bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Identification of Alternative Splicing Markers for Breast Cancer
Authors: Venables Julian P. , Klinck Roscoe , Bramard Anne , Inkel Lyna , Dufresne-Martin Geneviève , Koh ChuShin , Gervais-Bird Julien , Lapointe Elvy , Froehlich Ulrike , Durand Mathieu , Gendron Daniel , Brosseau Jean-Philippe , Thibault Philippe , Lucier Jean-Francois , Tremblay Karine , Prinos Panagiotis , Wellinger Raymund J. , Chabot Benoit , Rancourt Claudine , Elela Sherif Abou
Published: March 2024

Abstract
Breast cancer is the most common cause of cancer death among women under age 50 years, so it is imperative to identify molecular markers to improve diagnosis and prognosis of this disease. Here, we present a new approach for the identification of breast cancer markers that does not measure gene expression but instead uses the ratio of alternatively spliced mRNAs as its indicator. Using a high-throughput reverse transcription-PCR–based system for splicing annotation, we monitored the alternative splicing profiles of 600 cancer-associated genes in a panel of 21 normal and 26 cancerous breast tissues. We validated 41 alternative splicing events that significantly differed in breast tumors relative to normal breast tissues. Most cancer-specific changes in splicing that disrupt known protein domains support an increase in cell proliferation or survival consistent with a functional role for alternative splicing in cancer. In a blind screen, a classifier based on the 12 best cancer-associated splicing events correctly identified cancer tissues with 96% accuracy. Moreover, a subset of these alternative splicing events could order tissues according to histopathologic grade, and 5 markers were validated in a further blind set of 19 grade 1 and 19 grade 3 tumor samples. These results provide a simple alternative for the classification of normal and cancerous breast tumor tissues and underscore the putative role of alternative splicing in the biology of cancer. [Cancer Res 2008;68(22):9525–31]
Data from Type 2 Bias of T Cells Expanded from the Blood of Melanoma Patients Switched to Type 1 by IL-12p70 mRNA–Transfected Dendritic Cells
Authors: Minkis Kira , Kavanagh Daniel G. , Alter Galit , Bogunovic Dusan , O'Neill David , Adams Sylvia , Pavlick Anna , Walker Bruce D. , Brockman Mark A. , Gandhi Rajesh T. , Bhardwaj Nina
Published: March 2024

Abstract
Melanoma patients may exhibit a TH2-skewed cytokine profile within blood and tumor-infiltrating lymphocytes. Therapies that induce beneficial TH1-type tumor-specific immune responses, therefore, are highly desirable. Dendritic cells (DC) are widely used as immune adjuvants for cancer. Before their administration, DC are generally induced to mature with a cocktail of recombinant cytokines [interleukin (IL)-1β, tumor necrosis factor α, and IL-6] and prostaglandin E2 (PGE2), which is added to preserve the ability of DC to migrate to draining lymph nodes. However, PGE2 suppresses the production of IL-12p70, a cytokine essential for differentiation of TH1 responses. In this study, human DC were transfected with IL-12p70 mRNA and tested for their ability to alter the TH2 type bias manifested by blood T cells of patients with melanoma. Transfected DC secreted high levels of bioactive IL-12p70, as indicated by their capacity to enhance natural killer cell activity, skew TH1 responses in allogeneic mixed lymphocyte reactions through reduction of IL-4 and IL-5, and prime CD8+ T cells to the melanoma-associated antigen Melan A/MART-1. Furthermore, T-cell lines primed in vitro from the blood of melanoma patients showed strong type 2 skewing that was dramatically reversed by IL-12p70 transfection of autologous DC. Thus, IL-12p70 transfection of clinical DC preparations may enhance type 1 antitumor responses and may thereby contribute to effective immune-based therapy. [Cancer Res 2008;68(22):9441–50]
Data from PI3K Pathway Activation Mediates Resistance to MEK Inhibitors in KRAS Mutant Cancers
Authors: Wee Susan , Jagani Zainab , Xiang Kay Xiaoqin , Loo Alice , Dorsch Marion , Yao Yung-Mae , Sellers William R. , Lengauer Christoph , Stegmeier Frank
Published: March 2024

Abstract
The RAS pathway is one of the most frequently deregulated pathways in cancer. RAS signals through multiple effector pathways, including the RAF/mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK)/ERK MAPK and phosphatidylinositol 3-kinase (PI3K)-AKT signaling cascades. The oncogenic potential of these effector pathways is illustrated by the frequent occurrence of activating mutations in BRAF and PIK3CA as well as loss-of-function mutations in the tumor suppressor PTEN, a negative regulator of PI3K. Previous studies have found that whereas BRAF mutant cancers are highly sensitive to MEK inhibition, RAS mutant cancers exhibit a more variable response. The molecular mechanisms responsible for this heterogeneous response remain unclear. In this study, we show that PI3K pathway activation strongly influences the sensitivity of RAS mutant cells to MEK inhibitors. Activating mutations in PIK3CA reduce the sensitivity to MEK inhibition, whereas PTEN mutations seem to cause complete resistance. We further show that down-regulation of PIK3CA resensitizes cells with co-occurring KRAS and PIK3CA mutations to MEK inhibition. At the molecular level, the dual inhibition of both pathways seems to be required for complete inhibition of the downstream mammalian target of rapamycin effector pathway and results in the induction of cell death. Finally, we show that whereas inactivation of either the MEK or PI3K pathway leads to partial tumor growth inhibition, targeted inhibition of both pathways is required to achieve tumor stasis. Our study provides molecular insights that help explain the heterogeneous response of KRAS mutant cancers to MEK pathway inhibition and presents a strong rationale for the clinical testing of combination MEK and PI3K targeted therapies. [Cancer Res 2009;69(10):4286–93]
Data from Efficacy of Weekly Docetaxel and Bevacizumab in Mesenchymal Chondrosarcoma: A New Theranostic Method Combining Xenografted Biopsies with a Mathematical Model
Authors: Gorelik Boris , Ziv Irit , Shohat Revital , Wick Michael , Hankins W. David , Sidransky David , Agur Zvia
Published: March 2024

Abstract
The paucity of clinical treatment data on rare tumors, such as mesenchymal chondrosarcoma (MCS), emphasizes the need in theranostic tools for these diseases. We put forward and validated a new theranostic method, combining tumor xenografts and mathematical models, and used it to suggest an improved treatment schedule for a particular MCS patient. Growth curves and gene expression analysis of xenografts, derived from a patient's lung metastasis, served for creating a mathematical model of MCS progression and adapting it to the xenograft setting. The pharmacokinetics and pharmacodynamics of six drugs were modeled, with model variables being adjusted by patient-specific chemosensitivity tests. The xenografted animals were treated by various monotherapy and combination schedules, and the MCS xenograft model was computer simulated under the same treatment scenario. The mathematical model for xenograft growth was then up-scaled to retrieve the MCS patient's tumor progression under different treatment schedules. An average accuracy of 87.1% was obtained when comparing model predictions with the observed tumor growth inhibition in the xenografted animals. Simulation results suggested that a regimen containing bevacizumab applied i.v. in combination with once-weekly docetaxel would be more efficacious in the MCS patient than all other simulated schedules. Weekly docetaxel in the patient resulted in stable metastatic disease and relief of pancytopenia due to tumor infiltration. We suggest that the advantage of weekly docetaxel on the triweekly regimen is directly related to the angiogenesis rate of the tumor. Further validation of this conclusion, and the theranostic method we provide, may facilitate personalization of solid cancer pharmacotherapy. [Cancer Res 2008;68(21):9033–40]
Data from Genome-wide Allelic State Analysis on Flow-Sorted Tumor Fractions Provides an Accurate Measure of Chromosomal Aberrations
Authors: Corver Willem E. , Middeldorp Anneke , ter Haar Natalja T. , Jordanova Ekaterina S. , van Puijenbroek Marjo , van Eijk Ronald , Cornelisse Cees J. , Fleuren Gert Jan , Morreau Hans , Oosting Jan , van Wezel Tom
Published: March 2024

Abstract
Chromosomal aberrations are a common characteristic of cancer and are associated with copy number abnormalities and loss of heterozygosity (LOH). Tumor heterogeneity, low tumor cell percentage, and lack of knowledge of the DNA content impair the identification of these alterations especially in aneuploid tumors. To accurately detect allelic changes in carcinomas, we combined flow-sorting and single nucleotide polymorphism arrays. Cells derived from archival cervical and colon cancers were flow-sorted based on differential vimentin and keratin expression and DNA content and analyzed on single nucleotide polymorphism arrays. A new algorithm, the lesser allele intensity ratio, was used to generate a molecular measure of chromosomal aberrations for each case. Flow-sorting significantly improved the detection of copy number abnormalities; 31.8% showed an increase in amplitude and 23.2% were missed in the unsorted fraction, whereas 15.9% were detected but interpreted differently. Integration of the DNA index in the analysis enabled the identification of the allelic state of chromosomal aberrations, such as LOH ([A]), copy-neutral LOH ([AA]), balanced amplifications ([AABB]), and allelic imbalances ([AAB] or [AAAB], etc.). Chromosomal segments were sharply defined. Fluorescence in situ hybridization copy numbers, as well as the high similarity between the DNA index and the allelic state index, which is the average of the allelic states across the genome, validated the method. This new approach provides an individual molecular measure of chromosomal aberrations and will likely have repercussions for preoperative molecular staging, classification, and prognostic profiling of tumors, particularly for heterogeneous aneuploid tumors, and allows the study of the underlying molecular genetic mechanisms and clonal evolution of tumor subpopulations. [Cancer Res 2008;68(24):10333–40]
Data from Effects of Dasatinib on Src Kinase Activity and Downstream Intracellular Signaling in Primitive Chronic Myelogenous Leukemia Hematopoietic Cells
Authors: Konig Heiko , Copland Mhairi , Chu Su , Jove Richard , Holyoake Tessa L. , Bhatia Ravi
Published: March 2024

Abstract
Bcr-Abl tyrosine kinase inhibitors (TKI) are effective in inducing remissions in chronic myelogenous leukemia (CML) patients but do not eliminate primitive CML hematopoietic cells. There is a need to identify mechanisms that contribute to retention of CML progenitors. Src family tyrosine kinases have been identified as potential mediators of Bcr-Abl–induced leukemogenesis. Dasatinib (BMS-354825) is a potent dual Abl/Src kinase inhibitor approved for clinical use in CML patients. We evaluated Src activity in primitive human CML progenitors from different stages of disease and investigated effects of Dasatinib on Src activity and downstream signaling pathways. P-Src expression was increased in CD34+ cells and CD34+CD38− cells in all phases of CML. Dasatinib showed potent Src inhibitory activity in CML progenitors, inhibiting both Bcr-Abl–dependent and –independent Src activity. In contrast, Imatinib inhibited only Bcr-Abl–dependent Src activity. Dasatinib inhibited P–mitogen-activated protein kinase (MAPK), P-Akt, and P-STAT5 levels in CML progenitors in the absence of growth factors but not in the presence of growth factors. A marked increase in P-MAPK levels seen in the presence of growth factors with Imatinib was much less prominent with Dasatinib. Dasatinib significantly suppressed CML colony-forming cells and long-term culture-initiating cells but did not significantly alter the level of apoptosis-regulating proteins in CML CD34+ cells. Our results indicate that Dasatinib, in addition to potent anti–Bcr-Abl kinase activity, effectively inhibits Src kinase activity and downstream signaling pathways in CML progenitors but does not induce a strong proapoptotic response. These observations argue against a prominent role for Src kinases in persistence of primitive CML cells in TKI-treated patients. [Cancer Res 2008;68(23):9624–33]
Data from Activated Gα13 Impairs Cell Invasiveness through p190RhoGAP-Mediated Inhibition of RhoA Activity
Authors: Bartolomé Rubén A. , Wright Natalia , Molina-Ortiz Isabel , Sánchez-Luque Francisco J. , Teixidó Joaquin
Published: March 2024

Abstract
The GTPase RhoA is a downstream target of heterotrimeric G13 proteins and plays key roles in cell migration and invasion. Here, we show that expression in human melanoma cells of a constitutively active, GTPase-deficient Gα13 form (Gα13QL) or lysophosphatidylcholine (LPC)-promoted signaling through Gα13-coupled receptors led to a blockade of chemokine-stimulated RhoA activation and cell invasion that was rescued by active RhoA. Melanoma cells expressing Gα13QL or cells stimulated with LPC displayed an increase in p190RhoGAP activation, and defects in RhoA activation and invasion were recovered by knocking down p190RhoGAP expression, thus identifying this GTPase-activating protein (GAP) protein as a downstream Gα13 target that is responsible for these inhibitory responses. In addition, defective stress fiber assembly and reduced migration speed underlay inefficient invasion of Gα13QL melanoma cells. Importantly, Gα13QL expression in melanoma cells led to impairment in lung metastasis associated with prolonged survival in SCID mice. The data indicate that Gα13-dependent downstream effects on RhoA activation and invasion tightly depend on cell type–specific GAP activities and that Gα13-p190RhoGAP signaling might represent a potential target for intervention in melanoma metastasis. [Cancer Res 2008;68(20):8221–30]
Data from Platelet Granule Secretion Continuously Prevents Intratumor Hemorrhage
Authors: Ho-Tin-Noé Benoit , Goerge Tobias , Cifuni Stephen M. , Duerschmied Daniel , Wagner Denisa D.
Published: March 2024

Abstract
Cancer is associated with a prothrombogenic state capable of platelet activation. Platelets, on the other hand, can support angiogenesis, a process involved in the progression of tumor growth and metastasis. However, it is unclear whether platelet/tumor interactions substantially contribute to tumor physiology. We investigated whether platelets stabilize tumor vessels and studied the underlying mechanisms. We induced severe acute thrombocytopenia in mice bearing s.c. Lewis lung carcinoma or B16F10 melanoma. Intravital microscopy revealed that platelet depletion led to a rapid destabilization of tumor vessels with intratumor hemorrhage starting as soon as 30 min after induction of thrombocytopenia. Using an inhibitor of glycoprotein Ibα (GPIbα) and genetically engineered mice with platelet adhesion defects, we investigated the role of platelet adhesion receptors in stabilizing tumor vessels. We found that a single defect in either GPIbα, von Willebrand factor, P-selectin, or platelet integrin activation did not lead to intratumor hemorrhage. We then compared the ability of transfused resting and degranulated platelets to prevent intratumor hemorrhage. Whereas resting platelets prevented thrombocytopenia-induced tumor bleeding, circulating degranulated platelets did not. This suggests that the prevention of intratumor hemorrhage by platelets relies on the secretion of the content of platelet granules. Supporting this hypothesis, we further found that thrombocytopenia dramatically impairs the balance between propermeability and antipermeability factors in tumor-bearing animals, in particular depleting blood of angiopoietin-1 and serotonin. Our results show a crucial contribution of platelets to tumor homeostasis through continuous prevention of severe intratumor hemorrhage and consequent cell death. The study also suggests platelet function as a reasonable target for specific destabilization of tumor vessels. [Cancer Res 2008;68(16):6851–8]
Data from Hypoxia-Inducible Factor-2α Correlates to Distant Recurrence and Poor Outcome in Invasive Breast Cancer
Authors: Helczynska Karolina , Larsson Anna-Maria , Holmquist Mengelbier Linda , Bridges Esther , Fredlund Erik , Borgquist Signe , Landberg Göran , Påhlman Sven , Jirström Karin
Published: March 2024

Abstract
Differential regulation as well as target gene specificity of the two hypoxia-inducible factor (HIF)-α subunits HIF-1α and HIF-2α in various tumors and cell lines have been suggested. In breast cancer, the prognostic significance of HIF-1α is not clear-cut and that of HIF-2α is largely unknown. Using IHC analyses of HIF-1α, HIF-2α, and vascular endothelial growth factor (VEGF) expression in a tissue microarray of invasive breast cancer specimens from 512 patients, we investigated the expression patterns of the 2 HIF-α subunits in relation to established clinicopathologic variables, VEGF expression, and survival. HIF-1α and HIF-2α protein levels and their effect on survival were additionally analyzed in a second cohort of 179 patients. To evaluate the individual role of each subunit in the hypoxic response and induction of VEGF, HIF-α protein and HIF-α and VEGF mRNA levels were further studied in cultured breast cancer cells after hypoxic induction and/or knockdown of HIF-α subunits by siRNA by Western blot and Quantitative Real-Time PCR techniques. We showed that although HIF-1α and HIF-2α protein levels in breast cancer specimens were not interrelated, high levels of both HIF-1α and HIF-2α associated to high VEGF expression. HIF-2α expression was an independent prognostic factor associated to reduced recurrence-free and breast cancer–specific survival, whereas HIF-1α did not exhibit these correlations. In cultured cells, acute hypoxia induced both HIF-proteins. At prolonged hypoxia, HIF-2α remained accumulated, whereas HIF-1α protein levels decreased, in agreement with the oxygen level and time-dependent induction of HIFs recently reported in neuroblastoma. [Cancer Res 2008;68(22):9212–20]
Data from Illegitimate WNT Pathway Activation by β-Catenin Mutation or Autocrine Stimulation in T-Cell Malignancies
Authors: Groen Richard W.J. , Oud Monique E.C.M. , Schilder-Tol Esther J.M. , Overdijk Marije B. , ten Berge Derk , Nusse Roel , Spaargaren Marcel , Pals Steven T.
Published: March 2024

Abstract
Recent studies in mice have shown a role for the canonical WNT pathway in lymphocyte development. Because cancers often arise as a result of aberrant activation of signaling cascades that normally promote the self-renewal and expansion of their progenitor cells, we hypothesized that activation of the WNT pathway might contribute to the pathogenesis of lymphoproliferative disease. Therefore, we screened a large panel (n = 162) of non–Hodgkin lymphomas (NHL), including all major WHO categories, for nuclear expression of β-catenin, a hallmark of “active” WNT signaling. In 16 lymphomas, mostly of T-lineage origin, nuclear localization of β-catenin was detected. Interestingly, some of these tumors contained established gain-of-function mutations in the gene encoding β-catenin (CTNNB1); however, in the majority, mutations in either CTNNB1 or APC were not detected. Functional analysis of WNT signaling in precursor T-lymphoblastic lymphomas/leukemias, the NHL subset in which β-catenin accumulation was most prevalent (33% positive), revealed a constitutively activated, but still responsive, WNT pathway, which controlled T-cell factor–mediated gene transcription and cell growth. Our data indicate that activation of the WNT pathway, either by CTNNB1 mutation or autocrine stimulation, plays a role in the pathogenesis of a subset of NHLs, in particular, those of T-cell origin. [Cancer Res 2008;68(17):6969–77]
Data from Dietary Induction of Colonic Tumors in a Mouse Model of Sporadic Colon Cancer
Authors: Yang Kan , Kurihara Naoto , Fan Kunhua , Newmark Harold , Rigas Basil , Bancroft Laura , Corner Georgia , Livote Elayne , Lesser Martin , Edelmann Winfried , Velcich Anna , Lipkin Martin , Augenlicht Leonard
Published: March 2024

Abstract
A defined rodent “new Western diet” (NWD), which recapitulates intake levels of nutrients that are major dietary risk factors for human colon cancer, induced colonic tumors when fed to wild-type C57Bl/6 mice for 1.5 to 2 years from age 6 weeks (two-thirds of their life span). Colonic tumors were prevented by elevating dietary calcium and vitamin D3 to levels comparable with upper levels consumed by humans, but tumorigenesis was not altered by similarly increasing folate, choline, methionine, or fiber, each of which was also at the lower levels in the NWD that are associated with risk for colon cancer. The NWD significantly altered profiles of gene expression in the flat colonic mucosa that exhibited heterogeneity among the mice, but unsupervised clustering of the data and novel statistical analyses showed reprogramming of colonic epithelial cells in the flat mucosa by the NWD was similar to that initiated by inheritance of a mutant Apc allele. The NWD also caused general down-regulation of genes encoding enzymes involved in lipid metabolism and the tricarboxylic acid cycle in colonic epithelial cells before tumor formation, which was prevented by the supplementation of the NWD with calcium and vitamin D3 that prevented colon tumor development, demonstrating profound interaction among nutrients. This mouse model of dietary induction of colon cancer recapitulates levels and length of exposure to nutrients linked to relative risk for human sporadic colon cancer, which represents the etiology of >90% of colon cancer in the United States and other Western countries. [Cancer Res 2008;68(19):7803–10]
Data from Heparanase Augments Epidermal Growth Factor Receptor Phosphorylation: Correlation with Head and Neck Tumor Progression
Authors: Cohen-Kaplan Victoria , Doweck Ilana , Naroditsky Inna , Vlodavsky Israel , Ilan Neta
Published: March 2024

Abstract
Heparanase is an endoglycosidase that specifically cleaves heparan sulfate side chains, a class of glycosaminoglycans abundantly present in the extracellular matrix and on the cell surface. Heparanase activity is strongly implicated in tumor metastasis attributed to remodeling of the subepithelial and subendothelial basement membranes, resulting in dissemination of metastatic cancer cells. Moreover, heparanase up-regulation was noted in an increasing number of primary human tumors, correlating with tumors larger in size, increased microvessel density, and reduced postoperative survival rate, implying that heparanase function is not limited to tumor metastasis. This notion is supported by recent findings revealing induction of signaling molecules (i.e., Akt, p38) and gene transcription [i.e., tissue factor, vascular endothelial growth factor (VEGF)] by enzymatically-inactive heparanase. Here, we provide evidence that active and inactive heparanase proteins enhance epidermal growth factor receptor (EGFR) phosphorylation. Enhanced EGFR phosphorylation was associated with increased cell migration, cell proliferation, and colony formation, which were attenuated by Src inhibitors. Similarly, heparanase gene silencing by means of siRNA was associated with reduced Src and EGFR phosphorylation levels and decreased cell proliferation. Moreover, heparanase expression correlated with increased phospho-EGFR levels and progression of head and neck carcinoma, providing a strong clinical support for EGFR modulation by heparanase. Thus, heparanase seems to modulate two critical systems involved in tumor progression, namely VEGF expression and EGFR activation. Neutralizing heparanase enzymatic and nonenzymatic functions is therefore expected to profoundly affect tumor growth, angiogenesis, and metastasis. [Cancer Res 2008;68(24):10077–85
Data from Mutational Profile of Advanced Primary and Metastatic Radioactive Iodine-Refractory Thyroid Cancers Reveals Distinct Pathogenetic Roles for BRAF, PIK3CA, and AKT1
Authors: Ricarte-Filho Julio C. , Ryder Mabel , Chitale Dhananjay A. , Rivera Michael , Heguy Adriana , Ladanyi Marc , Janakiraman Manickam , Solit David , Knauf Jeffrey A. , Tuttle R. Michael , Ghossein Ronald A. , Fagin James A.
Published: March 2024

Abstract
Patients with poorly differentiated thyroid cancers (PDTC), anaplastic thyroid cancers (ATC), and radioactive iodine-refractory (RAIR) differentiated thyroid cancers have a high mortality, particularly if positive on [18F]fluorodeoxyglucose (FDG)-positron emission tomography (PET). To obtain comprehensive genetic information on advanced thyroid cancers, we designed an assay panel for mass spectrometry genotyping encompassing the most significant oncogenes in this disease: 111 mutations in RET, BRAF, NRAS, HRAS, KRAS, PIK3CA, AKT1, and other related genes were surveyed in 31 cell lines, 52 primary tumors (34 PDTC and 18 ATC), and 55 RAIR, FDG-PET-positive recurrences and metastases (nodal and distant) from 42 patients. RAS mutations were more prevalent than BRAF (44 versus 12%; P = 0.002) in primary PDTC, whereas BRAF was more common than RAS (39 versus 13%; P = 0.04) in PET-positive metastatic PDTC. BRAF mutations were highly prevalent in ATC (44%) and in metastatic tumors from RAIR PTC patients (95%). Among patients with multiple metastases, 9 of 10 showed between-sample concordance for BRAF or RAS mutations. By contrast, 5 of 6 patients were discordant for mutations of PIK3CA or AKT1. AKT1_G49A was found in 9 specimens, exclusively in metastases. This is the first documentation of AKT1 mutation in thyroid cancer. Thus, RAIR, FDG-PET–positive metastases are enriched for BRAF mutations. If BRAF is mutated in the primary, it is likely that the metastases will harbor the defect. By contrast, absence of PIK3CA/AKT1 mutations in one specimen may not reflect the status at other sites because these mutations arise during progression, an important consideration for therapies directed at phosphoinositide 3-kinase effectors. [Cancer Res 2009;69(11):4885–93]
Data from K-ras and Wnt Signaling Synergize to Accelerate Prostate Tumorigenesis in the Mouse
Authors: Pearson Helen B. , Phesse Toby J. , Clarke Alan R.
Published: March 2024

Abstract
Aberrant Ras and Wnt signaling are emerging as key events in the multistep nature of prostate tumorigenesis and progression. Here, we report the generation of a compound model of prostate cancer to define the synergism of activated K-ras (K-ras+/V12) and dominant stabilized β-catenin (Catnb+/lox(ex3)) in the murine prostate. Recombination of floxed alleles and subsequent expression of oncogenic transgenes was mediated by Cre recombinase expression governed by the composite Probasin (PB) promoter (termed PBCre). Concomitant with elevated mitogen-activated protein kinase (MAPK) signaling, PBCre+K-ras+/V12 mice developed AH at 100 days (100% incidence) and low-grade prostate intraepithelial neoplasia and adenocarcinoma (60% and 7% incidence) by 500 days. PBCre+Catnb+/lox(ex3) mice showed reduced longevity (average 428 days) and were predisposed to PIN-like keratinized squamous metaplasia at 100 days (100% incidence) and adenocarcinoma (100% incidence) at end-point. These lesions displayed elevated Wnt signaling and basal levels of MAPK signaling. Synchronous activation of K-ras and β-catenin significantly reduced survival (average 189 days), reflecting accelerated tumorigenesis and the development of invasive carcinoma that displayed activated Wnt and MAPK signaling. Notably, expression of the basal cell marker p63 negatively correlated with tumor grade, resembling human prostate adenocarcinoma. Taken together, our data show that combinatorial oncogenic mutations of K-ras and β-catenin drive rapid progression of prostate tumorigenesis to invasive carcinoma, characterized by the synergistic elevation of androgen receptor, cyclooxygenase-2, and c-Myc. [Cancer Res 2009;69(1):94–101]
Data from Sperm-Associated Antigen 9 Is Associated With Tumor Growth, Migration, and Invasion in Renal Cell Carcinoma
Authors: Garg Manoj , Kanojia Deepika , Khosla Aashima , Dudha Namrata , Sati Satish , Chaurasiya Dipak , Jagadish Nirmala , Seth Amlesh , Kumar Rajive , Gupta Samir , Gupta Anju , Lohiya Nirmal Kumar , Suri Anil
Published: March 2024

Abstract
Renal cell carcinoma (RCC) represents one of the most resistant tumors to radiation and chemotherapy. Current therapies for RCC patients are inefficient due to the lack of diagnostic and therapeutic markers. Our recent studies have suggested an association of sperm-associated antigen 9 (SPAG9) with ovarian carcinomas. In the present study, we investigated the clinical relevance of SPAG9 in RCC patients. RT-PCR analysis showed expression of SPAG9 transcript in RCC tissues and RCC cell lines. In situ RNA hybridization and immunohistochemistry analyses confirmed the expression of SPAG9 in 88% of cancer patients, suggesting that SPAG9 participates in renal cancer. In addition, immunoblotting and ELISA analyses revealed a humoral immune response against SPAG9 in the sera of RCC patients but not in healthy individuals. Consistent with the clinical findings, knockdown of SPAG9 expression in RCC cells with specific siRNA significantly reduced cell growth and colony formation. Using in vitro wound healing and Matrigel invasion assays, we found that cell migration and invasive ability were also significantly inhibited. Furthermore, in vivo xenograft studies in nude mice revealed that administration of a SPAG9 siRNA plasmid significantly inhibited tumor growth. In conclusion, SPAG9 expression is associated with clinicopathologic features of tumors, suggesting that SPAG9 could contribute to the early spread of cancer. These results indicate that SPAG9 may have a role in tumor development and metastasis and thus could serve as a novel target for early detection and treatment of RCC. [Cancer Res 2008;68(20):8240–8]
Data from A Novel Association between p130Cas and Resistance to the Chemotherapeutic Drug Adriamycin in Human Breast Cancer Cells
Authors: Ta Huy Q. , Thomas Keena S. , Schrecengost Randy S. , Bouton Amy H.
Published: March 2024

Abstract
Resistance to chemotherapy remains a major obstacle for the treatment of breast cancer. Understanding the molecular mechanism(s) of resistance is crucial for the development of new effective therapies to treat this disease. This study examines the putative role of p130Cas (Cas) in resistance to the cytotoxic agent Adriamycin. High expression of Cas in primary breast tumors is associated with the failure to respond to the antiestrogen tamoxifen and poor prognosis, highlighting the potential clinical importance of this molecule. Here, we show a novel association between Cas and resistance to Adriamycin. We show that Cas overexpression renders MCF-7 breast cancer cells less sensitive to the growth inhibitory and proapoptotic effects of Adriamycin. The catalytic activity of the nonreceptor tyrosine kinase c-Src, but not the epidermal growth factor receptor, is critical for Cas-mediated protection from Adriamycin-induced death. The phosphorylation of Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) is elevated in Cas-overexpressing cells treated with Adriamycin, whereas expression of the proapoptotic protein Bak is decreased. Conversely, Cas depletion in the more resistant T47D and MDA-MB-231 cell lines increases sensitivity to Adriamycin. Based on these data, we propose that Cas activates growth and survival pathways regulated by c-Src, Akt, and ERK1/2 that lead to the inhibition of mitochondrial-mediated apoptosis in the presence of Adriamycin. Because Cas is frequently expressed at high levels in breast cancers, these findings raise the possibility of resensitizing Cas-overexpressing tumors to chemotherapy through perturbation of Cas signaling pathways. [Cancer Res 2008;68(21):8796–804]
Data from Netrin-1 Induces Apoptosis in Human Cervical Tumor Cells via the TAp73α Tumor Suppressor
Authors: Roperch Jean-Pierre , El Ouadrani Karima , Hendrix Ann , Emami Shahin , De Wever Olivier , Melino Gerry , Gespach Christian
Published: March 2024

Abstract
Netrins and their receptors deleted in colon cancer (DCC), neogenin, UNC5, and integrins are involved in axon guidance, epithelial morphogenesis, vascular pattering, cancer cell survival, invasion, tumor angiogenesis, and metastasis. Here, we considered the possible contribution of the p53-related apoptosis mediators p63 and p73 in the mechanisms underlying the antagonism between netrin-1 and DCC at the cell death control. We have showed that ectopic expression and external addition of netrin-1 in HeLa and HEK-293 cells with inactive p53 lead to impaired cell viability and induction of apoptosis. These responses were associated with up-regulation of the proapoptotic protein TAp73α, decreased Bcl-2/Bax ratio, and caspase-3 cleavage, with no change in protein levels of the antiapoptotic NH2-terminal–truncated ΔNp73α isoform, p73 adapter Yap-1 and p73 E3 ubiquitin ligase Itch, and p63, as well as the transcripts encoding p63, TAp73α, and ΔNp73α. However, the proteasome inhibitor MG132 potentiated, while DCC counteracted, netrin-1–induced TAp73α. Consistently, netrin-1 expression correlated with stabilization of the TAp73α protein and lower levels of TAp73α ubiquitination that was conversely enhanced by DCC, in a netrin-dependent manner. Our data indicate that netrin-1 selectively up-regulates TAp73α by preventing its ubiquitination and degradation. Targeted repression of p73α by shRNA reversed TAp73α and the apoptosis induced by netrin-1, and exacerbated the growth of HeLa tumor xenografts. Apoptosis induced by cisplatin was markedly enhanced in netrin-1 or DCC-expressing cells. Collectively, our data reveal that the transcriptionally active TAp73α tumor suppressor is implicated in the apoptosis induced by netrin-1 in a p53-independent and DCC/ubiquitin-proteasome dependent manner. [Cancer Res 2008;68(20):8231–9]
Data from Antigen Presented by Tumors In vivo Determines the Nature of CD8+ T-Cell Cytotoxicity
Authors: Shanker Anil , Brooks Alan D. , Jacobsen Kristen M. , Wine John W. , Wiltrout Robert H. , Yagita Hideo , Sayers Thomas J.
Published: March 2024

Abstract
The biological relevance of the perforin and Fas ligand (FasL) cytolytic pathways of CD8+ T lymphocytes (CTL) for cancer immunotherapy is controversial. We investigated the importance of these pathways in a murine renal cell carcinoma expressing influenza viral hemagglutinin as a defined surrogate antigen (Renca-HA). Following Renca-HA injection, all FasL-dysfunctional FasLgld/gld mice (n = 54) died from Renca-HA tumors by day 62. By contrast, perforin−/− (51%; n = 45) and Faslpr/lpr (55%; n = 51) mice remained tumor-free at day 360. Blocking FasL in vivo inhibited tumor rejection in these mice. Moreover, established Renca-HA tumors were cleared more efficiently by adoptively transferred HA518-526–specific T-cell receptor–transgenic CTL using FasL rather than perforin. Strikingly, a range of mouse tumor cells presenting low concentrations of immunogenic peptide were all preferentially lysed by the FasL but not the Pfp-mediated effector pathway of CTL, whereas at higher peptide concentrations, the preference in effector pathway usage by CTL was lost. Interestingly, a number of human renal cancer lines were also susceptible to FasL-mediated cytotoxicity. Therefore, the FasL cytolytic pathway may be particularly important for eradicating Fas-sensitive tumors presenting low levels of MHC class I–associated antigens following adoptive T-cell therapy. [Cancer Res 2009;69(16):6615–23]
Data from Pomalidomide and Lenalidomide Induce p21WAF-1 Expression in Both Lymphoma and Multiple Myeloma through a LSD1-Mediated Epigenetic Mechanism
Authors: Escoubet-Lozach Laure , Lin I-Lin , Jensen-Pergakes Kristen , Brady Helen A. , Gandhi Anita K. , Schafer Peter H. , Muller George W. , Worland Peter J. , Chan Kyle W.H. , Verhelle Dominique
Published: March 2024

Abstract
Lenalidomide and pomalidomide have both been evaluated clinically for their properties as anticancer agents, with lenalidomide being available commercially. We previously reported that both compounds cause cell cycle arrest in Burkitt's lymphoma and multiple myeloma cell lines by increasing the level of p21WAF-1 expression. In the present study, we unravel the molecular mechanism responsible for p21WAF-1 up-regulation using Namalwa cells as a human lymphoma model. We show that the increase of p21WAF-1 expression is regulated at the transcriptional level through a mechanism independent of p53. Using a combination of approaches, we show that several GC-rich binding transcription factors are involved in pomalidomide-mediated up-regulation of p21WAF-1. Furthermore, we report that p21WAF-1 up-regulation is associated with a switch from methylated to acetylated histone H3 on p21WAF-1 promoter. Interestingly, lysine-specific demethylase-1 (LSD1) silencing reduced both pomalidomide and lenalidomide up-regulation of p21WAF-1, suggesting that this histone demethylase is involved in the priming of the p21WAF-1 promoter. Based on our findings, we propose a model in which pomalidomide and lenalidomide modify the chromatin structure of the p21WAF-1 promoter through demethylation and acetylation of H3K9. This effect, mediated via LSD1, provides GC-rich binding transcription factors better access to DNA, followed by recruitment of RNA polymerase II and transcription activation. Taken together, our results provide new insights on the mechanism of action of pomalidomide and lenalidomide in the regulation of gene transcription, imply possible efficacy in p53 mutated and deleted cancer, and suggest new potential clinical uses as an epigenetic therapy. [Cancer Res 2009;69(18):7347–56]
Data from Tgfbr1 Haploinsufficiency Is a Potent Modifier of Colorectal Cancer Development
Authors: Zeng Qinghua , Phukan Sharbani , Xu Yanfei , Sadim Maureen , Rosman Diana S. , Pennison Michael , Liao Jie , Yang Guang-Yu , Huang Chiang-Ching , Valle Laura , Di Cristofano Antonio , de la Chapelle Albert , Pasche Boris
Published: March 2024

Abstract
Transforming growth factor-β (TGF-β) signaling is frequently altered in colorectal cancer. Using a novel model of mice heterozygous for a targeted null mutation of Tgfbr1 crossed with ApcMin/+ mice, we show that ApcMin/+;Tgfbr1+/− mice develop twice as many intestinal tumors as ApcMin/+;Tgfbr1+/+ mice, as well as adenocarcinoma of the colon, without loss of heterozygosity at the Tgfbr1 locus. Decreased Smad2 and Smad3 phosphorylation and increased cellular proliferation are observed in the colonic epithelium crypts of ApcMin/+; Tgfbr1+/− mice. Smad-mediated TGF-β signaling is preserved in both ApcMin/+;Tgfbr1+/+ and ApcMin/+;Tgfbr1+/− intestinal tumors, but cyclin D1 expression and cellular proliferation are significantly higher in ApcMin/+;Tgfbr1+/− tumors. These results show that constitutively reduced Tgfbr1-mediated TGF-β signaling significantly enhances colorectal cancer development and results in increased tumor cell proliferation. These findings provide a plausible molecular mechanism for colorectal cancer development in individuals with constitutively altered TGFBR1 expression, a recently identified common form of human colorectal cancer. [Cancer Res 2009;69(2):678–86]
Data from Xenoestrogen-Induced Epigenetic Repression of microRNA-9-3 in Breast Epithelial Cells
Authors: Hsu Pei-Yin , Deatherage Daniel E. , Rodriguez Benjamin A.T. , Liyanarachchi Sandya , Weng Yu-I , Zuo Tao , Liu Joseph , Cheng Alfred S.L. , Huang Tim H-M.
Published: March 2024

Abstract
Early exposure to xenoestrogens may predispose to breast cancer risk later in adult life. It is likely that long-lived, self-regenerating epithelial progenitor cells are more susceptible to these exposure injuries over time and transmit the injured memory through epigenetic mechanisms to their differentiated progeny. Here, we used progenitor-containing mammospheres as an in vitro exposure model to study this epigenetic effect. Expression profiling identified that, relative to control cells, 9.1% of microRNAs (82 of 898 loci) were altered in epithelial progeny derived from mammospheres exposed to a synthetic estrogen, diethylstilbestrol. Repressive chromatin marks, trimethyl Lys27 of histone H3 (H3K27me3) and dimethyl Lys9 of histone H3 (H3K9me2), were found at a down-regulated locus, miR-9-3, in epithelial cells preexposed to diethylstilbestrol. This was accompanied by recruitment of DNA methyltransferase 1 that caused an aberrant increase in DNA methylation of its promoter CpG island in mammosphere-derived epithelial cells on diethylstilbestrol preexposure. Functional analyses suggest that miR-9-3 plays a role in the p53-related apoptotic pathway. Epigenetic silencing of this gene, therefore, reduces this cellular function and promotes the proliferation of breast cancer cells. Promoter hypermethylation of this microRNA may be a hallmark for early breast cancer development, and restoration of its expression by epigenetic and microRNA-based therapies is another viable option for future treatment of this disease. [Cancer Res 2009;69(14):5936–45]
Data from Plasminogen Kringle 5 Induces Apoptosis of Brain Microvessel Endothelial Cells: Sensitization by Radiation and Requirement for GRP78 and LRP1
Authors: McFarland Braden C. , Stewart Jerry , Hamza Amal , Nordal Robert , Davidson Donald J. , Henkin Jack , Gladson Candece L.
Published: March 2024

Abstract
Recombinant plasminogen kringle 5 (rK5) has been shown to induce apoptosis of dermal microvessel endothelial cells (MvEC) in a manner that requires glucose-regulated protein 78 (GRP78). As we are interested in antiangiogenic therapy for glioblastoma tumors, and the effectiveness of antiangiogenic therapy can be enhanced when combined with radiation, we investigated the proapoptotic effects of rK5 combined with radiation on brain MvEC. We found that rK5 treatment of brain MvEC induced apoptosis in a dose- and time-dependent manner and that prior irradiation significantly sensitized (500-fold) the cells to rK5-induced apoptosis. The rK5-induced apoptosis of both unirradiated and irradiated MvEC required expression of GRP78 and the low-density lipoprotein receptor-related protein 1 (LRP1), a scavenger receptor, based on down-regulation studies with small interfering RNA, and blocking studies with either a GRP78 antibody or a competitive inhibitor of ligand binding to LRP1. Furthermore, p38 mitogen-activated protein kinase was found to be a necessary downstream effector for rK5-induced apoptosis. These data suggest that irradiation sensitizes brain MvEC to the rK5-induced apoptosis and that this signal requires LRP1 internalization of GRP78 and the activation of p38 mitogen-activated protein kinase. Our findings suggest that prior irradiation would have a dose-sparing effect on rK5 antiangiogenic therapy for brain tumors and further suggest that the effects of rK5 would be tumor specific, as the expression of GRP78 protein is up-regulated on the brain MvEC in glioblastoma tumor biopsies compared with the normal brain. [Cancer Res 2009;69(13):5537–45]
Data from Immunity to Murine Prostatic Tumors: Continuous Provision of T-Cell Help Prevents CD8 T-Cell Tolerance and Activates Tumor-Infiltrating Dendritic Cells
Authors: Shafer-Weaver Kimberly A. , Watkins Stephanie K. , Anderson Michael J. , Draper Lauren J. , Malyguine Anatoli , Alvord W. Gregory , Greenberg Norman M. , Hurwitz Arthur A.
Published: March 2024

Abstract
We reported previously that tumor-specific CD8+ T cells (TcR-I) become tolerant in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. In this study, we show that CD4+ TcR transgenic (TcR-II) T cells transferred into TRAMP mice became activated in lymph nodes, trafficked to the prostate, and initially functioned as TH1 cells. Although a single cotransfer of TcR-II cells delayed TcR-I cell tolerization, repeated transfer of TcR-II cells was required to prevent TcR-I cell tolerization and significantly slowed progression of TRAMP prostate tumors. After transfer of TcR-II cells, dendritic cells within the tumor expressed higher levels of costimulatory molecules and displayed an enhanced ability to stimulate proliferation of naive T cells. Blockade of CD40-CD40L interactions during TcR-II transfer resulted in a profound reduction in dendritic cell stimulatory capacity and a partial loss of TcR-I effector functions and tumor immunity. These data show that sustained provision of activated tumor-specific CD4+ T cells alters the immunosuppressive tumor microenvironment, ultimately leading to the control of tumor growth. These findings will assist in the design of more effective immunotherapeutic approaches for cancer. [Cancer Res 2009;69(15):6256–64]
Data from Metformin Disrupts Crosstalk between G Protein–Coupled Receptor and Insulin Receptor Signaling Systems and Inhibits Pancreatic Cancer Growth
Authors: Kisfalvi Krisztina , Eibl Guido , Sinnett-Smith James , Rozengurt Enrique
Published: March 2024

Abstract
Recently, we identified a novel crosstalk between insulin and G protein–coupled receptor (GPCR) signaling pathways in human pancreatic cancer cells. Insulin enhanced GPCR signaling through a rapamycin-sensitive mTOR-dependent pathway. Metformin, the most widely used drug in the treatment of type 2 diabetes, activates AMP kinase (AMPK), which negatively regulates mTOR. Here, we determined whether metformin disrupts the crosstalk between insulin receptor and GPCR signaling in pancreatic cancer cells. Treatment of human pancreatic cancer cells (PANC-1, MIAPaCa-2, and BxPC-3) with insulin (10 ng/mL) for 5 minutes markedly enhanced the increase in intracellular [Ca2+] induced by GPCR agonists (e.g., neurotensin, bradykinin, and angiotensin II). Metformin pretreatment completely abrogated insulin-induced potentiation of Ca2+ signaling but did not interfere with the effect of GPCR agonists alone. Insulin also enhanced GPCR agonist–induced growth, measured by DNA synthesis, and the number of cells cultured in adherent or nonadherent conditions. Low doses of metformin (0.1–0.5 mmol/L) blocked the stimulation of DNA synthesis, and the anchorage-dependent and anchorage-independent growth induced by insulin and GPCR agonists. Treatment with metformin induced striking and sustained increase in the phosphorylation of AMPK at Thr172 and a selective AMPK inhibitor (compound C, at 5 μmol/L) reversed the effects of metformin on [Ca2+]i and DNA synthesis, indicating that metformin acts through AMPK activation. In view of these results, we tested whether metformin inhibits pancreatic cancer growth. Administration of metformin significantly decreased the growth of MIAPaCa-2 and PANC-1 cells xenografted on the flank of nude mice. These results raise the possibility that metformin could be a potential candidate in novel treatment strategies for human pancreatic cancer. [Cancer Res 2009;69(16):6539–45]