bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Novel STAT3 Phosphorylation Inhibitors Exhibit Potent Growth-Suppressive Activity in Pancreatic and Breast Cancer Cells
Authors: Lin Li , Hutzen Brian , Zuo Mingxin , Ball Sarah , Deangelis Stephanie , Foust Elizabeth , Pandit Bulbul , Ihnat Michael A. , Shenoy Satyendra S. , Kulp Samuel , Li Pui-Kai , Li Chenglong , Fuchs James , Lin Jiayuh
Published: March 2024

Abstract
The constitutive activation of signal transducer and activator of transcription 3 (STAT3) is frequently detected in most types of human cancer where it plays important roles in survival, drug resistance, angiogenesis, and other functions. Targeting constitutive STAT3 signaling is thus an attractive therapeutic approach for these cancers. We have recently developed novel small-molecule STAT3 inhibitors, known as FLLL31 and FLLL32, which are derived from curcumin (the primary bioactive compound of turmeric). These compounds are designed to bind selectively to Janus kinase 2 and the STAT3 Src homology-2 domain, which serve crucial roles in STAT3 dimerization and signal transduction. Here we show that FLLL31 and FLLL32 are effective inhibitors of STAT3 phosphorylation, DNA-binding activity, and transactivation in vitro, leading to the impediment of multiple oncogenic processes and the induction of apoptosis in pancreatic and breast cancer cell lines. FLLL31 and FLLL32 also inhibit colony formation in soft agar and cell invasion and exhibit synergy with the anticancer drug doxorubicin against breast cancer cells. In addition, we show that FLLL32 can inhibit the induction of STAT3 phosphorylation by IFNα and interleukin-6 in breast cancer cells. We also show that administration of FLLL32 can inhibit tumor growth and vascularity in chicken embryo xenografts as well as substantially reduce tumor volumes in mouse xenografts. Our findings highlight the potential of these new compounds and their efficacy in targeting pancreatic and breast cancers that exhibit constitutive STAT3 signaling. Cancer Res; 70(6); 2445–54
Data from A Susceptibility Locus on Chromosome 6q Greatly Increases Lung Cancer Risk among Light and Never Smokers
Authors: Amos Christopher I. , Pinney Susan M. , Li Yafang , Kupert Elena , Lee Juwon , de Andrade Mariza A. , Yang Ping , Schwartz Ann G. , Fain Pam R. , Gazdar Adi , Minna John , Wiest Jonathan S. , Zeng Dong , Rothschild Henry , Mandal Diptasri , You Ming , Coons Teresa , Gaba Colette , Bailey-Wilson Joan E. , Anderson Marshall W.
Published: March 2024

Abstract
Cigarette smoking is the major cause for lung cancer, but genetic factors also affect susceptibility. We studied families that included multiple relatives affected by lung cancer. Results from linkage analysis showed strong evidence that a region of chromosome 6q affects lung cancer risk. To characterize the effects that this region of chromosome 6q region has on lung cancer risk, we identified a haplotype that segregated with lung cancer. We then performed Cox regression analysis to estimate the differential effects that smoking behaviors have on lung cancer risk according to whether each individual carried a risk-associated haplotype or could not be classified and was assigned unknown haplotypic status. We divided smoking exposures into never smokers, light smokers (<20 pack-years), moderate smokers (20 to <40 pack-years), and heavy smokers (≥40 pack-years). Comparing results according to smoking behavior stratified by carrier status, compared with never smokers, there was weakly increasing risk for increasing smoking behaviors, with the hazards ratios being 3.44, 4.91, and 5.18, respectively, for light, moderate, or heavy smokers, whereas among the individuals from families without the risk haplotype, the risks associated with smoking increased strongly with exposure, the hazards ratios being, respectively, 4.25, 9.17, and 11.89 for light, moderate, and heavy smokers. The never smoking carriers had a 4.71-fold higher risk than the never smoking individuals without known risk haplotypes. These results identify a region of chromosome 6q that increases risk for lung cancer and that confers particularly higher risks to never and light smokers. Cancer Res; 70(6); 2359–67
Data from An Ets2-Driven Transcriptional Program in Tumor-Associated Macrophages Promotes Tumor Metastasis
Authors: Zabuawala Tahera , Taffany David A. , Sharma Sudarshana M. , Merchant Anand , Adair Brett , Srinivasan Ruchika , Rosol Thomas J. , Fernandez Soledad , Huang Kun , Leone Gustavo , Ostrowski Michael C.
Published: March 2024

Abstract
Tumor-associated macrophages (TAM) are implicated in breast cancer metastasis, but relatively little is known about the underlying genes and pathways that are involved. The transcription factor Ets2 is a direct target of signaling pathways involved in regulating macrophage functions during inflammation. We conditionally deleted Ets in TAMs to determine its function at this level on mouse mammary tumor growth and metastasis. Ets2 deletion in TAMs decreased the frequency and size of lung metastases in three different mouse models of breast cancer metastasis. Expression profiling and chromatin immunoprecipitation assays in isolated TAMs established that Ets2 repressed a gene program that included several well-characterized inhibitors of angiogenesis. Consistent with these results, Ets2 ablation in TAMs led to decreased angiogenesis and decreased growth of tumors. An Ets2-TAM expression signature consisting of 133 genes was identified within human breast cancer expression data which could retrospectively predict overall survival of patients with breast cancer in two independent data sets. In summary, we identified Ets2 as a central driver of a transcriptional program in TAMs that acts to promote lung metastasis of breast tumors. Cancer Res; 70(4); 1323–33
Data from Interleukin-8 Mediates Resistance to Antiangiogenic Agent Sunitinib in Renal Cell Carcinoma
Authors: Huang Dan , Ding Yan , Zhou Ming , Rini Brian I. , Petillo David , Qian Chao-Nan , Kahnoski Richard , Futreal P. Andrew , Furge Kyle A. , Teh Bin Tean
Published: March 2024

Abstract
The broad spectrum kinase inhibitor sunitinib is a first-line therapy for advanced clear cell renal cell carcinoma (ccRCC), a deadly form of kidney cancer. Unfortunately, most patients develop sunitinib resistance and progressive disease after about 1 year of treatment. In this study, we evaluated the mechanisms of resistance to sunitinib to identify the potential tactics to overcome it. Xenograft models were generated that mimicked clinical resistance to sunitinib. Higher microvessel density was found in sunitinib-resistant tumors, indicating that an escape from antiangiogenesis occurred. Notably, escape coincided with increased secretion of interleukin-8 (IL-8) from tumors into the plasma, and coadministration of an IL-8 neutralizing antibody resensitized tumors to sunitinib treatment. In patients who were refractory to sunitinib treatment, IL-8 expression was elevated in ccRCC tumors, supporting the concept that IL-8 levels might predict clinical response to sunitinib. Our results reveal IL-8 as an important contributor to sunitinib resistance in ccRCC and a candidate therapeutic target to reverse acquired or intrinsic resistance to sunitinib in this malignancy. Cancer Res; 70(3); 1063–71
Data from Prolactin Inhibits BCL6 Expression in Breast Cancer through a Stat5a-Dependent Mechanism
Authors: Tran Thai H. , Utama Fransiscus E. , Lin Justin , Yang Ning , Sjolund Ashley B. , Ryder Amy , Johnson Kevin J. , Neilson Lynn M. , Liu Chengbao , Brill Kristin L. , Rosenberg Anne L. , Witkiewicz Agnieszka K. , Rui Hallgeir
Published: March 2024

Abstract
BCL6 is a transcriptional repressor that recognizes DNA target sequences similar to those recognized by signal transducer and activator of transcriptions 5 (Stat5). BCL6 disrupts differentiation of breast epithelia, is downregulated during lactation, and is upregulated in poorly differentiated breast cancer. In contrast, Stat5a mediates prolactin-induced differentiation of mammary epithelia, and loss of Stat5 signaling in human breast cancer is associated with undifferentiated histology and poor prognosis. Here, we identify the mammary cell growth factor prolactin as a potent suppressor of BCL6 protein expression in human breast cancer through a mechanism that requires Stat5a, but not prolactin-activated Stat5b, MEK-ERK, or PI3K-AKT pathways. Prolactin rapidly suppressed BCL6 mRNA in T47D, MCF7, ZR75.1, and SKBr3 breast cancer cell lines, followed by prolonged reduction of BCL6 protein levels within 3 hours. Prolactin suppression of BCL6 was enhanced by overexpression of Stat5a but not Stat5b, was mimicked by constitutively active Stat5a, but did not require the transactivation domain of Stat5a. Stat5 chromatin immunoprecipitation demonstrated physical interaction with a BCL6 gene regulatory region, and BCL6 transcript repression required histone deacetylase activity based on sensitivity to trichostatin A. Functionally, BCL6 overexpression disrupted prolactin induction of Stat5 reporter genes. Prolactin suppression of BCL6 was extended to xenotransplant tumors in nude mice in vivo and to freshly isolated human breast cancer explants ex vivo. Quantitative immunohistochemistry revealed elevated BCL6 in high-grade and metastatic breast cancer compared with ductal carcinoma in situ and nonmalignant breast, and cellular BCL6 protein levels correlated negatively with nuclear Stat5a (r = −0.52; P < 0.001) but not with Stat5b. Loss of prolactin-Stat5a signaling and concomitant upregulation of BCL6 may represent a regulatory switch facilitating undifferentiated histology and poor prognosis of breast cancer. Cancer Res; 70(4); 1711–21
Data from Phosphatase PRL-3 Is a Direct Regulatory Target of TGFβ in Colon Cancer Metastasis
Authors: Jiang Yanjun , Liu Xiao-Qiong , Rajput Ashwani , Geng Liying , Ongchin Melanie , Zeng Qi , Taylor Gregory S. , Wang Jing
Published: March 2024

Abstract
Metastasis causes most deaths from cancer yet mechanistic understanding and therapeutic options remain limited. Overexpression of the phosphatase PRL-3 (phosphatase of regenerating liver) is associated with metastasis of colon cancer. Here, we show that PRL-3 is a direct target of signaling by TGFβ, which is broadly implicated in progression and metastasis. We found that suppression of PRL-3 expression by TGFβ was mediated by Smad-dependent inhibition of PRL-3 transcription at the level of promoter activity. PRL-3 activation stimulated PI3K/AKT signaling that caused resistance to stress-induced apoptosis. PRL-3 overexpression promoted metastatic colonization in an orthotopic mouse model of colon cancer, whereas PRL-3 knockdown reduced metastatic potential. Altered metastatic phenotypes were not derivative of primary tumor development or local invasion but could be attributed to PRL-3–mediated cell survival. Our findings suggest that inhibiting PRL-3 expression might be an important mechanism through which TGFβ suppresses metastasis in colon cancer. In addition, our findings suggest that loss of TGFβ signaling, which occurs commonly during colon cancer progression, is sufficient to activate a PRL-3–mediated cell survival pathway that can selectively promote metastasis. Therefore, a major implication of our findings is that PRL-3 antagonists may offer significant value for antimetastatic therapy in patients with colon cancer. Cancer Res; 71(1); 234–44. ©2010 AACR.
Data from Kallikrein-Related Peptidase 7 Promotes Multicellular Aggregation via the α5β1 Integrin Pathway and Paclitaxel Chemoresistance in Serous Epithelial Ovarian Carcinoma
Authors: Dong Ying , Tan Olivia L. , Loessner Daniela , Stephens Carson , Walpole Carina , Boyle Glen M. , Parsons Peter G. , Clements Judith A.
Published: March 2024

Abstract
Kallikrein-related peptidase 7 (KLK7) is upregulated in epithelial ovarian carcinoma (EOC) with high levels correlated with poor prognosis. However, the mechanisms underlying this relationship and the role of KLK7 in EOC progression are unknown. We report that two different KLK7 transcripts, KLK7-253 and KLK7-181, are simultaneously expressed in high-grade serous EOC. Multicellular aggregates (MCA), which promote cell survival and chemoresistance, were observed in SKOV-3 cells stably overexpressing KLK7-253 in particular. Importantly, these MCAs invade into a monolayer of mesothelial cells and form cancer cell foci. Blocking MCA using antibodies against KLK7 and α5β1 and β1 integrins confirmed the involvement of KLK7 and integrin-regulated cell adhesion. Increased levels of α5/β1 integrins and enhanced attachment to fibronectin and vitronectin, which was blocked with an anti–β1 integrin antibody, were also observed. Finally, Western blot and immunohistochemistry showed higher KLK7 and α5/β1 integrin levels in serous EOC cells from ascites and tumor samples from chemotherapy nonresponders with short postsurvival times. Additionally, both KLK7-253 and KLK7-181 clones were more resistant to paclitaxel treatment in vitro. These findings suggest a mechanism for the association of high KLK7 levels with chemoresistance and poor prognosis for serous EOC patients by promotion of peritoneal dissemination and reinvasion via increased MCA and α5β1 integrin–dependent cell adhesion. Cancer Res; 70(7); 2624–33
Data from Cancer Stem Cells in Squamous Cell Carcinoma Switch between Two Distinct Phenotypes That Are Preferentially Migratory or Proliferative
Authors: Biddle Adrian , Liang Xiao , Gammon Luke , Fazil Bilal , Harper Lisa J. , Emich Helena , Costea Daniela Elena , Mackenzie Ian C.
Published: March 2024

Abstract
Epithelial-to-mesenchymal transition (EMT) is an important driver of tumor invasion and metastasis, which causes many cancer deaths. Cancer stem cells (CSC) that maintain and initiate tumors have also been implicated in invasion and metastasis, but whether EMT is an important contributor to CSC function is unclear. In this study, we investigated whether a population of CSCs that have undergone EMT (EMT CSCs) exists in squamous cell carcinoma (SCC). We also determined whether a separate population of CSCs that retain epithelial characteristics (non-EMT CSCs) is also present. Our studies revealed that self-renewing CSCs in SCC include two biologically-distinct phenotypes. One phenotype, termed CD44highESAhigh, was proliferative and retained epithelial characteristics (non-EMT CSCs), whereas the other phenotype, termed CD44highESAlow, was migratory and had mesenchymal traits characteristic of EMT CSCs. We found that non-EMT and EMT CSCs could switch their epithelial or mesenchymal traits to reconstitute the cellular heterogeneity which was characteristic of CSCs. However, the ability of EMT CSCs to switch to non-EMT character was restricted to cells that were also ALDH1+, implying that only ALDH1+ EMT cells had the ability to seed a new epithelial tumor. Taken together, our findings highlight the identification of two distinct CSC phenotypes and suggest a need to define therapeutic targets that can eradicate both of these variants to achieve effective SCC treatment. Cancer Res; 71(15); 5317–26. ©2011 AACR.
Data from Prognostic Value and Function of KLF4 in Prostate Cancer: RNAa and Vector-Mediated Overexpression Identify KLF4 as an Inhibitor of Tumor Cell Growth and Migration
Authors: Wang Ji , Place Robert F. , Huang Vera , Wang Xiaoling , Noonan Emily J. , Magyar Clara E. , Huang Jiaoti , Li Long-Cheng
Published: March 2024

Abstract
KLF4/GLKF4 is a transcription factor that can have divergent functions in different malignancies. The role of KLF4 in prostate cancer etiology remains unclear. We have recently reported that small double-stranded RNA can induce gene expression by targeting promoter sequence in a phenomenon referred to as RNA activation (RNAa). In this study, we examine KLF4 levels in prostate cancer tissue and utilize RNAa as a tool for gene overexpression to investigate its function. Expression analysis indicated that KLF4 is significantly downregulated in prostate cancer cell lines compared with nontumorigenic prostate cells. Meta-analysis of existing cDNA microarray data also revealed that KLF4 is frequently depleted in prostate cancer tissue with more pronounced reduction in metastases. In support, tissue microarray analysis of tumors and patient-matched controls indicated downregulation of KLF4 in metastatic tumor samples. Logistic regression analysis found that tumors with a KLF4 staining score less than 5 had a 15-fold higher risk for developing metastatic prostate cancer (P = 0.001; 95% confidence interval, 3.0–79.0). In vitro analysis indicated that RNAa-mediated overexpression of KLF4 inhibited prostate cancer cell proliferation and survival and altered the expression of several downstream cell-cycle–related genes. Ectopic expression of KLF4 via viral transduction recapitulated the RNAa results, validating its inhibitory effects on cancer growth. Reactivation of KLF4 also suppressed migration and invasion of prostate cancer cells. These results suggest that KLF4 functions as an inhibitor of tumor cell growth and migration in prostate cancer and decreased expression has prognostic value for predicting prostate cancer metastasis. Cancer Res; 70(24); 10182–91. ©2010 AACR.
Data from The Role of Calcium in the Activation of Estrogen Receptor-Alpha
Authors: Divekar Shailaja D. , Storchan Geoffrey B. , Sperle Katherine , Veselik David J. , Johnson Earl , Dakshanamurthy Sivanesan , Lajiminmuhip Yuse N. , Nakles Rebecca E. , Huang Li , Martin Mary Beth
Published: March 2024

Abstract
Environmental estrogen mimics, including metalloestrogens that can activate estrogen receptor-alpha (ERα), may contribute to breast cancer risk. However, the underlying mechanisms through which these molecular mimics activate the ERα are generally poorly understood. With concern to this important question, we investigated whether intracellular calcium may mediate the cross-talk between signaling pathways that activate ERα and the ligand-binding domain of ERα. MCF-7 cells treated with EGF, ATP, extracellular calcium, or caffeine to increase intracellular calcium triggered a rapid recruitment of ERα to estrogen-responsive promoters and stimulated expression of estrogen-responsive genes including pS2, complement C3, and progesterone receptor. Induction was blocked by an antiestrogen but also by the chelation of intracellular calcium. Treatment with extracellular calcium also increased the growth of MCF-7 cells through an ER-dependent mechanism. We found that EGF and extracellular calcium activated the C-terminus of ERα and the activation was blocked by the antiestrogen. Mechanistic investigations identified four potential sites on the solvent-accessible surface of the ERα ligand-binding domain as important for calcium activation of the receptor. Taken together, our results suggest that calcium mediates the cross-talk between ERα-activating signaling pathways and the ligand-binding domain of ERα providing a potential explanation for the ability of certain environmental metalloestrogens to activate the receptor. Cancer Res; 71(5); 1658–68. ©2011 AACR.
Data from ABCB5 Identifies a Therapy-Refractory Tumor Cell Population in Colorectal Cancer Patients
Authors: Wilson Brian J. , Schatton Tobias , Zhan Qian , Gasser Martin , Ma Jie , Saab Karim R. , Schanche Robin , Waaga-Gasser Ana-Maria , Gold Jason S. , Huang Qin , Murphy George F. , Frank Markus H. , Frank Natasha Y.
Published: March 2024

Abstract
Identification and reversal of treatment resistance mechanisms of clinically refractory tumor cells is critical for successful cancer therapy. Here we show that ATP-binding cassette member B5 (ABCB5) identifies therapy-refractory tumor cells in colorectal cancer patients following fluorouracil (5-FU)–based chemoradiation therapy and provide evidence for a functional role of ABCB5 in colorectal cancer 5-FU resistance. Examination of human colon and colorectal cancer specimens revealed ABCB5 to be expressed only on rare cells within healthy intestinal tissue, whereas clinical colorectal cancers exhibited substantially increased levels of ABCB5 expression. Analysis of successive, patient-matched biopsy specimens obtained prior to and following neoadjuvant 5-FU–based chemoradiation therapy in a series of colorectal cancer patients revealed markedly enhanced abundance of ABCB5-positive tumor cells when residual disease was detected. Consistent with this finding, the ABCB5-expressing tumor cell population was also treatment refractory and exhibited resistance to 5-FU–induced apoptosis in a colorectal cancer xenograft model of 5-FU monotherapy. Mechanistically, short hairpin RNA–mediated ABCB5 knockdown significantly inhibited tumorigenic xenograft growth and sensitized colorectal cancer cells to 5-FU–induced cell killing. Our results identify ABCB5 as a novel molecular marker of therapy-refractory tumor cells in colorectal cancer patients and point to a need for consistent eradication of ABCB5-positive resistant tumor cell populations for more effective colorectal cancer therapy. Cancer Res; 71(15); 5307–16. ©2011 AACR.
Data from Inhibition of Histone Lysine Methylation Enhances Cancer–Testis Antigen Expression in Lung Cancer Cells: Implications for Adoptive Immunotherapy of Cancer
Authors: Rao Mahadev , Chinnasamy Nachimuthu , Hong Julie A. , Zhang Yuwei , Zhang Mary , Xi Sichuan , Liu Fang , Marquez Victor E. , Morgan Richard A. , Schrump David S.
Published: March 2024

Abstract
Cancer–testis antigens (CTA), such as NY-ESO-1, MAGE-A1, and MAGE-A3, are immunogenic proteins encoded by genes, which are normally expressed only in male germ cells but are activated by ill-defined epigenetic mechanisms in human tumors, including lung cancers. Previously, we reported induction of these CTAs in cancer cells, but not normal cells, by DNA-demethylating agents and histone deacetylase inhibitors using clinically achievable exposure conditions. In the present study, we evaluated chromatin alterations associated with repression/activation of cancer–testis genes in lung cancer cells to further develop gene-induction regimens for cancer immunotherapy. Repression of NY-ESO-1, MAGE-A1, and MAGE-A3 coincided with DNA hypermethylation, recruitment, and binding of polycomb-group proteins, and histone heterochromatin modifications within the promoters of these genes. Derepression coincided with DNA demethylation, dissociation of polycomb proteins, and presence of euchromatin marks within the respective promoters. Short hairpin RNAs were used to inhibit several histone methyltransferases (KMT) and histone demethylases (KDM) that mediate histone methylation and repress gene expression. Knockdown of KMT6, KDM1, or KDM5B markedly enhanced deoxyazacytidine (DAC)-mediated activation of these cancer–testis genes in lung cancer cells. DZNep, a pharmacologic inhibitor of KMT6 expression, recapitulated the effects of KMT6 knockdown. Following DAC–DZNep exposure, lung cancer cells were specifically recognized and lysed by allogeneic lymphocytes expressing recombinant T-cell receptors recognizing NY-ESO-1 and MAGE-A3. Combining DNA-demethylating agents with compounds, such as DZNep, that modulate histone lysine methylation may provide a novel epigenetic strategy to augment cancer–testis gene expression as an adjunct to adoptive cancer immunotherapy. Cancer Res; 71(12); 4192–204. ©2011 AACR.
Data from Cytokine Receptor CXCR4 Mediates Estrogen-Independent Tumorigenesis, Metastasis, and Resistance to Endocrine Therapy in Human Breast Cancer
Authors: Rhodes Lyndsay V. , Short Sarah P. , Neel Nicole F. , Salvo Virgilio A. , Zhu Yun , Elliott Steven , Wei Yongkun , Yu Dihua , Sun Menghong , Muir Shannon E. , Fonseca Juan P. , Bratton Melyssa R. , Segar Chris , Tilghman Syreeta L. , Sobolik-Delmaire Tammy , Horton Linda W. , Zaja-Milatovic Snjezana , Collins-Burow Bridgette M. , Wadsworth Scott , Beckman Barbara S. , Wood Charles E. , Fuqua Suzanne A. , Nephew Kenneth P. , Dent Paul , Worthylake Rebecca A. , Curiel Tyler J. , Hung Mien-Chie , Richmond Ann , Burow Matthew E.
Published: March 2024

Abstract
Estrogen independence and progression to a metastatic phenotype are hallmarks of therapeutic resistance and mortality in breast cancer patients. Metastasis has been associated with chemokine signaling through the SDF-1–CXCR4 axis. Thus, the development of estrogen independence and endocrine therapy resistance in breast cancer patients may be driven by SDF-1–CXCR4 signaling. Here we report that CXCR4 overexpression is indeed correlated with worse prognosis and decreased patient survival irrespective of the status of the estrogen receptor (ER). Constitutive activation of CXCR4 in poorly metastatic MCF-7 cells led to enhanced tumor growth and metastases that could be reversed by CXCR4 inhibition. CXCR4 overexpression in MCF-7 cells promoted estrogen independence in vivo, whereas exogenous SDF-1 treatment negated the inhibitory effects of treatment with the anti-estrogen ICI 182,780 on CXCR4-mediated tumor growth. The effects of CXCR4 overexpression were correlated with SDF-1–mediated activation of downstream signaling via ERK1/2 and p38 MAPK (mitogen activated protein kinase) and with an enhancement of ER-mediated gene expression. Together, these results show that enhanced CXCR4 signaling is sufficient to drive ER-positive breast cancers to a metastatic and endocrine therapy-resistant phenotype via increased MAPK signaling. Our findings highlight CXCR4 signaling as a rational therapeutic target for the treatment of ER-positive, estrogen-independent breast carcinomas needing improved clinical management. Cancer Res; 71(2); 603–13. ©2010 AACR.
Data from p53-Dependent BRCA1 Nuclear Export Controls Cellular Susceptibility to DNA Damage
Authors: Jiang Juhong , Yang Eddy S. , Jiang Guochun , Nowsheen Somaira , Wang Hong , Wang Tong , Wang Yihan , Billheimer Dean , Chakravarthy A. Bapsi , Brown Melissa , Haffty Bruce , Xia Fen
Published: March 2024

Abstract
Subcellular localization regulates BRCA1 function, and BRCA1 is exported to the cytoplasm following DNA damage in a p53-dependent manner. Because more than 50% of solid tumors harbor p53 mutations, it is possible that genetically wild-type (wt) BRCA1 is functionally abnormal through compromised nuclear-cytoplasmic shuttling in sporadic breast cancer patients with dysfunctional p53. In this study, we have investigated the mechanisms of p53-dependent BRCA1 subcellular distribution and DNA damage-induced nuclear export, as well as the impact on the resulting cytotoxic response to therapy in human breast cancer. We first show that p53 mediates BRCA1 nuclear export via protein–protein binding, rather than by modulation of its transcription. Furthermore, it is the C-terminal (BRCT) region of BRCA1 that is critical for its interaction with p53, and p53 may promote BRCA1 nuclear export by interrupting the association of BRCA1 with BARD1. In sporadic breast cancer specimens, dysfunctional p53 strongly correlates with nuclear retention of sequence-verified wt BRCA1. This p53-dependent BRCA1 shuttling determines cellular susceptibility to DNA damage as augmentation of cytosolic BRCA1 significantly enhances cancer cell susceptibility to ionizing radiation. Taken together, our data suggest that p53 dysfunction compromises nuclear export of wt BRCA1 as a mechanism to increase cellular resistance to DNA damage in sporadic breast cancer. We propose that targeting nuclear BRCA1 to the cytoplasm may offer a unique strategy to sensitize p53-deficient sporadic breast cancers to DNA damage–based therapy. Cancer Res; 71(16); 5546–57. ©2011 AACR.
Data from Celecoxib Promotes c-FLIP Degradation through Akt-Independent Inhibition of GSK3
Authors: Chen Shuzhen , Cao Wei , Yue Ping , Hao Chunhai , Khuri Fadlo R. , Sun Shi-Yong
Published: March 2024

Abstract
Celecoxib is a COX-2 inhibitor that reduces the risk of colon cancer. However, the basis for its cancer chemopreventive activity is not fully understood. In this study, we defined a mechanism of celecoxib action based on degradation of cellular FLICE-inhibitory protein (c-FLIP), a major regulator of the death receptor pathway of apoptosis. c-FLIP protein levels are regulated by ubiquitination and proteasome-mediated degradation. We found that celecoxib controlled c-FLIP ubiquitination through Akt-independent inhibition of glycogen synthase kinase-3 (GSK3), itself a candidate therapeutic target of interest in colon cancer. Celecoxib increased the levels of phosphorylated GSK3, including the α and β forms, even in cell lines, where phosphorylated Akt levels were not increased. Phosphoinositide 3-kinase inhibitors abrogated Akt phosphorylation as expected but had no effect on celecoxib-induced GSK3 phosphorylation. In contrast, protein kinase C (PKC) inhibitors abolished celecoxib-induced GSK3 phosphorylation, implying that celecoxib influenced GSK3 phosphorylation through a mechanism that relied upon PKC and not Akt. GSK3 blockade either by siRNA or kinase inhibitors was sufficient to attenuate c-FLIP levels. Combining celecoxib with GSK3 inhibition enhanced attenuation of c-FLIP and increased apoptosis. Proteasome inhibitor MG132 reversed the effects of GSK3 inhibition and increased c-FLIP ubiquitination, confirming that c-FLIP attenuation was mediated by proteasomal turnover as expected. Our findings reveal a novel mechanism through which the regulatory effects of c-FLIP on death receptor signaling are controlled by GSK3, which celecoxib acts at an upstream level to control independently of Akt. Cancer Res; 71(19); 6270–81. ©2011 AACR.
Data from The Mef/Elf4 Transcription Factor Fine Tunes the DNA Damage Response
Authors: Sashida Goro , Bae Narae , Di Giandomenico Silvana , Asai Takashi , Gurvich Nadia , Bazzoli Elena , Liu Yan , Huang Gang , Zhao Xinyang , Menendez Silvia , Nimer Stephen D.
Published: March 2024

Abstract
The ATM kinase plays a critical role in initiating the DNA damage response that is triggered by genotoxic stresses capable of inducing DNA double-strand breaks. Here, we show that ELF4/MEF, a member of the ETS family of transcription factors, contributes to the persistence of γH2AX DNA damage foci and promotes the DNA damage response leading to the induction of apoptosis. Conversely, the absence of ELF4 promotes the faster repair of damaged DNA and more rapid disappearance of γH2AX foci in response to γ-irradiation, leading to a radio-resistant phenotype despite normal ATM phosphorylation. Following γ-irradiation, ATM phosphorylates ELF4, leading to its degradation; a mutant form of ELF4 that cannot be phosphorylated by ATM persists following γ-irradiation, delaying the resolution of γH2AX foci and triggering an excessive DNA damage response. Thus, although ELF4 promotes the phosphorylation of H2AX by ATM, its activity must be dampened by ATM-dependent phosphorylation and degradation to avoid an excessive DNA damage response. Cancer Res; 71(14); 4857–65. ©2011 AACR.
Data from Navitoclax (ABT-263) Accelerates Apoptosis during Drug-Induced Mitotic Arrest by Antagonizing Bcl-xL
Authors: Shi Jue , Zhou Yuan , Huang Hsiao-Chun , Mitchison Timothy J.
Published: March 2024

Abstract
Combining microtubule-targeting antimitotic drugs with targeted apoptosis potentiators is a promising new chemotherapeutic strategy to treat cancer. In this study, we investigate the cellular mechanism by which navitoclax (previously called ABT-263), a Bcl-2 family inhibitor, potentiates apoptosis triggered by paclitaxel and an inhibitor of kinesin-5 (K5I, also called a KSP inhibitor), across a panel of epithelial cancer lines. By using time-lapse microscopy, we showed that navitoclax has little effect on cell death during interphase, but strongly accelerates apoptosis during mitotic arrest, and greatly increases the fraction of apoptosis-resistant cells that die. By systematically knocking down individual Bcl-2 proteins, we determined that Mcl-1 and Bcl-xL are the primary negative regulators of apoptosis during prolonged mitotic arrest. Mcl-1 levels decrease during mitotic arrest because of an imbalance between synthesis and turnover, and turnover depends in part on the MULE/HUWE1 E3 ligase. The combination of Mcl-1 loss with inhibition of Bcl-xL by navitoclax causes rapid apoptosis in all lines tested. Variation in expression levels of Mcl-1 and Bcl-xL largely determines variation in response to antimitotics alone, and antimitotics combined with navitoclax, across our panel. We concluded that Bcl-xL is a critical target of Bcl-2 family inhibitors for enhancing the lethality of antimitotic drugs in epithelial cancers, and combination treatment with navitoclax and a spindle specific antimitotic, such as a K5I, might be more effective than paclitaxel alone. Cancer Res; 71(13); 4518–26. ©2011 AACR.
Data from Protein Kinase D3 Sensitizes RAF Inhibitor RAF265 in Melanoma Cells by Preventing Reactivation of MAPK Signaling
Authors: Chen Jian , Shen Qiong , Labow Mark , Gaither L. Alex
Published: March 2024

Abstract
RAS mutations occur in more than 30% of all human cancers but efforts to directly target mutant RAS signaling as a cancer therapy have yet to succeed. As alternative strategies, RAF and MEK inhibitors have been developed to block oncogenic signaling downstream of RAS. As might be expected, studies of these inhibitors have indicated that tumors with RAS or BRAF mutations display resistance RAF or MEK inhibitors. In order to better understand the mechanistic basis for this resistance, we conducted a RNAi-based screen to identify genes that mediated chemoresistance to the RAF kinase inhibitor RAF265 in a BRAF (V600E) mutant melanoma cell line that is resistant to this drug. In this way, we found that knockdown of protein kinase D3 (PRKD3) could enhance cell killing of RAF and MEK inhibitors across multiple melanoma cell lines of various genotypes and sensitivities to RAF265. PRKD3 blockade cooperated with RAF265 to prevent reactivation of the MAPK signaling pathway, interrupt cell cycle progression, trigger apoptosis, and inhibit colony formation growth. Our findings offer initial proof-of-concept that PRKD3 is a valid target to overcome drug resistance being encountered widely in the clinic with RAF or MEK inhibitors. Cancer Res; 71(12); 4280–91. ©2011 AACR.
Data from Caveolin-1 Upregulation Mediates Suppression of Primary Breast Tumor Growth and Brain Metastases by Stat3 Inhibition
Authors: Chiu Wen-Tai , Lee Hsueh-Te , Huang Feng-Ju , Aldape Kenneth D. , Yao Jun , Steeg Patricia S. , Chou Cheng-Yang , Lu Zhimin , Xie Keping , Huang Suyun
Published: March 2024

Abstract
Stat3 activation has been implicated as an important driver of brain metastasis in breast cancer, but the critical targets of Stat3 in this process are yet to be fully defined. In this study, we identified the lipid raft organizing protein Caveolin-1 (Cav-1) as a critical genetic target of Stat3 in this process. In human breast cancers, we found that activated Stat3 correlated with attenuation of Cav-1 in brain metastases relative to primary tumors. Cav-1 promoter activity and gene expression were increased by overexpressing an activated form of Stat3 but decreased by attenuation of Stat3 activity or expression. We identified putative Stat3-binding elements in the Cav-1 promoter and showed a direct repression of Cav-1 transcription by Stat3. Reciprocally, we showed that strategies to increase or decrease Cav-1 expression were sufficient to attenuate or promote breast cancer cell invasion. Furthermore, increased expression of Cav-1 phenocopied the effects of Stat3 activation in blocking primary tumor growth and abrogating formation of brain metastases. Collectively, our findings provide clinical and mechanistic evidence that Cav-1 is a critical target for suppression by Stat3 in driving invasion and metastasis of breast cancer cells. Cancer Res; 71(14); 4932–43. ©2011 AACR.
Data from 18F-Fluorodeoxy-glucose Positron Emission Tomography Marks MYC-Overexpressing Human Basal-Like Breast Cancers
Authors: Palaskas Nicolaos , Larson Steven M. , Schultz Nikolaus , Komisopoulou Evangelia , Wong Justin , Rohle Dan , Campos Carl , Yannuzzi Nicolas , Osborne Joseph R. , Linkov Irina , Kastenhuber Edward R. , Taschereau Richard , Plaisier Seema B. , Tran Chris , Heguy Adriana , Wu Hong , Sander Chris , Phelps Michael E. , Brennan Cameron , Port Elisa , Huse Jason T. , Graeber Thomas G. , Mellinghoff Ingo K.
Published: March 2024

Abstract
In contrast to normal cells, cancer cells avidly take up glucose and metabolize it to lactate even when oxygen is abundant, a phenomenon referred to as the Warburg effect. This fundamental alteration in glucose metabolism in cancer cells enables their specific detection by positron emission tomography (PET) following i.v. injection of the glucose analogue 18F-fluorodeoxy-glucose (18FDG). However, this useful imaging technique is limited by the fact that not all cancers avidly take up FDG. To identify molecular determinants of 18FDG retention, we interrogated the transcriptomes of human-cancer cell lines and primary tumors for metabolic pathways associated with 18FDG radiotracer uptake. From ninety-five metabolic pathways that were interrogated, the glycolysis, and several glycolysis-related pathways (pentose phosphate, carbon fixation, aminoacyl-tRNA biosynthesis, one-carbon-pool by folate) showed the greatest transcriptional enrichment. This “FDG signature” predicted FDG uptake in breast cancer cell lines and overlapped with established gene expression signatures for the “basal-like” breast cancer subtype and MYC-induced tumorigenesis in mice. Human breast cancers with nuclear MYC staining and high RNA expression of MYC target genes showed high 18FDG-PET uptake (P < 0.005). Presence of the FDG signature was similarly associated with MYC gene copy gain, increased MYC transcript levels, and elevated expression of metabolic MYC target genes in a human breast cancer genomic dataset. Together, our findings link clinical observations of glucose uptake with a pathologic and molecular subtype of human breast cancer. Furthermore, they suggest related approaches to derive molecular determinants of radiotracer retention for other PET-imaging probes. Cancer Res; 71(15); 5164–74. ©2011 AACR.
Data from TMPRSS2–ERG-Mediated Feed-Forward Regulation of Wild-Type ERG in Human Prostate Cancers
Authors: Mani Ram-Shankar , Iyer Matthew K. , Cao Qi , Brenner J. Chad , Wang Lei , Ghosh Aparna , Cao Xuhong , Lonigro Robert J. , Tomlins Scott A. , Varambally Sooryanarayana , Chinnaiyan Arul M.
Published: March 2024

Abstract
Recurrent gene fusions involving ETS family genes are a distinguishing feature of human prostate cancers, with TMPRSS2–ERG fusions representing the most common subtype. The TMPRSS2–ERG fusion transcript and its splice variants are well characterized in prostate cancers; however, not much is known about the levels and regulation of wild-type ERG. By employing an integrative approach, we show that the TMPRSS2–ERG gene fusion product binds to the ERG locus and drives the overexpression of wild-type ERG in prostate cancers. Knockdown of TMPRSS2–ERG in VCaP cells resulted in the downregulation of wild-type ERG transcription, whereas stable overexpression of TMPRSS2–ERG in the gene fusion-negative PC3 cells was associated with the upregulation of wild-type ERG transcript. Further, androgen signaling-mediated upregulation of TMPRSS2–ERG resulted in the concomitant upregulation of wild-type ERG transcription in VCaP cells. The loss of wild-type ERG expression was associated with a decrease in the invasive potential of VCaP cells. Importantly, 38% of clinically localized prostate cancers and 27% of metastatic prostate cancers harboring the TMPRSS2–ERG gene fusions exhibited overexpression of wild-type ERG. Taken together, these results provide novel insights into the regulation of ERG in human prostate cancers. Cancer Res; 71(16); 5387–92. ©2011 AACR.
Data from Verticillin A Overcomes Apoptosis Resistance in Human Colon Carcinoma through DNA Methylation-Dependent Upregulation of BNIP3
Authors: Liu Feiyan , Liu Qianqian , Yang Dafeng , Bollag Wendy B. , Robertson Keith , Wu Ping , Liu Kebin
Published: March 2024

Abstract
Drug resistance is a major cause of failure in cancer chemotherapy. Therefore, identification and combined use of adjuvant compounds that can overcome drug resistance may improve the efficacy of cancer therapy. We screened extracts of Verticillium species-infected mushrooms for antitumor compounds and identified the compound Verticillin A as an inducer of hepatoma cell apoptosis in vitro and an inhibitor of tumor xenograft growth in vivo. Verticillin A exhibited a potent apoptosis-sensitizing activity in human colon carcinoma cells exposed to TRAIL or Fas in vitro. Furthermore, Verticillin A effectively sensitized metastatic human colon carcinoma xenograft to TRAIL-mediated growth inhibition in vivo. At the molecular level, we observed that Verticillin A induces cell-cycle arrest in the G2 phase of the cell cycle in human colon carcinoma cells, markedly upregulating BNIP3 in both hepatoma and colon carcinoma cells. Notably, silencing BNIP3 decreased the sensitivity of tumor cells to Verticillin A–induced apoptosis in the absence or presence of TRAIL. We found that the BNIP3 promoter is methylated in both human hepatoma and colon carcinoma cells and tumor specimens. Verticillin A upregulated the expression of a panel of genes known to be regulated at the level of DNA methylation, in support of the concept that Verticillin A may act by demethylating the BNIP3 promoter to upregulate BNIP3 expression. Taken together, our findings identify Verticillin A as a potent apoptosis sensitizer with great promise for further development as an adjuvant agent to overcome drug resistance in human cancer therapy. Cancer Res; 71(21); 6807–16. ©2011 AACR.
Data from Quantitative, Noninvasive Imaging of Radiation-Induced DNA Double-Strand Breaks In Vivo
Authors: Li Wenrong , Li Fang , Huang Qian , Shen Jingping , Wolf Frank , He Yujun , Liu Xinjian , Hu Y. Angela , Bedford Joel S. , Li Chuan-Yuan
Published: March 2024

Abstract
DNA double-strand breaks (DSB) are a major form of DNA damage and a key mechanism through which radiotherapy and some chemotherapeutic agents kill cancer cells. Despite its importance, measuring DNA DSBs is still a tedious task that is normally carried out by gel electrophoresis or immunofluorescence staining. Here, we report a novel approach to image and quantify DSBs in live mammalian cells through bifragment luciferase reconstitution. N- and C-terminal fragments of firefly luciferase genes were fused with H2AX and MDC1 genes, respectively. Our strategy was based on the established fact that at the sites of DSBs, H2AX protein is phosphoryated and physically associates with the MDC1 protein, thus bringing together N- and C-luciferase fragments and reconstituting luciferase activity. Our strategy allowed serial, noninvasive quantification of DSBs in cells irradiated with X-rays and 56Fe ions. Furthermore, it allowed for the evaluation of DSBs noninvasively in vivo in irradiated tumors over 2 weeks. Surprisingly, we detected a second wave of DSB induction in irradiated tumor cells days after radiation exposure in addition to the initial rapid induction of DSBs. We conclude that our new split-luciferase–based method for imaging γ-H2AX–MDC1 interaction is a powerful new tool to study DSB repair kinetics in vivo with considerable advantage for experiments requiring observations over an extended period of time. Cancer Res; 71(12); 4130–7. ©2011 AACR.
Data from GPR56 Regulates VEGF Production and Angiogenesis during Melanoma Progression
Authors: Yang Liquan , Chen Guangchun , Mohanty Sonali , Scott Glynis , Fazal Fabeha , Rahman Arshad , Begum Shahinoor , Hynes Richard O. , Xu Lei
Published: March 2024

Abstract
Angiogenesis is a critical step during cancer progression. The VEGF is a major stimulator for angiogenesis and is predominantly contributed by cancer cells in tumors. Inhibition of the VEGF signaling pathway has shown promising therapeutic benefits for cancer patients, but adaptive tumor responses are often observed, indicating the need for further understanding of VEGF regulation. We report that a novel G protein–coupled receptor, GPR56, inhibits VEGF production from the melanoma cell lines and impedes melanoma angiogenesis and growth, through the serine threonine proline-rich segment in its N-terminus and a signaling pathway involving protein kinase Cα. We also present evidence that the two fragments of GPR56, which are generated by autocatalyzed cleavage, played distinct roles in regulating VEGF production and melanoma progression. Finally, consistent with its suppressive roles in melanoma progression, the expression levels of GPR56 are inversely correlated with the malignancy of melanomas in human subjects. We propose that components of the GPR56-mediated signaling pathway may serve as new targets for antiangiogenic treatment of melanoma. Cancer Res; 71(16); 5558–68. ©2011 AACR.
Data from Following Cytochrome c Release, Autophagy Is Inhibited during Chemotherapy-Induced Apoptosis by Caspase 8–Mediated Cleavage of Beclin 1
Authors: Li Hua , Wang Peng , Sun Quanhong , Ding Wen-Xing , Yin Xiao-Ming , Sobol Robert W. , Stolz Donna B. , Yu Jian , Zhang Lin
Published: March 2024

Abstract
Autophagy is an evolutionarily conserved stress response mechanism that often occurs in apoptosis-defective cancer cells and can protect against cell death. In this study, we investigated how apoptosis and autophagy affect each other in cancer cells in response to chemotherapeutic treatment. We found that specific ablation of the proapoptotic function of cytochrome c, a key regulator of mitochondria-mediated apoptosis, enhanced autophagy following chemotherapeutic treatment. Induction of autophagy required Beclin 1 and was associated with blockage of Beclin 1 cleavage by caspase 8 at two sites. To investigate the role of Beclin 1 cleavage in the suppression of autophagy and cell survival, a caspase-resistant mutant of Beclin 1 was knocked into HCT116 colon cancer cells. Beclin 1 mutant knockin resulted in markedly increased autophagy and improved long-term cell survival after chemotherapeutic treatment but without affecting apoptosis and caspase activation. Furthermore, Beclin 1 mutant tumors were significantly less responsive to chemotherapeutic treatment than were wild-type tumors. These results show that chemotherapy-induced apoptosis inhibits autophagy at the execution stage subsequent to cytochrome c release through caspase 8–mediated cleavage of Beclin 1. If apoptosis fails to execute, autophagy is unleashed due to lack of Beclin 1 cleavage by caspases and can contribute to cancer cell survival and therapeutic resistance. Therefore, Beclin 1 may be a useful target for inhibiting autophagy to sensitize chemotherapy. Cancer Res; 71(10); 3625–34. ©2011 AACR.