bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Rapid Detection of Epidermal Growth Factor Receptor Mutations in Lung Cancer by the SMart-Amplification Process
Authors: Hoshi Kanako , Takakura Hideki , Mitani Yasumasa , Tatsumi Kenji , Momiyama Nobuyoshi , Ichikawa Yasushi , Togo Shinji , Miyagi Toru , Kawai Yuki , Kogo Yasushi , Kikuchi Takeshi , Kato Chiaki , Arakawa Takahiro , Uno Syuji , Cizdziel Paul E. , Lezhava Alexander , Ogawa Noburou , Hayashizaki Yoshihide , Shimada Hiroshi
Published: April 2023

Abstract
Purpose: A positive response to gefitinib in non–small cell lung cancer (NSCLC) has been correlated to mutations in epidermal growth factor receptor (EGFR) gene. Previous reports have been based mainly on diagnostic screening by sequencing. However, sequencing is a time-consuming and complicated procedure, not suitable for routine clinical use.

Experimental Design: We have developed rapid, simple, and sensitive mutation detection assays based on the SMart Amplification Process (SMAP) and applied it for analyzing EGFR gene mutations in clinical samples. By using SMAP, we can detect mutations within 30 min including sample preparation. To validate the assay system for potential use in clinical diagnostics, we examined 45 NSCLC patients for EGFR mutations using sequencing and SMAP.

Results: The outcomes of the SMAP assay perfectly matched the sequencing results, except in one case where SMAP was able to identify a mutation that was not detected by sequencing. We also evaluated the sensitivity and specificity of SMAP in mutation detection for EGFR. In a serial dilution study, SMAP was able to find a mutation in a sample containing only 0.1% of the mutant allele in a mixture of wild-type genomic DNA. We also could show amplification of mutated DNA with only 30 copies per reaction.

Conclusions: The SMAP method offers higher sensitivity and specificity than alternative technologies, while eliminating the need for sequencing to identify mutations in the EGFR gene of NSCLC. It provides a robust and point-of-care accessible approach for a rapid identification of most patients likely to respond to gefitinib.
Data from Clinical Significance of Cytokeratin 20-Positive Circulating Tumor Cells Detected by a Refined Immunomagnetic Enrichment Assay in Colorectal Cancer Patients
Authors: Wong Sze Chuen Cesar , Chan Charles Ming Lok , Ma Brigette Buig Yue , Hui Edwin Pun , Ng Simon Siu Man , Lai Paul Bo San , Cheung Moon Tong , Lo Elena Siu Fong , Chan Amanda Kit Ching , Lam Money Yan Yee , Au Thomas Chi Chuen , Chan Anthony Tak Cheung
Published: April 2023

Abstract
Purpose: Current immunomagnetic enrichment method can only detect general epithelial antigens of circulating tumor cells (CTC). Further characterization of the CTCs to provide specific information on the tumor type is not possible. We attempted to overcome this drawback by developing the methodology for using a gastrointestinal-specific anti-cytokeratin (CK) 20 antibody to detect CTCs in colorectal cancer patients' blood.

Experimental Design: The protocol was validated using a colorectal cancer SW480 cell line. The clinical significance of findings in colorectal cancer was investigated by detecting CK20-positive CTCs (pCTC) in patients with colorectal cancer, other common cancers, colorectal adenoma, benign colorectal diseases, and normal subjects. Moreover, the malignant nature of CK20 pCTCs was examined by comparing chromosome 17 aberration patterns with those from the corresponding primary tumors.

Results: The assay successfully showed CK20-positive SW480 cells. When applied in patient samples, the detection rates were 62% (132 colorectal cancer patients; median number = 11 CTCs), 0% (120 patients with other common cancers), 6% (50 colorectal adenoma patients), 0% (120 patients with benign colorectal diseases), and 0% (40 normal subjects). Furthermore, statistical analysis showed that CK20 pCTC numbers were associated with tumor-node-metastasis stage and lymph node status. Using the median CK20 pCTC numbers as the cutoff points, stratified groups of colorectal cancer patients had significant differences in their recurrence, metastasis, and survival. Finally, chromosome 17 aneusomy in 90% of colorectal cancer patients with CK20 pCTCs matched with those from the primary tumors.

Conclusions: Detection of CK20 pCTCs using the new protocol could generate clinically important information for colorectal cancer patients.
Data from Expression of Mutated IGHV3-23 Genes in Chronic Lymphocytic Leukemia Identifies a Disease Subset with Peculiar Clinical and Biological Features
Authors: Bomben Riccardo , Dal-Bo Michele , Benedetti Dania , Capello Daniela , Forconi Francesco , Marconi Daniela , Bertoni Francesco , Maffei Rossana , Laurenti Luca , Rossi Davide , Del Principe Maria Ilaria , Luciano Fabrizio , Sozzi Elisa , Cattarossi Ilaria , Zucchetto Antonella , Rossi Francesca Maria , Bulian Pietro , Zucca Emanuele , Nicoloso Milena S. , Degan Massimo , Marasca Roberto , Efremov Dimitar G. , Del Poeta Giovanni , Gaidano Gianluca , Gattei Valter
Published: April 2023

Abstract
Purpose: B-cell chronic lymphocytic leukemia (CLL) is a clinically heterogeneous disease whose outcome can be foreseen by investigating the mutational status of immunoglobulin heavy chain variable (IGHV) genes. Moreover, a different prognosis was reported for CLL expressing specific IGHV genes in the context or not of stereotyped B-cell receptors. Here we investigated novel associations between usage of specific IGHV genes and clinical features in CLL.

Experimental Design: Among 1,426 CLL-specific IG-rearrangements, stereotyped B-cell receptor clusters never utilized the IGHV3-23 gene. Given this notion, this study was aimed at characterizing the IGHV3-23 gene in CLL, and identifying the properties of IGHV3-23–expressing CLL.

Results: IGHV3-23 was the second most frequently used (134 of 1,426) and usually mutated (M; 109 of 134) IGHV gene in our CLL series. In the vast majority of M IGHV3-23 sequences, the configuration of the 13 amino acids involved in superantigen recognition was consistent with superantigen binding. Clinically, M IGHV3-23 CLL had shorter time-to-treatment than other M non–IGHV3-23 CLL, and multivariate analyses selected IGHV3-23 gene usage, Rai staging, and chromosomal abnormalities as independent prognosticators for M CLL. Compared with M non–IGHV3-23 CLL, the gene expression profile of M IGHV3-23 CLL was deprived in genes, including the growth/tumor suppressor genes PDCD4, TIA1, and RASSF5, whose downregulation is under control of miR-15a and miR-16-1. Accordingly, relatively higher levels of miR-15a and miR-16-1 were found in M IGHV3-23 compared with M non–IGHV3-23 CLL.

Conclusions: Altogether, expression of the IGHV3-23 gene characterizes a CLL subset with distinct clinical and biological features. Clin Cancer Res; 16(2); 620–8
Data from Partial CD4 Depletion Reduces Regulatory T Cells Induced by Multiple Vaccinations and Restores Therapeutic Efficacy
Authors: LaCelle Michael G. , Jensen Shawn M. , Fox Bernard A.
Published: April 2023

Abstract
Purpose: A single vaccination of intact or reconstituted-lymphopenic mice (RLM) with a granulocyte macrophage colony-stimulating factorsecreting B16BL6-D5 melanoma cell line induces protective antitumor immunity and T cells that mediate the regression of established melanoma in adoptive immunotherapy studies. We wanted to study if multiple vaccinations during immune reconstitution of the lymphopenic host would maintain a potent antitumor immune response.

Experimental Design: RLM were vaccinated multiple times over a 40-day period. Spleens were isolated from these mice, activated in vitro, and adoptively transferred into mice bearing 3-day experimental pulmonary metastases.

Results: Multiple vaccinations, rather than boosting the immune response, significantly reduced therapeutic efficacy of adoptive immunotherapy and were associated with an increased frequency and absolute number of CD3+CD4+Foxp3+ T regulatory (Treg) cells. Anti-CD4 administration reduced the absolute number of Treg cells 9-fold. Effector T-cells generated from anti-CD4treated mice were significantly (P < 0.0001) more therapeutic in adoptive transfer studies than T cells from multiply vaccinated animals with a full complement of CD4+ cells.

Conclusion: These results suggest that CD4+ Treg cells limit the efficacy of multiple vaccinations and that timed partial depletion of CD4+ T cells may reduce suppression and tip-the-balance in favor of therapeutic antitumor immunity. The recent failure of large phase III cancer vaccine clinical trials, wherein patients received multiple vaccines, underscores the potential clinical relevance of these findings. (Clin Cancer Res 2009;15(22):688190)
Data from Herbal Extract of Wedelia chinensis Attenuates Androgen Receptor Activity and Orthotopic Growth of Prostate Cancer in Nude Mice
Authors: Tsai Chin-Hsien , Lin Feng-Min , Yang Yu-Chih , Lee Ming-Ting , Cha Tai-Lung , Wu Guan-James , Hsieh Shih-Chuan , Hsiao Pei-Wen
Published: April 2023

Abstract
Purpose: Wedelia chinensis is a common ingredient of anti-inflammatory herbal medicines in Taiwan and southern China. Inflammation is involved in promoting tumor growth, invasion, and metastasis. This study aims to test the biological effects in vivo of W. chinensis extract on prostate cancer.

Experimental Design: The in vivo efficacy and mechanisms of action of oral administration of a standardized extract of W. chinensis were analyzed in animals bearing a subcutaneous or orthotopic prostate cancer xenograft.

Results: Exposure of prostate cancer cells to W. chinensis extract induced apoptosis selectively in androgen receptor (AR)–positive prostate cancer cells and shifted the proportion in each phase of cell cycle toward G2-M phase in AR-negative prostate cancer cells. Oral herbal extract (4 or 40 mg/kg/d for 24-28 days) attenuated the growth of prostate tumors in nude mice implanted at both subcutaneous (31% and 44%, respectively) and orthotopic (49% and 49%, respectively) sites. The tumor suppression effects were associated with increased apoptosis and lower proliferation in tumor cells as well as reduced tumor angiogenesis. The antitumor effect of W. chinensis extract was correlated with accumulation of the principle active compounds wedelolactone, luteolin, and apigenin in vivo.

Conclusion: Anticancer action of W. chinensis extract was due to three active compounds that inhibit the AR signaling pathway. Oral administration of W. chinensis extract impeded prostate cancer tumorigenesis. Future studies of W. chinensis for chemoprevention or complementary medicine against prostate cancer in humans are thus warranted. (Clin Cancer Res 2009;15(17):5435–44)
Data from ERBB1 and ERBB2 Have Distinct Functions in Tumor Cell Invasion and Intravasation
Authors: Kedrin Dmitriy , Wyckoff Jeffrey , Boimel Pamela J. , Coniglio Salvatore J. , Hynes Nancy E. , Arteaga Carlos L. , Segall Jeffrey E.
Published: April 2023

Abstract
Purpose: The epidermal growth factor receptor (ERBB1) and related family member HER-2/neu (ERBB2) are often overexpressed in aggressive breast cancers and their overexpression is correlated with poor prognosis. Clinical studies using ERBB inhibitors have focused on tumor growth effects, but ERBBs can contribute to malignancy independent of their effects on tumor growth. Our studies were designed to evaluate the effect of ERBB inhibition on tumor cell motility and intravasation in vivo using clinically relevant small-molecule inhibitors.

Experimental Design: Using in vivo mouse models of breast cancer, we test the effects of ERBB1 and ERBB2 inhibitors AC480 and lapatinib, ERBB1 inhibitor gefitinib, and ERBB2 inhibitor AG825 on in vivo tumor cell invasive properties in mammary fat pad tumors.

Results: ERBB1 and ERBB2 inhibition rapidly (within 3 h) inhibits both tumor cell motility and intravasation. Using gefitinib, ERBB1 inhibition rapidly inhibits tumor cell motility and invasion but not intravasation, whereas ERBB2 inhibition by AG825 rapidly blocks intravasation.

Conclusions: ERBB1 and ERBB2 inhibition can rapidly block tumor cell invasive properties. In addition, we differentiate for the first time the contributions of ERBB1 and ERBB2 to the key metastatic properties of in vivo tumor cell invasion and intravasation. These experiments temporally and molecularly separate two key stages in tumor cell entry into blood vessels: invasion and intravasation. These results indicate that ERBB inhibition should be considered for blocking other tumor cell malignant properties besides growth.
Data from Rituximab-Mediated Cell Signaling and Chemo/Immuno-sensitization of Drug-Resistant B-NHL Is Independent of Its Fc Functions
Authors: Vega Mario I. , Huerta-Yepez Sara , Martinez-Paniagua Melisa , Martinez-Miguel Bernardo , Hernandez-Pando Rogelio , González-Bonilla Cesar R. , Chinn Paul , Hanna Nabil , Hariharan Kandasamy , Jazirehi Ali R. , Bonavida Benjamin
Published: April 2023

Abstract
Purpose: Rituximab [chimeric anti-CD20 monoclonal antibody], alone or combined with chemotherapy, is used in the treatment of non–Hodgkin's lymphoma (NHL). Rituximab binds to CD20 and inhibits intracellular survival/growth pathways leading to chemo/immunosensitization of tumor cells in vitro. The contribution of rituximab Fc-FcR interaction in signaling is not known. This study examined the role of Fc-FcR interactions in rituximab-induced signaling using rituximab (Fab')2 fragments as well as rituximab devoid of the CH2 Fc-binding domain (CH2−).

Experimental Design: Rituximab (CH2−) and rituximab (Fab')2 were tested for their activity on B-NHL cell lines. Cell signaling and sensitization to chemotherapy and immunotherapy were examined. The in vitro studies were validated in mice bearing tumor xenografts.

Results: Although the modified antibodies were defective in antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity functions, they retained all other biological activities such as inhibition of cell proliferation, induction of cell aggregation, and apoptosis induction. In addition, similar to rituximab, the modified antibodies inhibited the activity of cell survival/growth pathways and their associated transcription factors (e.g., NF-κB, YY1, SP-1), and signal transducers and activators of transcription 3 (STAT-3), and downregulated the expression of antiapoptotic gene products, such as Bcl-2/Bclxl, which regulate drug resistance. The modified antibodies, similar to rituximab, sensitized resistant B-NHL cells to both CDDP and Fas ligand–induced apoptosis. Furthermore, treatment of nude mice bearing Raji tumor cell xenografts with the combination of rituximab (Fab')2 or rituximab and CDDP resulted in similar and significant inhibition of tumor growth.

Conclusion: These findings reveal that rituximab-mediated inhibition of intracellular signaling pathways and leading to chemo/immuno-sensitization of resistant B-NHL is Fc independent. (Clin Cancer Res 2009;15(21):6582–94)
Data from A CYP3A4 Phenotype–Based Dosing Algorithm for Individualized Treatment of Irinotecan
Authors: van der Bol Jessica M. , Mathijssen Ron H.J. , Creemers Geert-Jan M. , Planting André S.Th. , Loos Walter J. , Wiemer Erik A.C. , Friberg Lena E. , Verweij Jaap , Sparreboom Alex , de Jong Floris A.
Published: April 2023

Abstract
Purpose: Irinotecan, the prodrug of SN-38, is extensively metabolized by cytochrome P450-3A4 (CYP3A4). A randomized trial was done to assess the utility of an algorithm for individualized irinotecan dose calculation based on a priori CYP3A4 activity measurements by the midazolam clearance test.

Experimental Design: Patients were randomized to receive irinotecan at a conventional dose level of 350 mg/m2 (group A) or doses based on an equation consisting of midazolam clearance, γ-glutamyl-transferase, and height (group B). Pharmacokinetics and toxicities were obtained during the first treatment course.

Results: Demographics of 40 evaluable cancer patients were balanced between both groups, including UGT1A1*28 genotype and smoking status. The absolute dose of irinotecan ranged from 480 to 800 mg in group A and 380 to 1,060 mg in group B. The mean absolute dose and area under the curve of irinotecan and SN-38 were not significantly different in either group (P > 0.18). In group B, the interindividual variability in the area under the curve of irinotecan and SN-38 was reduced by 19% and 25%, respectively (P > 0.22). Compared with group A, the incidence of grades 3 to 4 neutropenia was >4-fold lower in group B (45 versus 10%; P = 0.013). The incidence of grades 3 to 4 diarrhea was equal in both groups (10%).

Conclusions: Incorporation of CYP3A4 phenotyping in dose calculation resulted in an improved predictability of the pharmacokinetic and toxicity profile of irinotecan, thereby lowering the incidence of severe neutropenia. In combination with UGT1A1*28 genotyping, CYP3A4 phenotype determination should be explored further as a strategy for the individualization of irinotecan treatment. Clin Cancer Res; 16(2); 736–42
Data from Identification of Cyclooxygenase-2 as a Major Actor of the Transcriptomic Adaptation of Endothelial and Tumor Cells to Cyclic Hypoxia: Effect on Angiogenesis and Metastases
Authors: Daneau Géraldine , Boidot Romain , Martinive Philippe , Feron Olivier
Published: April 2023

Abstract
Purpose: Cyclic hypoxia in tumors originates from heterogeneities in RBC flux and influences not only tumor cells but also endothelial cells lining tumor blood vessels. Whether pO2 fluctuations, particularly transient reoxygenation periods, alter the well-known hypoxia-inducible factor (HIF)–dependent gene program is largely unknown.

Experimental Design: We compared the transcriptomic profiles of endothelial and tumor cells exposed to cyclic hypoxia versus continuous hypoxia to uncover a possible differential effect on angiogenesis and metastases.

Results: Microarray analyses identified early genes that were selectively induced by cyclic hypoxia in endothelial cells. Among them, we focused on PTGS2 because the observed increase in mRNA expression led to a significant increase in the expression and activity of cyclooxygenase-2 (COX-2; the protein product of PTGS2). HIF-1α was shown to be stabilized by cyclic hypoxia (despite reoxygenation periods) and to favor COX-2 induction as validated by the use of echinomycin and HIF-1α targeting small interfering RNA. Using a specific COX-2 inhibitor and a dedicated COX-2 targeting small interfering RNA, we documented that COX-2 accounted for the higher endothelial cell survival and angiogenic potential conferred by cyclic hypoxia. Cyclic hypoxia also led to a preferential COX-2 induction in tumor cells and, contrary to continuous hypoxia, fostered a higher metastatic take of prechallenged tumor cells.

Conclusions: Our study documents that PTGS2/COX-2 is part of a cyclic hypoxia gene signature and largely accounts for the unique phenotype of endothelial and tumor cells exposed to fluctuations in pO2, thereby offering new perspectives for the clustering of tumors expressing COX-2 together with other cyclic hypoxia-responsive genes. Clin Cancer Res; 16(2); 410–9
Data from Characterizing the Clinical Relevance of an Embryonic Stem Cell Phenotype in Lung Adenocarcinoma
Authors: Stevenson Marvaretta , Mostertz William , Acharya Chaitanya , Kim William , Walters Kelli , Barry William , Higgins Kristin , Tuchman Sascha A. , Crawford Jeffrey , Vlahovic Gordana , Ready Neal , Onaitis Mark , Potti Anil
Published: April 2023

Abstract
Purpose: Cancer cells possess traits reminiscent of those ascribed to normal stem cells. It is unclear whether these phenotypic similarities are the result of a common biological phenotype, such as regulatory pathways.

Experimental Design: Lung cancer cell lines with corresponding gene expression data and genes associated with an embryonic stem cell identity were used to develop a signature of embryonic stemness (ES) activity specific to lung adenocarcinoma. Biological characteristics were elucidated as a function of cancer biology/oncogenic pathway dysregulation. The ES signature was applied to three independent early-stage (I-IIIa) lung adenocarcinoma data sets with clinically annotated gene expression data. The relationship between the ES phenotype and cisplatin (current standard of care) sensitivity was evaluated.

Results: Pathway analysis identified specific regulatory networks [Ras (P = 0.0005), Myc (P = 0.0224), wound healing (P < 0.0001), chromosomal instability (P < 0.0001), and invasiveness (P < 0.0001)] associated with the ES phenotype. The prognostic relevance of the ES signature, as related to patient survival, was characterized in three cohorts [CALGB 9761 (n = 82; P = 0.0001), National Cancer Institute Director's Challenge Consortium (n = 442; P = 0.0002), and Duke (n = 45; P = 0.06)]. The ES signature was not prognostic in prostate, breast, or ovarian adenocarcinomas. Lung tumors (n = 569) and adenocarcinoma cell lines (n = 31) expressing the ES phenotype were more likely to be resistant to cisplatin (P < 0.0001 and P = 0.006, respectively).

Conclusions: Lung adenocarcinomas that share a common gene expression pattern with normal human embryonic stem cells were associated with decreased survival, increased biological complexity, and increased likelihood of resistance to cisplatin. This indicates the aggressiveness of these tumors. (Clin Cancer Res 2009;15(24):7553–61)
Data from IGFBP2 and IGFBP3 Protein Expressions in Human Breast Cancer: Association with Hormonal Factors and Obesity
Authors: Probst-Hensch Nicole M. , Steiner Julia H.B. , Schraml Peter , Varga Zsuzsanna , Zürrer-Härdi Ursina , Storz Martina , Korol Dimitri , Fehr Mathias K. , Fink Daniel , Pestalozzi Bernhard C. , Lütolf Urs M. , Theurillat Jean-Philippe , Moch Holger
Published: April 2023

Abstract
Purpose: The insulin-like growth factor (IGF) signaling system is involved in breast cancer initiation and progression. The prognostic relevance of tumor expression patterns of IGFI-related proteins remains poorly understood. This study associates the expression of selected IGF proteins with breast tumor and patient characteristics.

Experimental Design: IGFI, IGFI receptor, IGF-binding protein (IGFBP)2, and IGFBP3 expression was measured in 855 primary breast carcinomas by immunohistochemistry using tissue microarrays. We investigated the association of tumor and nodal stage, grade, hormone receptor status, HER2 gene amplification, menopausal status, body mass index, and survival with IGF protein expression.

Results: In contrast to IGFI, the expression of IGFI receptor, IGFBP2, and IGFBP3 was associated with estrogen receptor status. In addition, IGFBP3 was positively correlated with body mass index and premenopausal status. Importantly, IGFBP2 was an independent and positive predictor of overall survival (hazard ratio, 0.48; 95% confidence interval, 0.24-0.95; P = 0.04). There was a weak suggestion for IGFBP2 and overweight to modify each other's effect on survival.

Conclusions: According to these results, which need confirmation in larger patient series, the prognostic relevance of IGFBP2 and IGFBP3 protein expressions in breast cancer may depend on the hormonal context and body weight. Clin Cancer Res; 16(3); 1025–32
Data from Immunomodulation of FOXP3+ Regulatory T Cells by the Aromatase Inhibitor Letrozole in Breast Cancer Patients
Authors: Generali Daniele , Bates Gaynor , Berruti Alfredo , Brizzi Maria P. , Campo Leticia , Bonardi Simone , Bersiga Alessandra , Allevi Giovanni , Milani Manuela , Aguggini Sergio , Dogliotti Luigi , Banham Alison H. , Harris Adrian L. , Bottini Alberto , Fox Stephen B.
Published: April 2023

Abstract
Purpose: We have shown previously that tumor infiltration by FOXP3+ regulatory T cells (Treg) is associated with increased relapse and shorter survival of patients with both in situ and invasive breast cancer. Because estrogen regulates Treg numbers in mice and promotes the proliferation of human Tregs, we hypothesized that blocking estrogen receptor-α signaling would abrogate Tregs and be associated with response to hormonal therapy and increased survival.

Experimental Design: FOXP3+ Tregs were quantified in tumor samples collected at baseline by incisional biopsy and after 6 months at definitive surgery in 83 elderly breast cancer patients (T2-4 N0-1) enrolled in a randomized phase II trial based on 6 months of primary letrozole (2.5 mg/d) or 6 months of letrozole plus oral “metronomic” cyclophosphamide (50 mg/d).

Results: Treg number ranged from 0 to 380 (median, 30) before treatment and from 0 to 300 (median, 8) after treatment. There was a significant reduction in Tregs in letrozole and letrozole-cyclophosphamide patients (P < 0.0001 and P < 0.002, respectively) after treatment. Treg number at residual histology was inversely related with response (P < 0.03 and P = 0.50, respectively) and a greater Treg reduction was observed in responding patients (P < 0.03).

Conclusion: This study suggests that aromatase inhibitors may have an indirect antitumor mechanism of action through reducing Tregs in breast tumors and may be of use in estrogen receptor-α-negative tumors in combination with immunotherapy approaches.
Data from Prostate Cancer–Associated Gene Expression Alterations Determined from Needle Biopsies
Authors: Qian David Z. , Huang Chung-Ying , O'Brien Catherine A. , Coleman Ilsa M. , Garzotto Mark , True Lawrence D. , Higano Celestia S. , Vessella Robert , Lange Paul H. , Nelson Peter S. , Beer Tomasz M.
Published: April 2023

Abstract
Purpose: To accurately identify gene expression alterations that differentiate neoplastic from normal prostate epithelium using an approach that avoids contamination by unwanted cellular components and is not compromised by acute gene expression changes associated with tumor devascularization and resulting ischemia.

Experimental Design: Approximately 3,000 neoplastic and benign prostate epithelial cells were isolated using laser capture microdissection from snap-frozen prostate biopsy specimens provided by 31 patients who subsequently participated in a clinical trial of preoperative chemotherapy. cDNA synthesized from amplified total RNA was hybridized to custom-made microarrays composed of 6,200 clones derived from the Prostate Expression Database. Expression differences for selected genes were verified using quantitative reverse transcription-PCR.

Results: Comparative analyses identified 954 transcript alterations associated with cancer (q < 0.01%), including 149 differentially expressed genes with no known functional roles. Gene expression changes associated with ischemia and surgical removal of the prostate gland were absent. Genes up-regulated in prostate cancer were statistically enriched in categories related to cellular metabolism, energy use, signal transduction, and molecular transport. Genes down-regulated in prostate cancers were enriched in categories related to immune response, cellular responses to pathogens, and apoptosis. A heterogeneous pattern of androgen receptor expression changes was noted. In exploratory analyses, androgen receptor down-regulation was associated with a lower probability of cancer relapse after neoadjuvant chemotherapy followed by radical prostatectomy.

Conclusions: Assessments of tumor phenotypes based on gene expression for treatment stratification and drug targeting of oncogenic alterations may best be ascertained using biopsy-based analyses where the effects of ischemia do not complicate interpretation.
Data from Crosstalk between Mast Cells and Pancreatic Cancer Cells Contributes to Pancreatic Tumor Progression
Authors: Strouch Matthew J. , Cheon Eric C. , Salabat Mohammad R. , Krantz Seth B. , Gounaris Elias , Melstrom Laleh G. , Dangi-Garimella Surabhi , Wang Edward , Munshi Hidayatullah G. , Khazaie Khashayarsha , Bentrem David J.
Published: April 2023

Abstract
Purpose: To assess the clinical and pathologic significance of mast cell infiltration in human pancreatic cancer and evaluate crosstalk between mast cells and cancer cells in vitro.

Experimental Design: Immunohistochemistry for tryptase was done on 53 pancreatic cancer specimens. Mast cell counts were correlated with clinical variables and survival. Serum tryptase activity from patients with cancer was compared with patients with benign pancreatic disease. In vitro, the effect of pancreatic cancer–conditioned medium on mast cell migration was assessed. The effect of conditioned medium from the human mast cell line, LAD-2, on cancer and normal ductal cell proliferation was assessed by thymidine incorporation. Matrigel invasion assays were used to evaluate the effect of mast cell–conditioned medium on cancer cell invasion in the presence and absence of a matrix metalloproteinase inhibitor, GM6001.

Results: Mast cell infiltration was significantly increased in pancreatic cancer compared with normal pancreatic tissue (11.4 ± 6.7 versus 2.0 ± 1.4, P < 0.001). Increased infiltrating mast cells correlated with higher grade tumors (P < 0.0001) and worse survival. Patients with pancreatic cancer had elevated serum tryptase activity (P < 0.05). In vitro, AsPC1 and PANC-1 cells induced mast cell migration. Mast cell–conditioned medium induced pancreatic cancer cell migration, proliferation, and invasion but had no effect on normal ductal cells. Furthermore, the effect of mast cells on cancer cell invasion was, in large part, matrix metalloproteinase–dependent.

Conclusions: Tumor-infiltrating mast cells are associated with worse prognosis in pancreatic cancer. In vitro, the interaction between mast cells and pancreatic cancer cells promotes tumor growth and invasion. Clin Cancer Res; 16(8); 2257–65. ©2010 AACR.
Data from C-Raf Is Associated with Disease Progression and Cell Proliferation in a Subset of Melanomas
Authors: Jilaveanu Lucia B. , Zito Christopher R. , Aziz Saadia A. , Conrad Patricia J. , Schmitz John C. , Sznol Mario , Camp Robert L. , Rimm David L. , Kluger Harriet M.
Published: April 2023

Abstract
Purpose: Raf-kinases include three major isoforms. Although the role of B-Raf in melanoma is well established, little is known about C-Raf. We studied effects of C-Raf knockdown in vitro and assessed expression of C-Raf in a large cohort of melanomas and nevi.

Experimental Design: Using specific siRNAs, we knocked down C-Raf expression, and determined the effect on viability, MAP extracellular signal-regulated kinase (ERK)/ERK kinase signaling, and apoptosis in seven melanoma cell lines. We determined the IC50 of the C-Raf inhibitors sorafenib and GW5074, and studied the effects of GW5074 on cell signaling. Using an automated method to measure in situ protein expression, we quantified C-Raf expression in 263 nevi and 523 melanomas.

Results: C-Raf was knocked down in three cell lines with detectable phospho-C-Raf, resulting in decreased viability in two of the three (YULAC and YUROB). This resulted in decreased Bcl-2 expression and phospho-Bad cleavage, without affecting phospho-MEK and phospho-ERK. Sensitivity to sorafenib and GW5074 varied. GW5074 inhibited mitogen-activated protein kinase signaling without Bcl-2 and phospho-Bad down-regulation. C-Raf was highly expressed in melanomas compared with nevi (P < 0.0001), and no nevi had high C-Raf expression. C-Raf expression was higher in metastatic than primary specimens (P = 0.0225).

Conclusions: C-Raf siRNA knock-down results in decreased viability of YULAC (B-RafV600K) and YUROB (B-RafWT) melanoma cells, likely mediated by Bcl-2 inhibition rather than mitogen-activated protein kinase inhibition. Cotargeting C-Raf and parallel pathways might be an effective therapeutic approach for melanoma. C-Raf expression is up-regulated in a subset of melanomas but not in nevi, suggesting that it might be a valuable diagnostic marker and therapeutic target. (Clin Cancer Res 2009;15(18):5704–13)
Data from Helicobacter pylori Infection Induces Genetic Instability of Nuclear and Mitochondrial DNA in Gastric Cells
Authors: Machado Ana Manuel Dantas , Figueiredo Ceu , Touati Eliette , Máximo Valdemar , Sousa Sonia , Michel Valérie , Carneiro Fátima , Nielsen Finn Cilius , Seruca Raquel , Rasmussen Lene Juel
Published: April 2023

Abstract
Purpose:Helicobacter pylori is a major cause of gastric carcinoma. To investigate a possible link between bacterial infection and genetic instability of the host genome, we examined the effect of H. pylori infection on known cellular repair pathways in vitro and in vivo. Moreover, various types of genetic instabilities in the nuclear and mitochondrial DNA (mtDNA) were examined.

Experimental Design: We observed the effects of H. pylori infection on a gastric cell line (AGS), on C57BL/6 mice, and on individuals with chronic gastritis. In AGS cells, the effect of H. pylori infection on base excision repair and mismatch repair (MMR) was analyzed by reverse transcription-PCR, Western blot, and activity assays. In mice, MMR expression was analyzed by reverse transcription-PCR and the CA repeat instabilities were examined by Mutation Detection Enhancement gel electrophoresis. Mutation spectra in AGS cells and chronic gastritis tissue were determined by PCR, single-stranded conformation polymorphism, and sequencing. H. pylori vacA and cagA genotyping was determined by multiplex PCR and reverse hybridization.

Results: Following H. pylori infection, the activity and expression of base excision repair and MMR are down-regulated both in vitro and in vivo. Moreover, H. pylori induces genomic instability in nuclear CA repeats in mice and in mtDNA of AGS cells and chronic gastritis tissue, and this effect in mtDNA is associated with bacterial virulence.

Conclusions: Our results suggest that H. pylori impairs central DNA repair mechanisms, inducing a transient mutator phenotype, rendering gastric epithelial cells vulnerable to the accumulation of genetic instability and thus contributing to gastric carcinogenesis in infected individuals.
Data from Genetic and Expression Analysis of HER-2 and EGFR Genes in Salivary Duct Carcinoma: Empirical and Therapeutic Significance
Authors: Williams Michelle D. , Roberts Dianna B. , Kies Merrill S. , Mao Li , Weber Randal S. , El-Naggar Adel K.
Published: April 2023

Abstract
Purpose: Salivary duct carcinoma overexpresses epidermal growth factor receptor (EGFR) and HER-2, although the underlying mechanisms remain undefined. Because of the potential utilization of these markers as treatment targets, we evaluated protein and gene status by several techniques to determine complementary value.

Experimental Design: A tissue microarray of 66 salivary duct carcinomas was used for immunohistochemical analysis of HER-2 and EGFR expression (semiquantitatively evaluated into a three-tiered system), and fluorescence in situ hybridization for gene copy number, and chromosomes 7 and 17 ploidy status. Sequencing of exons 18, 19, and 21 of the EGFR gene for mutations was carried out.

Result: For EGFR, 46 (69.7%) of the 66 tumors showed some form of EGFR expression (17 at 3+, 17 at 2+, 12 at 1+) but none gene amplification. Five (9.4%) of 53 tumors showed mutations in exon 18 (n = 3) and exon 19 (n = 2). Polysomy of chromosome 7 (average >2.5 copies/cell) was detected in 15 (25.0%) of 60 tumors (6 at 3+, 5 at 2+, 2 at 1+, 2 at 0+ expression) and correlated with poor 3-year survival (P = 0.015). For HER-2, 17 (25.8%) of 66 tumors expressed HER-2 (10 at 3+, 3 at 2+, 4 at 1+). Eight tumors showed HER-2 gene amplification (6 at 3+, 1 at 1+, 1 at 0+ protein expression). Chromosome 17 polysomy was found in 8 (15.7%) of 51 tumors; two had HER-2 expression (3+, 1+).

Conclusion: Our study shows that salivary duct carcinomas (a) harbor EGFR gene mutations in a subset of tumors that may guide therapy, (b) pursue an aggressive clinical course in cases with chromosome 7 polysomy and high EGFR expression, and (c) with HER-2 gene amplification and protein high expression, may be selected for targeted therapy. Clin Cancer Res; 16(8); 2266–74. ©2010 AACR.
Data from Polyphenon E Inhibits the Growth of Human Barrett's and Aerodigestive Adenocarcinoma Cells by Suppressing Cyclin D1 Expression
Authors: Song Shumei , Krishnan Koyamangalath , Liu Kaifeng , Bresalier Robert S.
Published: April 2023

Abstract
Purpose: Green tea consumption has been shown to exhibit cancer-preventive activities in preclinical studies. Polyphenon E (Poly E) is a well-defined green tea–derived catechin mixture. This study was designed to determine the effects of Poly E on the growth of human Barrett's and aerodigestive adenocarcinoma cells and the mechanisms involved in growth regulation by this agent.

Experimental Design: Human adenocarcinoma cells and immortalized Barrett's epithelial cells were used as model systems.

Results: Poly E inhibited the proliferation of immortalized Barrett's cells as well as various adenocarcinoma cells, and this was associated with the down-regulation of cyclin D1 protein expression. Inhibition of cyclin D1 led to dephosphorylation of the retinoblastoma protein in a dose-dependent manner; these changes were associated with G1 cell cycle arrest. Poly E down-regulated cyclin D1 promoter activity and mRNA expression, suggesting transcriptional repression, and this correlated with decreased nuclear β-catenin and β-catenin/TCF4 transcriptional activity. MG132, an inhibitor of 26S proteosome, blocked the Poly E–induced down-regulation of cyclin D1, and Poly E promoted cyclin D1 polyubiquitination, suggesting that Poly E also inhibits cyclin D1 expression by promoting its degradation.

Conclusion: Poly E inhibits growth of transformed aerodigestive epithelial cells by suppressing cyclin D1 expression through both transcriptional and posttranslational mechanisms. These results provide insight into the mechanisms by which Poly E inhibits growth of Barrett's and adenocarcinoma cells, and provides a rationale for using this agent as a potential chemopreventive and therapeutic strategy for esophageal adenocarcinoma and its precursor, Barrett's esophagus.
Data from Association of MDM2 SNP309, Age of Onset, and Gender in Cutaneous Melanoma
Authors: Firoz Elnaz F. , Warycha Melanie , Zakrzewski Jan , Pollens Danuta , Wang Guimin , Shapiro Richard , Berman Russell , Pavlick Anna , Manga Prashiela , Ostrer Harry , Celebi Julide Tok , Kamino Hideko , Darvishian Farbod , Rolnitzky Linda , Goldberg Judith D. , Osman Iman , Polsky David
Published: April 2023

Abstract
Purpose: In certain cancers, MDM2 SNP309 has been associated with early tumor onset in women. In melanoma, incidence rates are higher in women than in men among individuals less than 40 years of age, but among those older than 50 years of age, melanoma is more frequent in men than in women. To investigate this difference, we examined the association among MDM2 SNP309, age at diagnosis, and gender among melanoma patients.

Experimental Design: Prospectively enrolled melanoma patients (N = 227) were evaluated for MDM2 SNP309 and the related polymorphism, p53 Arg72Pro. DNA was isolated from patient blood samples, and genotypes were analyzed by PCR-restriction fragment length polymorphism. Associations among MDM2 SNP309, p53 Arg72Pro, age at diagnosis, and clinicopathologic features of melanoma were analyzed.

Results: The median age at diagnosis was 13 years earlier among women with a SNP309 GG genotype (46 years) compared with women with TG+TT genotypes (59 years; P = 0.19). Analyses using age dichotomized at each decade indicated that women with a GG genotype had significantly higher risks of being diagnosed with melanoma at ages <50 years compared with women ≥50 years, but not when the comparison was made between women <60 and ≥60 years. At ages <50 years, women with a GG genotype had a 3.89 times greater chance of being diagnosed compared with women with TG+TT genotypes (P = 0.01). Similar observations were not seen among men.

Conclusions: Our data suggest that MDM2 may play an important role in the development of melanoma in women. The MDM2 SNP309 genotype may help identify women at risk of developing melanoma at a young age.
Data from Personalized Tamoxifen: A Step Closer but Miles To Go
Authors: Bardia Aditya , Stearns Vered
Published: April 2023

Abstract
Genetic variants in CYP2D6 lead to reduced conversion of tamoxifen to the active metabolite endoxifen. However, the role of the CYP2D6 genotype in predicting tamoxifen-associated outcomes remains controversial. Accurate assignment of the CYP2D6 genotype in archival tissues is crucial for future studies attempting to determine risk prediction of outcomes in tamoxifen-treated individuals. Clin Cancer Res; 16(17); 4308–10. ©2010 AACR.
Data from Chromosomal Instability in Microsatellite-Unstable and Stable Colon Cancer
Authors: Trautmann Karolin , Terdiman Jonathan P. , French Amy J. , Roydasgupta Ritu , Sein Nancy , Kakar Sanjay , Fridlyand Jane , Snijders Antoine M. , Albertson Donna G. , Thibodeau Stephen N. , Waldman Frederic M.
Published: April 2023

Abstract
Purpose: The genomic instability in colon cancer can be divided into at least two major types, microsatellite instability (MSI) or chromosomal instability (CIN). Although initially felt to be mutually exclusive, recent evidence suggests that there may be overlap between the two. The aim of this study was to identify chromosomal alterations at high resolution in sporadic colon cancers with high-level microsatellite instability (MSI-H) and to compare them to those present in a set of matched microsatellite stable (MSS) tumors.

Experimental Design: Array-based comparative genomic hybridization was used to analyze a set of 23 sporadic MSI-H and 23 MSS colon cancers matched for location, gender, stage, and age. The arrays consisted of 2,464 bacterial artificial chromosome clones.

Results: MSI and MSS colon cancers differed significantly with respect to frequency and type of chromosomal alterations. The median fraction of genome altered was lower among MSI-H tumors than MSS tumors (2.8% versus 30.7%, P = 0.00006). However, the MSI-H tumors displayed a range of genomic alterations, from the absence of detectable alterations to extensive alterations. Frequent alterations in MSI-H tumors included gains of chromosomes 8, 12, and 13, and loss of 15q14. In contrast, the most frequent alterations in MSS tumors were gains of 7, 13, 8q, and 20, and losses of 8p, 17p, and 18. A small, previously uncharacterized, genomic deletion on 16p13.2, found in 35% of MSI-H and 21% of MSS tumors, was confirmed by fluorescence in situ hybridization.

Conclusion: MSI and CIN are not mutually exclusive forms of genomic instability in sporadic colon cancer, with MSI tumors also showing varying degrees of CIN.
Data from Pharmacokinetic-Pharmacodynamic Relationships for the Heat Shock Protein 90 Molecular Chaperone Inhibitor 17-Allylamino, 17-Demethoxygeldanamycin in Human Ovarian Cancer Xenograft Models
Authors: Banerji Udai , Walton Michael , Raynaud Florence , Grimshaw Rachel , Kelland Lloyd , Valenti Melani , Judson Ian , Workman Paul
Published: April 2023

Abstract
Purpose: To establish the pharmacokinetic and pharmacodynamic profile of the heat shock protein 90 (HSP90) inhibitor 17-allylamino, 17-demethoxygeldanamycin (17-AAG) in ovarian cancer xenograft models.

Experimental Design: The effects of 17-AAG on growth inhibition and the expression of pharmacodynamic biomarkers c-RAF-1, CDK4, and HSP70 were studied in human ovarian cancer cell lines A2780 and CH1. Corresponding experiments were conducted with established tumor xenografts. The variability and specificity of pharmacodynamic markers in human peripheral blood lymphocytes (PBL) were studied.

Results: The IC50 values of 17-AAG in A2780 and CH1 cells were 18.3 nmol/L (SD, 2.3) and 410.1 nmol/L (SD, 9.4), respectively. Pharmacodynamic changes indicative of HSP90 inhibition were demonstrable at greater than or equal the IC50 concentration in both cell lines. Xenograft experiments confirmed tumor growth inhibition in vivo. Peak concentrations of 17-AAG achieved in A2780 and CH1 tumors were 15.6 and 16.5 μmol/L, respectively, and there was no significant difference between day 1 and 11 pharmacokinetic profiles. Reversible changes in pharmacodynamic biomarkers were shown in tumor and murine PBLs in both xenograft models. Expression of pharmacodynamic markers varied between human PBLs from different human volunteers but not within the same individual. Pharmacodynamic biomarker changes consistent with HSP90 inhibition were shown in human PBLs exposed ex vivo to 17-AAG but not to selected cytotoxic drugs.

Conclusion: Pharmacokinetic-pharmacodynamic relationships were established for 17-AAG. This information formed the basis of a pharmacokinetic-pharmacodynamic-driven phase I trial.
Data from Plasma Levels of Trefoil Factors are Increased in Patients with Advanced Prostate Cancer
Authors: Vestergaard Else Marie , Borre Michael , Poulsen Steen Seier , Nexø Ebba , Tørring Niels
Published: April 2023

Abstract
Purpose: Through cDNA array analyses and immunohistochemistry on tissue microarrays, trefoil factor 3 (TFF3) was recently shown to be overexpressed in prostate cancer. The purpose of this study was to test the feasibility of using the levels of trefoil factors as a plasma marker for prostate cancer.

Experimental Design: In 79 patients with prostate cancer, 23 patients with benign prostatic hyperplasia, and 44 healthy individuals plasma TFF1, TFF2, and TFF3 were determined with ELISAs and compared with clinical stage and prostate-specific antigen (PSA) values. Plasma levels of TFF were compared with the immunohistochemical expression of TFF and chromogranin A in 30 prostate cancer tissue samples.

Results: Patients with advanced prostate cancer had significantly higher plasma concentrations of TFF1, TFF2, and TFF3 (P < 0.01) compared with patients with localized disease. Using a cutoff of 200 pmol/L, the sensitivity and specificity of plasma TFF3 in differentiating between patients with localized and advanced disease was 74% (59-85%) and 81% (66-91%). Plasma levels of TFF3 were highest in patients with bone metastases (P = 0.008). Patients with serum PSA >10 μg/L had significantly higher plasma TFF3 values than patients with serum PSA <10 μg/L (P = 0.03) and TFF3 levels were higher in patients with Gleason sums of ≥7 (P = 0.02). Expression of TFF1 and TFF3 determined by immunohistochemistry was increased in patients with prostate cancer but did not correlate with plasma trefoil factor values.

Conclusions: Plasma levels of trefoil factors are increased in patients with advanced prostate cancer. Prospective studies are needed to confirm the predictive utility of trefoil factors in prostate cancer.
Data from Modeling the Pharmacodynamics of Highly Schedule-Dependent Agents: Exemplified by Cytarabine-Based Regimens in Acute Myeloid Leukemia
Authors: Braess Jan , Fiegl Michael , Lorenz Isolde , Waxenberger Karin , Hiddemann Wolfgang
Published: April 2023

Abstract
Background: Many agents in antineoplastic chemotherapy are highly schedule dependent. Therefore, variables such as total dose and also the area under the curve (AUC) that are schedule insensitive are generally insufficient to adequately represent treatment strength.

Purpose: To establish a descriptor of treatment strength that takes into account the differential contribution of plasma concentrations (C) and exposure times (T) towards the cytotoxic effect and to investigate whether such a pharmacodynamically weighed descriptor is better correlated to the clinical effect than conventional variables.

Patients and Methods: The paradigm “CN × T = constant” (for an isoeffect) incorporates a weighing factor N (concentration coefficient) into the conventional description of the AUC that quantitates the differential contribution of C and T towards the cytotoxic effect. N was to be numerically derived from a multitude of in vitro isoeffect analyses of the major agents in acute myeloid leukemia (AML) therapy from patient samples (n = 57).

Results: For cytarabine, N was 0.45, numerically expressing the substantially higher relevance of T versus C for its cytotoxic effect. In a meta-analysis of 49 study arms involving >10,000 patients, neither total dose, dose intensity, nor AUC was correlated to the clinical effect. However, when AUC was pharmacodynamically weighed (N-weighed AUC, N-AUC = C0.45 × T), this new descriptor was highly significantly correlated to the clinical effect (P < 0.001).

Conclusion: The N-AUC concept is able to characterize schedule-dependent agents and is the only descriptor of cytarabine treatment strength actually correlated to the clinical effect in AML.