mutational status in the primary tumor. PIK3CA mutants overall showed a significantly longer time to first recurrence than wild type cases (P = 0.03).

Conclusion:PIK3CA mutations occur at high frequency in primary and metastatic breast cancer; these may not necessarily confer increased aggressiveness as mutants had a longer time to recurrence. Because PIK3CA status quite frequently changes between primary and metastatic disease, it emphasizes the necessity of assessing the PIK3CA status in the metastatic lesion for selection of PIK3CA inhibitor therapy. Clin Cancer Res; 17(4); 667–77. ©2010 AACR.
Data from MicroRNAs Are Independent Predictors of Outcome in Diffuse Large B-Cell Lymphoma Patients Treated with R-CHOP
Authors: Alencar Alvaro J. , Malumbres Raquel , Kozloski Goldi A. , Advani Ranjana , Talreja Neha , Chinichian Shideh , Briones Javier , Natkunam Yasodha , Sehn Laurie H. , Gascoyne Randy D. , Tibshirani Rob , Lossos Izidore S.
Published: April 2023

Abstract
Purpose: Diffuse large B-cell lymphoma (DLBCL) heterogeneity has prompted investigations for new biomarkers that can accurately predict survival. A previously reported 6-gene model combined with the International Prognostic Index (IPI) could predict patients' outcome. However, even these predictors are not capable of unambiguously identifying outcome, suggesting that additional biomarkers might improve their predictive power.

Experimental Design: We studied expression of 11 microRNAs (miRNA) that had previously been reported to have variable expression in DLBCL tumors. We measured the expression of each miRNA by quantitative real-time PCR analyses in 176 samples from uniformly treated DLBCL patients and correlated the results to survival.

Results: In a univariate analysis, the expression of miR-18a correlated with overall survival (OS), whereas the expression of miR-181a and miR-222 correlated with progression-free survival (PFS). A multivariate Cox regression analysis including the IPI, the 6-gene model–derived mortality predictor score and expression of the miR-18a, miR-181a, and miR-222, revealed that all variables were independent predictors of survival except the expression of miR-222 for OS and the expression of miR-18a for PFS.

Conclusion: The expression of specific miRNAs may be useful for DLBCL survival prediction and their role in the pathogenesis of this disease should be examined further. Clin Cancer Res; 17(12); 4125–35. ©2011 AACR.
Data from HDAC Inhibitors Potentiate the Activity of the BCR/ABL Kinase Inhibitor KW-2449 in Imatinib-Sensitive or -Resistant BCR/ABL+ Leukemia Cells In Vitro and In Vivo
Authors: Nguyen Tri , Dai Yun , Attkisson Elisa , Kramer Lora , Jordan Nicholas , Nguyen Nguyen , Kolluri Nikhil , Muschen Markus , Grant Steven
Published: April 2023

Abstract
Purpose: The purpose of this study was to determine whether histone deacetylase (HDAC) inhibitors (HDACI) such as vorinostat or entinostat (SNDX-275) could increase the lethality of the dual Bcr/Abl-Aurora kinase inhibitor KW-2449 in various Bcr/Abl+ human leukemia cells, including those resistant to imatinib mesylate (IM).

Experimental Design: Bcr/Abl+ chronic myelogenous leukemia (CML) and acute lymphoblastic leukemia (ALL) cells, including those resistant to IM (T315I, E255K), were exposed to KW-2449 in the presence or absence of vorinostat or SNDX-275, after which apoptosis and effects on signaling pathways were examined. In vivo studies combining HDACIs and KW2449 were carried out by using a systemic IM-resistant ALL xenograft model.

Results: Coadministration of HDACIs synergistically increased KW-2449 lethality in vitro in multiple CML and Ph+ ALL cell types including human IM resistant cells (e.g., BV-173/E255K and Adult/T315I). Combined treatment resulted in inactivation of Bcr/Abl and downstream targets (e.g., STAT5 and CRKL), as well as increased reactive oxygen species (ROS) generation and DNA damage (γH2A.X). The latter events and cell death were significantly attenuated by free radical scavengers (TBAP). Increased lethality was also observed in primary CD34+ cells from patients with CML, but not in normal CD34+ cells. Finally, minimally active vorinostat or SNDX275 doses markedly increased KW2449 antitumor effects and significantly prolonged the survival of murine xenografts bearing IM-resistant ALL cells (BV173/E255K).

Conclusions: HDACIs increase KW-2449 lethality in Bcr/Abl+ cells in association with inhibition of Bcr/Abl, generation of ROS, and induction of DNA damage. This strategy preferentially targets primary Bcr/Abl+ hematopoietic cells and exhibits enhanced in vivo activity. Combining KW-2449 with HDACIs warrants attention in IM-resistant Bcr/Abl+ leukemias. Clin Cancer Res; 17(10); 3219–32. ©2011 AACR.
Data from Clinical Significance of CD99 Down-Regulation in Gastric Adenocarcinoma
Authors: Lee Jung Hyun , Kim Seok-Hyung , Wang Li-Hui , Choi Yoon-La , Kim Young Chul , Kim Jin Hee , Park Tae Sung , Hong Yun-Chul , Shin Young Kee
Published: April 2023

Abstract
Purpose: CD99 is a cell adhesion molecule associated with human tumors. The aim of the present study was to characterize its role in the development and progression of human gastric adenocarcinoma.

Experimental Design: The expression of CD99 was investigated in 283 gastric adenocarcinomas and related lesions and 9 gastric carcinoma cell lines. We also analyzed the methylation status of CD99 gene by using methylation-specific PCR and examined loss of heterozygosity (LOH) of this gene locus by using an intragenic marker. Moreover, we assessed whether SP1, a positive transcription factor for CD99, is expressed in these samples.

Results: We found that the decreased expression of CD99 was strongly associated with poor survival and unfavorable clinicopathologic variables. Promoter region methylation (15 of 89, 16.9%) and LOH (21 of 74, 28.4%) were observed and significantly associated with CD99 down-regulation (P < 0.05). In addition, most of the gastric adenocarcinoma cases with CD99 down-regulation had reduced expression of SP1 (47 of 103, 45.6%; P < 0.01). This relationship between CD99 and SP1 was consolidated by using SP1 small interfering RNA transfection experiment and CD99 promoter luciferase assay. Furthermore, we showed that CD99 down-regulation was associated with proliferation and migration in gastric carcinoma cell line.

Conclusion: These observations suggest that CD99 down-regulation is a critical event in the progression of gastric adenocarcinoma, and CD99 promoter methylation, CD99 LOH, and SP1 down-regulation were responsible for the down-regulation of CD99.
Data from SMAD4 Levels and Response to 5-Fluorouracil in Colorectal Cancer
Authors: Alhopuro Pia , Alazzouzi Hafid , Sammalkorpi Heli , Dávalos Verónica , Salovaara Reijo , Hemminki Akseli , Järvinen Heikki , Mecklin Jukka-Pekka , Schwartz Simo , Aaltonen Lauri A. , Arango Diego
Published: April 2023

Abstract
We have recently reported that low tumor levels of SMAD4, a key mediator of transforming growth factor-β superfamily signaling, can predict the probability of recurrence in patients with Dukes C colorectal cancer who had surgery as the only form of treatment. However, standard treatment for Dukes C colorectal cancer patients currently involves the administration of 5-fluorouracil (5-FU)–based adjuvant chemotherapy after surgery. Approximately 30% to 40% of these patients present with recurrence and die within 5 years, and there is great need for markers capable of predicting poor prognosis after the combined surgery/adjuvant treatment. In this study, we evaluate the prognostic value of SMAD4 in patients treated with surgery and 5-FU-based adjuvant therapy. We used immunohistochemistry and quantitative real-time reverse transcription-PCR to measure the levels of SMAD4 protein and mRNA expression in the primary tumors and a number of lymph node metastases from a series of 75 Dukes C colorectal cancer patients with at least 6 years of follow-up. Patients with tumors expressing low levels of SMAD4 protein or mRNA showed significantly shorted disease-free and overall survival than patients with high tumor levels of SMAD4. The median survival of patients with low SMAD4 protein or mRNA tumor levels was 1.4 and 1.2 years, respectively, whereas patients with high SMAD4 tumor level had a median survival of >9.3 years. In addition, the protein and mRNA levels of SMAD4 in lymph node metastases was significantly lower than in primary tumors (P = 0.006). In contrast, allelic imbalance in chromosome 18q21 was of no prognostic significance in these patients. In conclusion, low SMAD4 tumor levels identified a subset of patients with poor prognosis following surgery and 5-FU-based adjuvant therapy; therefore, these patients could be good candidates to receive combined treatment with additional chemotherapeutic agents such as CPT-11 and/or oxaliplatin.
Data from ABCA3 as a Possible Cause of Drug Resistance in Childhood Acute Myeloid Leukemia
Authors: Steinbach Daniel , Gillet Jean-Pierre , Sauerbrey Axel , Gruhn Bernd , Dawczynski Kristin , Bertholet Vincent , de Longueville Françoise , Zintl Felix , Remacle Jose , Efferth Thomas
Published: April 2023

Abstract
Background: A major issue in the treatment of acute myeloid leukemia (AML) is resistance to chemotherapeutic drugs. Multidrug resistance can be caused by ATP-binding cassette (ABC) transporters that function as drug efflux pumps. The majority of these proteins have not yet been examined in malignant diseases.

Experimental Design: A newly developed microarray for the simultaneous quantification of 38 ABC transporter genes and Taqman real-time PCR was used to analyze the expression of ABC transporters in pediatric AML and healthy bone marrow. Small interfering RNA was used to verify the role of ABCA3 in drug resistance.

Results: Using the microarray, we identified four new ABC transporters, which were overexpressed in many AML samples compared with healthy bone marrow: ABCA2, ABCA3, ABCB2, and ABCC10. The overexpression of these four genes was verified by real-time PCR in 42 samples from children with AML and 18 samples of healthy bone marrow. The median expression of ABCA3 was three times higher in 21 patients who had failed to achieve remission after the first course of chemotherapy than in a well-matched group of 21 patients who had achieved remission at this stage (P = 0.023). Incubation of cell lines with a number of different cytostatic drugs induced an up-regulation of ABCA3. Down-regulation of ABCA3 by small interfering RNA sensitized cells to doxorubicin.

Conclusion: Our results show that ABCA2, ABCA3, ABCB2, and ABCC10 are overexpressed in childhood AML compared with healthy bone marrow. ABCA3 is the most likely transporter to cause drug resistance.
Data from BRCA1 Loss Preexisting in Small Subpopulations of Prostate Cancer Is Associated with Advanced Disease and Metastatic Spread to Lymph Nodes and Peripheral Blood
Authors: Bednarz Natalia , Eltze Elke , Semjonow Axel , Rink Michael , Andreas Antje , Mulder Lennart , Hannemann Juliane , Fisch Margit , Pantel Klaus , Weier Heinz-Ulrich G. , Bielawski Krzysztof P. , Brandt Burkhard
Published: April 2023

Abstract
Purpose: A preliminary study performed on a small cohort of multifocal prostate cancer (PCa) detected BRCA1 allelic imbalances among circulating tumor cells (CTC). The present analysis was aimed to elucidate the biological and clinical roles of BRCA1 losses in metastatic spread and tumor progression in PCa patients.

Experimental Design: To map molecular progression in PCa outgrowth, we used fluorescence in situ hybridization analysis of primary tumors and lymph node sections, and CTCs from peripheral blood.

Results: We found that 14% of 133 tested patients carried monoallelic BRCA1 loss in at least one tumor focus. Extended molecular analysis of chr17q revealed that this aberration was often a part of larger cytogenetic rearrangement involving chr17q21 accompanied by allelic imbalance of the tumor suppressor gene PTEN and lack of BRCA1 promoter methylation. The BRCA1 losses correlated with advanced T stage (P < 0.05), invasion to pelvic lymph nodes (P < 0.05), as well as biochemical recurrence (P < 0.01). Their prevalence was twice as high within 62 lymph node metastases (LNM) as in primary tumors (27%, P < 0.01). The analysis of 11 matched primary PCa-LNM pairs confirmed the suspected transmission of genetic abnormalities between these two sites. In four of seven patients with metastatic disease, BRCA1 losses appeared in a minute fraction of cytokeratin- and vimentin-positive CTCs.

Conclusions: Small subpopulations of PCa cells bearing BRCA1 losses might be one confounding factor initiating tumor dissemination and might provide an early indicator of shortened disease-free survival. Clin Cancer Res; 16(13); 3340–8. ©2010 AACR.
Data from HLA-DQB1*02–Restricted HPV-16 E7 Peptide–Specific CD4+ T-Cell Immune Responses Correlate with Regression of HPV-16–Associated High-Grade Squamous Intraepithelial Lesions
Authors: Peng Shiwen , Trimble Cornelia , Wu Lee , Pardoll Drew , Roden Richard , Hung Chien-Fu , Wu T.-C.
Published: April 2023

Abstract
Purpose: The fact that up to 30% of established high-grade squamous intraepithelial lesions (HSIL) of the cervix regress spontaneously presents the opportunity to identify clinically relevant human papillomavirus (HPV) viral epitopes associated with disease outcome. Two human HPV antigens, E6 and E7, are functionally required for initiation and maintenance of cervical cancer precursor lesions and invasive cervical cancer. The identification and characterization of endogenously processed HPV antigenic epitopes in closely characterized patient cohorts will provide insight into the reasons for success or failure of therapeutic approaches.

Experimental Design: We characterized the HPV-16 E6/E7–specific T-cell epitopes using E6/E7 overlapping peptide pools with peripheral blood lymphocytes obtained from normal healthy donors. We then analyzed the difference in the HPV-16 T-cell immune responses in HPV-16+ HSIL patients with or without spontaneous regression of lesions using the statistical methods.

Results: We have identified an HPV-16 E7–specific CD4+ T-cell epitope [amino acids (aa) 71-85] that was restricted by HLA-DQB1*0201. Analysis of peripheral blood lymphocytes obtained from 14 HLA-DQB1*02 patients with HPV-16+ HSILs showed that the HPV-16+ E7 peptide (aa 71-85)–specific CD4+ T-cell immune response was significantly higher in the group of patients with regression compared with the patients without regression (P value <0.05).

Conclusions: The HPV-16 E7 peptide–specific CD4+ T-cell immune response correlates with spontaneous regression of established HPV16+ HSILs. Thus, this E7 epitope may be useful for the characterization of HPV-specific immune responses in patients infected with HPV-16 or immunized with HPV vaccines.
Data from Proteomic Analysis of Malignant Ovarian Cancer Effusions as a Tool for Biologic and Prognostic Profiling
Authors: Davidson Ben , Espina Virginia , Steinberg Seth M. , Flørenes Vivi Ann , Liotta Lance A. , Kristensen Gunnar B. , Tropé Claes G. , Berner Aasmund , Kohn Elise C.
Published: April 2023

Abstract
Purpose: Malignant epithelial ovarian cancer effusions are important in disease dissemination and clinical outcome. The identification of biochemical events active in effusions may improve our identification and application of targeted therapeutics.

Experimental Design: Archival effusion samples for which outcome information was known were studied. Clinical variables were comparable between these groups. Two cohorts of patients with malignant effusion were assessed: those with effusion at presentation (Tap1) or at first recurrence (Tap2). Expression and activated fraction of selected signaling proteins were quantitated on serial protein microarrays using validated antibodies. Proteomic results and clinical variables were analyzed by univariate analysis followed by Cox proportional hazards model analysis.

Results: Malignant effusions (>80% malignant cells) were distinguished from benign effusions by higher expression of AKT, activated extracellular signal-regulated kinase, activated (P ≤ 0.001) and total cAMP-responsive element binding protein (P = 0.01), and JNK (P = 0.03). Malignant pleural effusions could not be differentiated from ascites by signaling profiles. Both had signal expression clusters for survival, proliferation and metastasis, and injury pathways. Cox proportional hazards model analysis revealed high p38 and pEGFR/EGFR ratio as jointly associated with poor survival in Tap1 cases (both P ≤ 0.002). Phospho-JNK quantity was associated with worse outcome in Tap2 patients (P = 0.004), when taking other factors into consideration.

Conclusions: Proliferation, survival, and apoptosis signaling dysregulation can be identified in ovarian cancer effusion samples. Biochemical characterization of clinical effusions may provide either predictive and/or correlative information on patient outcome from which to further understand the mechanisms of effusion development and target clinical intervention.
Data from Phase I Safety, Pharmacokinetic, and Pharmacodynamic Study of ENMD-2076, a Novel Angiogenic and Aurora Kinase Inhibitor,in Patients with Advanced Solid Tumors
Authors: Diamond Jennifer R. , Bastos Bruno R. , Hansen Ryan J. , Gustafson Daniel L. , Eckhardt S. Gail , Kwak Eunice L. , Pandya Shuchi S. , Fletcher Graham C. , Pitts Todd M. , Kulikowski Gillian N. , Morrow Mark , Arnott Jamie , Bray Mark R. , Sidor Carolyn , Messersmith Wells , Shapiro Geoffrey I.
Published: April 2023

Abstract
Purpose: ENMD-2076 is a unique orally bioavailable Aurora kinase and VEGFR inhibitor. The purpose of this phase 1 study of ENMD-2076 was to determine the MTD, pharmacokinetic, and pharmacodynamic profiles and preliminary antitumor activity.

Experimental Design: Patients with refractory advanced solid malignancies were treated with ENMD-2076 orally with continuous once daily dosing. Doses from 60 to 200 mg/m2 were evaluated using a standard 3 (to 4) + 3 design. Pharmacokinetic parameters were studied on days 1, 28, and 30 to 35 of cycle 1. Expanded MTD cohorts included patients with ovarian cancer, colorectal cancer, and refractory solid tumors.

Results: A total of 67 patients (46 F, 21M; ages 30–76) entered the study. Dose levels of 60, 80, 120, 200, and 160 mg/m2 were evaluated. Two patients experienced grade 3 hypertension at 200 mg/m2, and additional grade 3 neutropenia events limited tolerability at this dose. An intermediate dose of 160 mg/m2 was determined to be the MTD. The most common drug-related adverse events included hypertension, nausea/vomiting, and fatigue. The pharmacokinetics of ENMD-2076 were characterized by a rapid absorption phase (Tmax 3–7.8 hours), a t1/2 of 27.3 to 38.3 hours after a single dose, and dose proportional exposure. Decreased plasma sVEGFR2 was observed posttreatment. Two patients with platinum refractory/resistant ovarian cancer had RECIST partial responses.

Conclusions: ENMD-2076 was well tolerated, had a linear pharmacokinetic profile, and showed promising antitumor activity, particularly in ovarian cancer. The recommended phase 2 dose of ENMD-2076 is 160 mg/m2 administered orally once daily with continuous dosing. Clin Cancer Res; 17(4); 849–60. ©2010 AACR.
Data from Prospective Gene Signature Study Using microRNA to Identify the Tissue of Origin in Patients with Carcinoma of Unknown Primary
Authors: Varadhachary Gauri R. , Spector Yael , Abbruzzese James L. , Rosenwald Shai , Wang Huamin , Aharonov Ranit , Carlson Heather R. , Cohen Dalia , Karanth Siddharth , Macinskas Joanna , Lenzi Renato , Chajut Ayelet , Edmonston Tina B. , Raber Martin N.
Published: April 2023

Abstract
Purpose: Accurate identification of tissue of origin (ToO) for patients with carcinoma of unknown primary (CUP) may help customize therapy to the putative primary and thereby improve the clinical outcome. We prospectively studied the performance of a microRNA-based assay to identify the ToO in CUP patients.

Experimental Design: Formalin-fixed paraffin-embedded (FFPE) metastatic tissue from 104 patients was reviewed and 87 of these contained sufficient tumor for testing. The assay quantitates 48 microRNAs and assigns one of 25 tumor diagnoses by using a biologically motivated binary decision tree and a K-nearest neighbors (KNN). The assay predictions were compared with clinicopathologic features and, where suitable, to therapeutic response.

Results: Seventy-four of the 87 cases were processed successfully. The assay result was consistent or compatible with the clinicopathologic features in 84% of cases processed successfully (71% of all samples attempted). In 65 patients, pathology and immunohistochemistry (IHC) suggested a diagnosis or (more often) a differential diagnosis. Out of those, the assay was consistent or compatible with the clinicopathologic presentation in 55 (85%) cases. Of the 9 patients with noncontributory IHC, the assay provided a ToO prediction that was compatible with the clinical presentation in 7 cases.

Conclusions: In this prospective study, the microRNA diagnosis was compatible with the clinicopathologic picture in the majority of cases. Comparative effectiveness research trials evaluating the added benefit of molecular profiling in appropriate CUP subsets are warranted. MicroRNA profiling may be particularly helpful in patients in whom the IHC profile of the metastasis is nondiagnostic or leaves a large differential diagnosis. Clin Cancer Res; 17(12); 4063–70. ©2011 AACR.
Data from Blood-Based Gene Expression Signatures in Non–Small Cell Lung Cancer
Authors: Zander Thomas , Hofmann Andrea , Staratschek-Jox Andrea , Classen Sabine , Debey-Pascher Svenja , Maisel Daniela , Ansén Sascha , Hahn Moritz , Beyer Marc , Thomas Roman K. , Gathof Birgit , Mauch Cornelia , Delank Karl-Stefan , Engel-Riedel Walburga , Wichmann H.-Erich , Stoelben Erich , Schultze Joachim L. , Wolf Jürgen
Published: April 2023

Abstract
Purpose: Blood-based surrogate markers would be attractive biomarkers for early detection, diagnosis, prognosis, and prediction of therapeutic outcome in cancer. Disease-associated gene expression signatures in peripheral blood mononuclear cells (PBMC) have been described for several cancer types. However, RNA-stabilized whole blood–based technologies would be clinically more applicable and robust. We evaluated the applicability of whole blood–based gene expression profiling for the detection of non–small cell lung cancer (NSCLC).

Experimental Design: Expression profiles were generated from PAXgene-stabilized blood samples from three independent groups consisting of NSCLC cases and controls (n = 77, 54, and 102), using the Illumina WG6-VS2 system.

Results: Several genes are consistently differentially expressed in whole blood of NSCLC patients and controls. These expression profiles were used to build a diagnostic classifier for NSCLC, which was validated in an independent validation set of NSCLC patients (stages I–IV) and hospital-based controls. The area under the receiver operator curve was calculated to be 0.824 (P < 0.001). In a further independent dataset of stage I NSCLC patients and healthy controls the AUC was 0.977 (P < 0.001). Specificity of the classifier was validated by permutation analysis in both validation cohorts. Genes within the classifier are enriched in immune-associated genes and show specificity for NSCLC.

Conclusions: Our results show that gene expression profiles of whole blood allow for detection of manifest NSCLC. These results prompt further development of gene expression–based biomarker tests in peripheral blood for the diagnosis and early detection of NSCLC. Clin Cancer Res; 17(10); 3360–7. ©2011 AACR.
Data from Potent siRNA Inhibitors of Ribonucleotide Reductase Subunit RRM2 Reduce Cell Proliferation In vitro and In vivo
Authors: Heidel Jeremy D. , Liu Joanna Yi-Ching , Yen Yun , Zhou Bingsen , Heale Bret S.E. , Rossi John J. , Bartlett Derek W. , Davis Mark E.
Published: April 2023

Abstract
Purpose: Ribonucleotide reductase (RR) is a therapeutic target for DNA replication–dependent diseases such as cancer. Here, a potent small interfering RNA (siRNA) duplex against the M2 subunit of RR (RRM2) is developed and shown to reduce the growth potential of cancer cells both in vitro and in vivo.

Experimental Design: Three anti-RRM2 siRNAs were identified via computational methods, and the potency of these and additional “tiling” duplexes was analyzed in cultured cells via cotransfections using a RRM2-luciferase fusion construct. Knockdown of RRM2 by the best duplex candidates was confirmed directly by Western blotting. The effect of potent duplexes on cell growth was investigated by a real-time cell electronic sensing assay. Finally, duplex performance was tested in vivo in luciferase-expressing cells via whole animal bioluminescence imaging.

Results: Moderate anti-RRM2 effects are observed from the three duplexes identified by computational methods. However, the tiling experiments yielded an extremely potent duplex (siR2B+5). This duplex achieves significant knockdown of RRM2 protein in cultured cells and has pronounced antiproliferative activity. S.c. tumors of cells that had been transfected with siR2B+5 preinjection grew slower than those of control cells.

Conclusions: An anti-RRM2 siRNA duplex is identified that exhibits significant antiproliferative activity in cancer cells of varying human type and species (mouse, rat, monkey); these findings suggest that this duplex is a promising candidate for therapeutic development.
Data from In vitro Drug Sensitivity Predicts Response and Survival after Individualized Sensitivity-Directed Chemotherapy in Metastatic Melanoma: A Multicenter Phase II Trial of the Dermatologic Cooperative Oncology Group
Authors: Ugurel Selma , Schadendorf Dirk , Pföhler Claudia , Neuber Karsten , Thoelke Adina , Ulrich Jens , Hauschild Axel , Spieth Konstanze , Kaatz Martin , Rittgen Werner , Delorme Stefan , Tilgen Wolfgang , Reinhold Uwe
Published: April 2023

Abstract
Purpose:In vitro sensitivity assays are promising tools to predict the individual outcome of different chemotherapy regimens. However, a direct association between in vitro and in vivo chemosensitivity has to be shown by clinical studies. This multicenter phase II trial was aimed to investigate the efficacy of a sensitivity-directed, first-line chemotherapy in metastasized melanoma patients, and to prove an association between in vitro sensitivity and therapy outcome.

Patients and Methods: The primary study end point was objective response; secondary end points were safety, overall survival, and progression-free survival. Viable tumor cells obtained from metastatic lesions were tested for chemosensitivity to seven single drugs and five drug combinations using an ATP-based luminescence viability assay.

Results: Out of 82 recruited patients (intention-to-treat), 57 received assay-directed chemotherapy and 53 were evaluable for all study end points (per protocol). The drug combinations used were gemcitabine + treosulfan, paclitaxel + cisplatin, paclitaxel + doxorubicin, and gemcitabine + cisplatin. The per protocol population could be divided into 22 (42%) chemosensitive and 31 (58%) chemoresistant patients by an arbitrary chemosensitivity index. Objective response was 36.4% in chemosensitive patients compared with 16.1% in chemoresistant patients (P = 0.114); progression arrest (complete response + partial response + stable disease) was 59.1% versus 22.6% (P = 0.01). Chemosensitive patients showed an increased overall survival of 14.6 months compared with 7.4 months in chemoresistant patients (P = 0.041).

Conclusion:In vitro chemosensitivity testing may be worthy of further exploration to see if it could be a useful tool to predict the outcome of melanoma patients treated with a sensitivity-directed chemotherapy. Therefore, these preliminary results will be evaluated by a planned phase III trial using a randomized, standard-regimen controlled setting.
Data from The Use of a Tropism-Modified Measles Virus in Folate Receptor–Targeted Virotherapy of Ovarian Cancer
Authors: Hasegawa Kosei , Nakamura Takafumi , Harvey Mary , Ikeda Yasuhiro , Oberg Ann , Figini Mariangela , Canevari Silvana , Hartmann Lynn C. , Peng Kah-Whye
Published: April 2023

Abstract
Purpose: Attenuated measles viruses are promising experimental anticancer agents currently being evaluated in a phase I dose escalation trial for ovarian cancer patients. Virus attachment, entry, and subsequent intercellular fusion between infected and uninfected neighboring cells are mediated via the two measles receptors (CD46 and SLAM). To minimize potential toxicity due to measles virus–associated immunosuppression and infection of nontarget tissues, we sought to develop an ovarian cancer exclusive fully retargeted measles virus.

Experimental Design and Results: Interactions of measles virus with its natural receptors were ablated, and a single-chain antibody (scFv) specific for α-folate receptor (FRα), a target overexpressed on 90% of nonmucinous ovarian cancer, was genetically engineered on the viral attachment protein (MV-αFR). Specificity of virus tropism was tested on tumor and normal cells. Biodistribution of measles virus infection was evaluated in measles-susceptible CD46 transgenic mice, whereas antitumor activity was monitored noninvasively by bioluminescence imaging in xenograft models. Tropism and fusogenic activity of MV-αFR was redirected exclusively to FRα without compromise to virus infectivity. In contrast to the parental virus, MV-αFR has no background infectivity on normal human cells. The antitumor activity of MV-αFR, as assessed by tumor volume reduction and overall survival increase, was equal to the parental virus in two models of human ovarian cancer (s.c. and i.p.).

Conclusions: A FR-exclusive ovarian cancer targeted oncolytic virus was generated and shown to be therapeutically effective, thus introducing a new modality for FR targeting and a candidate measles virus for clinical testing.
Data from Metformin Amplifies Chemotherapy-Induced AMPK Activation and Antitumoral Growth
Authors: Rocha Guilherme Z. , Dias Marília M. , Ropelle Eduardo R. , Osório-Costa Felipe , Rossato Franco A. , Vercesi Anibal E. , Saad Mario J.A. , Carvalheira José B.C.
Published: April 2023

Abstract
Purpose: Metformin is a widely used antidiabetic drug whose anticancer effects, mediated by the activation of AMP-activated protein kinase (AMPK) and reduction of mTOR signaling, have become noteworthy. Chemotherapy produces genotoxic stress and induces p53 activity, which can cross-talk with AMPK/mTOR pathway. Herein, we investigate whether the combination of metformin and paclitaxel has an effect in cancer cell lines.

Experimental Design: Human tumors were xenografted into severe combined immunodeficient (SCID) mice and the cancer cell lines were treated with only paclitaxel or only metformin, or a combination of both drugs. Western blotting, flow cytometry, and immunohistochemistry were then used to characterize the effects of the different treatments.

Results: The results presented herein show that the addition of metformin to paclitaxel leads to quantitative potentialization of molecular signaling through AMPK and a subsequent potent inhibition of the mTOR signaling pathway. Treatment with metformin and paclitaxel resulted in an increase in the number of cells arrested in the G2–M phase of the cell cycle, and decreased the tumor growth and increased apoptosis in tumor-bearing mice, when compared with individual drug treatments.

Conclusion: We have provided evidence for a convergence of metformin and paclitaxel induced signaling at the level of AMPK. This mechanism shows how different drugs may cooperate to augment antigrowth signals, and suggests that target activation of AMPK by metformin may be a compelling ally in cancer treatment. Clin Cancer Res; 17(12); 3993–4005. ©2011 AACR.
Data from Mapping Geographic Zones of Cancer Risk with Epigenetic Biomarkers in Normal Breast Tissue
Authors: Yan Pearlly S. , Venkataramu Chinnambally , Ibrahim Ashraf , Liu Joseph C. , Shen Rulong Z. , Diaz Nils M. , Centeno Barbara , Weber Frank , Leu Yu-Wei , Shapiro Charles L. , Eng Charis , Yeatman Timothy J. , Huang Tim H.-M.
Published: April 2023

Abstract
Purpose: Genetic alterations were previously identified in normal epithelia adjacent to invasive cancers. The aim of this study was to determine DNA methylation in histologically normal tissues from multiple geographic zones adjacent to primary breast tumors.

Experimental Design: First, methylation status of a 4-kb region of RASSF1A promoter was interrogated using oligonucleotide-based microarray in 144 samples (primary tumors, 47; adjacent normals, 69; reduction mammoplasty tissues, 28). Second, allelic imbalance (AI)/loss of heterozygosity (LOH) surrounding RASSF1A promoter were analyzed in 30 samples (tumors, 8; adjacent normals, 22). Third, global methylation screening of 49 samples (tumors, 12; adjacent normals, 25; reduction mammoplasty, 12) was done by differential methylation hybridization. Real-time quantitative methylation-specific PCR was used to validate the microarray findings.

Results: DNA methylation in the core RASSF1A promoter was low in reduction mammoplasty tissues (P = 0.0001) when compared with primary tumors. The adjacent normals had an intermediate level of methylation. The regions surrounding the core were highly methylated in all sample types. Microsatellite markers showed AI/LOH in tumors and some of the adjacent normals. Concurrent AI/LOH and DNA methylation in RASSF1A promoter occurred in two of six tumors. Global methylation screening uncovered genes more methylated in adjacent normals than in reduction mammoplasty tissues. The methylation status of four genes was confirmed by quantitative methylation-specific PCR.

Conclusions: Our findings suggest a field of methylation changes extending as far as 4 cm from primary tumors. These frequent alterations may explain why normal tissues are at risk for local recurrence and are useful in disease prognostication.
Data from Tumor Regression and Growth Rates Determined in Five Intramural NCI Prostate Cancer Trials: The Growth Rate Constant as an Indicator of Therapeutic Efficacy
Authors: Stein Wilfred D. , Gulley James L. , Schlom Jeff , Madan Ravi A. , Dahut William , Figg William D. , Ning Yang-min , Arlen Phil M. , Price Doug , Bates Susan E. , Fojo Tito
Published: April 2023

Abstract
Purpose: In solid tumors such as prostate cancer, novel paradigms are needed to assess therapeutic efficacy. We utilized a method estimating tumor growth and regression rate constants from serial PSA measurements, and assessed its potential in patients with metastatic castration resistant prostate carcinoma (mCRPC).

Experimental Design: Patients were enrolled in five phase II studies, including an experimental vaccine trial, representing the evolution of therapy in mCRPC. PSA measurements obtained before, and during, therapy were used. Data analysis using a two-phase mathematical equation yielded concomitant PSA growth and regression rate constants.

Results: Growth rate constants (g) can be estimated while patients receive therapy and in such patients g is superior to PSA-DT in predicting OS. Incremental reductions in growth rate constants were recorded in successive trials with a 10-fold slower g in the most recent combination therapy trial (log g = 10−3.17) relative to single-agent thalidomide (log g = 10−2.08) more than a decade earlier. Growth rate constants correlated with survival, except in patients receiving vaccine-based therapy where the evidence demonstrates prolonged survival presumably due to immunity developing subsequent to vaccine administration.

Conclusion: Incremental reductions in tumor growth rate constants suggest increased efficacy in successive chemotherapy trials. The derived growth rate constant correlates with survival, and may be used to assess efficacy. The PSA-TRICOM vaccine appears to have provided marked benefit not apparent during vaccination, but consistent with subsequent development of a beneficial immune response. If validated as a surrogate for survival, growth rate constants would offer an important new efficacy endpoint for clinical trials. Clin Cancer Res; 17(4); 907–17. ©2010 AACR.
Data from Gene Expression Profile Associated with Response to Doxorubicin-Based Therapy in Breast Cancer
Authors: Koike Folgueira Maria Aparecida Azevedo , Carraro Dirce Maria , Brentani Helena , da Costa Patrão Diogo Ferreira , Barbosa Edson Mantovani , Netto Mário Mourão , Caldeira José Roberto Fígaro , Katayama Maria Lucia Hirata , Soares Fernando Augusto , Oliveira Célia Tosello , Reis Luiz Fernando Lima , Kaiano Jane Haruko Lima , Camargo Luiz Paulo , Vêncio Ricardo Zorzetto Nicoliello , Snitcovsky Igor Moysés Longo , Makdissi Fabiana Baroni Alves , da Silva e Silva Paulo José , Góes João Carlos Guedes Sampaio , Brentani Maria Mitzi
Published: April 2023

Abstract
Purpose: This study was designed to identify genes that could predict response to doxorubicin-based primary chemotherapy in breast cancer patients.

Experimental Design: Biopsy samples were obtained before primary treatment with doxorubicin and cyclophosphamide. RNA was extracted and amplified and gene expression was analyzed using cDNA microarrays.

Results: Response to chemotherapy was evaluated in 51 patients, and based on Response Evaluation Criteria in Solid Tumors guidelines, 42 patients, who presented at least a partial response (≥30% reduction in tumor dimension), were classified as responsive. Gene profile of samples, divided into training set (n = 38) and independent validation set (n = 13), were at first analyzed against a cDNA microarray platform containing 692 genes. Unsupervised clustering could not separate responders from nonresponders. A classifier was identified comprising EMILIN1, FAM14B, and PBEF, which however could not correctly classify samples included in the validation set. Our next step was to analyze gene profile in a more comprehensive cDNA microarray platform, containing 4,608 open reading frame expressed sequence tags. Seven samples of the initial training set (all responder patients) could not be analyzed. Unsupervised clustering could correctly group all the resistant samples as well as at least 85% of the sensitive samples. Additionally, a classifier, including PRSS11, MTSS1, and CLPTM1, could correctly distinguish 95.4% of the 44 samples analyzed, with only two misclassifications, one sensitive sample and one resistant tumor. The robustness of this classifier is 2.5 greater than the first one.

Conclusion: A trio of genes might potentially distinguish doxorubicin-responsive from nonresponsive tumors, but further validation by a larger number of samples is still needed.
Data from CTLA4 Blockade Induces Frequent Tumor Infiltration by Activated Lymphocytes Regardless of Clinical Responses in Humans
Authors: Huang Rong Rong , Jalil Jason , Economou James S. , Chmielowski Bartosz , Koya Richard C. , Mok Stephen , Sazegar Hooman , Seja Elizabeth , Villanueva Arturo , Gomez-Navarro Jesus , Glaspy John A. , Cochran Alistair J. , Ribas Antoni
Published: April 2023

Abstract
Background: CTLA4 blocking monoclonal antibodies provide durable clinical benefit in a subset of patients with advanced melanoma mediated by intratumoral lymphocytic infiltrates. A key question is defining whether the intratumoral infiltration (ITI) is a differentiating factor between patients with and without tumor responses.

Methods: Paired baseline and postdosing tumor biopsy specimens were prospectively collected from 19 patients with metastatic melanoma, including 3 patients with an objective tumor response, receiving the anti-CTLA4 antibody tremelimumab within a clinical trial with primary endpoint of quantitating CD8+ cytotoxic T-lymphocyte (CTL) infiltration in tumors. Samples were analyzed for cell density by automated imaging capture and further characterized for functional lymphocyte properties by assessing the cell activation markers HLA-DR and CD45RO, the cell proliferation marker Ki67, and the regulatory T-cell marker FOXP3.

Results: There was a highly significant increase in ITI by CD8+ cells in biopsy samples taken after tremelimumab treatment. This included increases between 1-fold and 100-fold changes in 14 of 18 evaluable cases regardless of clinical tumor response or progression. There was no difference between the absolute number, location, or cell density of infiltrating cells between clinical responders and patients with nonresponding lesions that showed acquired intratumoral infiltrates. There were similar levels of expression of T-cell activation markers (CD45RO, HLA-DR) in both groups and no difference in markers for cell replication (Ki67) or the suppressor cell marker FOXP3.

Conclusion: CTLA4 blockade induces frequent increases in ITI by T cells despite which only a minority of patients have objective tumor responses. Clin Cancer Res; 17(12); 4101–9. ©2011 AACR.
Data from Imaging Epidermal Growth Factor Receptor Expression In vivo: Pharmacokinetic and Biodistribution Characterization of a Bioconjugated Quantum Dot Nanoprobe
Authors: Diagaradjane Parmeswaran , Orenstein-Cardona Jacobo M. , E. Colón-Casasnovas Norman , Deorukhkar Amit , Shentu Shujun , Kuno Norihito , Schwartz David L. , Gelovani Juri G. , Krishnan Sunil
Published: April 2023

Abstract
Purpose: To develop and validate an optical imaging nanoprobe for the discrimination of epidermal growth factor (EGF) receptor (EGFR)–overexpressing tumors from surrounding normal tissues that also expresses EGFR.

Experimental Design: Near-infrared (NIR) quantum dots (QD) were coupled to EGF using thiol-maleimide conjugation to create EGF-QD nanoprobes. In vitro binding affinity of these nanoprobes and unconjugated QDs was evaluated in a panel of cell lines, with and without anti-EGFR antibody pretreatment. Serial optical imaging of HCT116 xenograft tumors was done after systemic injection of QD and EGF-QD.

Results: EGF-QD showed EGFR-specific binding in vitro. In vivo imaging showed three distinct phases, tumor influx (∼3 min), clearance (∼60 min), and accumulation (1-6 h), of EGF-QD nanoprobes. Both QD and EGF-QD showed comparable nonspecific rapid tumor influx and clearance followed by attainment of an apparent dynamic equilibrium at ∼60 min. Subsequently (1-6 h), whereas QD concentration gradually decreased in tumors, EGF-QDs progressively accumulated in tumors. On delayed imaging at 24 h, tumor fluorescence decreased to near-baseline levels for both QD and EGF-QD. Ex vivo whole-organ fluorescence, tissue homogenate fluorescence, and confocal microscopic analyses confirmed tumor-specific accumulation of EGF-QD at 4 h. Immunofluorescence images showed diffuse colocalization of EGF-QD fluorescence within EGFR-expressing tumor parenchyma compared with patchy perivascular sequestration of QD.

Conclusion: These results represent the first pharmacokinetic characterization of a robust EGFR imaging nanoprobe. The measurable contrast enhancement of tumors 4 h after systemic administration of EGF-QD and its subsequent normalization at 24 h imply that this nanoprobe may permit quantifiable and repetitive imaging of EGFR expression.