bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Flavokawain A, a Novel Chalcone from Kava Extract, Induces Apoptosis in Bladder Cancer Cells by Involvement of Bax Protein-Dependent and Mitochondria-Dependent Apoptotic Pathway and Suppresses Tumor Growth in Mice
Authors: Zi Xiaolin , Simoneau Anne R.
Published: March 2023

Abstract
Consumption of the traditional kava preparation was reported to correlate with low and uncustomary gender ratios (more cancer in women than men) of cancer incidences in three kava-drinking countries: Fiji, Vanuatu, and Western Samoa. We have identified flavokawain A, B, and C but not the major kavalactone, kawain, in kava extracts as causing strong antiproliferative and apoptotic effect in human bladder cancer cells. Flavokawain A results in a significant loss of mitochondrial membrane potential and release of cytochrome c into the cytosol in an invasive bladder cancer cell line T24. These effects of flavokawain A are accompanied by a time-dependent decrease in Bcl-xL, a decrease in the association of Bcl-xL to Bax, and an increase in the active form of Bax protein. Using the primary mouse embryo fibroblasts Bax knockout and wild-type cells as well as a Bax inhibitor peptide derived from the Bax-binding domain of Ku70, we showed that Bax protein was, at least in part, required for the apoptotic effect of flavokawain A. In addition, flavokawain A down-regulates the expression of X-linked inhibitor of apoptosis and survivin. Because both X-linked inhibitor of apoptosis and survivin are main factors for apoptosis resistance and are overexpressed in bladder tumors, our data suggest that flavokawain A may have a dual efficacy in induction of apoptosis preferentially in bladder tumors. Finally, the anticarcinogenic effect of flavokawain A was evident in its inhibitory growth of bladder tumor cells in a nude mice model (57% of inhibition) and in soft agar.
Data from NUP98-HOXA9 Induces Long-term Proliferation and Blocks Differentiation of Primary Human CD34+ Hematopoietic Cells
Authors: Takeda Akiko , Goolsby Charles , Yaseen Nabeel R.
Published: March 2023

Abstract
NUP98-HOXA9, the chimeric protein resulting from the t(7;11)(p15;p15) chromosomal translocation, is a prototype of several NUP98 fusions that occur in myelodysplastic syndromes and acute myeloid leukemia. We examined its effect on differentiation, proliferation, and gene expression in primary human CD34+ hematopoietic cells. Colony-forming cell (CFC) assays in semisolid medium combined with morphologic examination and flow cytometric immunophenotyping revealed that NUP98-HOXA9 increased the numbers of erythroid precursors and impaired both myeloid and erythroid differentiation. In continuous liquid culture, cells transduced with NUP98-HOXA9 exhibited a biphasic growth curve with initial growth inhibition followed by enhanced long-term proliferation, suggesting an increase in the numbers of primitive self-renewing cells. This was confirmed by a dramatic increase in the numbers of long-term culture-initiating cells, the most primitive hematopoietic cells detectable in vitro. To understand the molecular mechanisms underlying the effects of NUP98-HOXA9 on hematopoietic cell proliferation and differentiation, oligonucleotide microarray analysis was done at several time points over 16 days, starting at 6 hours posttransduction. The early growth suppression was preceded by up-regulation of IFNβ1 and accompanied by marked up-regulation of IFN-induced genes, peaking at 3 days posttransduction. In contrast, oncogenes such as homeobox transcription factors, FLT3, KIT, and WT1 peaked at 8 days or beyond, coinciding with increased proliferation. In addition, several putative tumor suppressors and genes associated with hematopoietic differentiation were repressed at later time points. These findings provide a comprehensive picture of the changes in proliferation, differentiation, and global gene expression that underlie the leukemic transformation of human hematopoietic cells by NUP98-HOXA9. (Cancer Res 2006; 66(13): 6628-37)
Data from Loss of the hSNF5 Gene Concomitantly Inactivates p21CIP/WAF1 and p16INK4a Activity Associated with Replicative Senescence in A204 Rhabdoid Tumor Cells
Authors: Chai Jingjing , Charboneau Aubri L. , Betz Bryan L. , Weissman Bernard E.
Published: March 2023

Abstract
hSNF5, the smallest member of the SWI/SNF chromatin remodeling complex, is lost in most malignant rhabdoid tumors (MRT). In MRT cell lines, reexpression of hSNF5 induces G1 cell cycle arrest, elevated p16INK4a, and activated replicative senescence markers, such as β-galactosidase (β-Gal) and plasminogen activator inhibitor-1. To compare the replicative senescence caused by hSNF5 in A204 cells to normal cellular senescence, we examined the activation of both p16INK4a and p21CIP/WAF1. Analogous to normal cellular senescence, both p16INK4a and p21CIP/WAF1 were up-regulated following hSNF5 restoration. Furthermore, we found that hSNF5 bound the p16INK4a and p21CIP/WAF1 promoters, suggesting that it directly regulates transcription of these genes. Using p16INK4a RNA interference, we showed its requirement for the replicative senescence caused by hSNF5 but not the growth arrest. Instead, p21CIP/WAF1 remained activated by hSNF5 in the absence of high p16INK4a expression, apparently causing the growth arrest in A204. Interestingly, we also found that, in the absence of p16INK4a, reexpression of hSNF5 also increased protein levels of a second cyclin-dependent kinase (CDK) inhibitor, p18INK4c. However, our data show that lack of hSNF5 does not abrogate cellular responsiveness to DNA damage or growth-inhibitory factors. In summary, our studies suggest that hSNF5 loss may influence the regulation of multiple CDK inhibitors involved in replicative senescence.
Data from The Synergistic Combination of the Farnesyl Transferase Inhibitor Lonafarnib and Paclitaxel Enhances Tubulin Acetylation and Requires a Functional Tubulin Deacetylase
Authors: Marcus Adam I. , Zhou Jun , O'Brate Aurora , Hamel Ernest , Wong Jason , Nivens Michael , El-Naggar Adel , Yao Tso-Pang , Khuri Fadlo R. , Giannakakou Paraskevi
Published: March 2023

Abstract
Farnesyl transferase (FT) inhibitors (FTI) are anticancer agents developed to target oncogenic Ras proteins by inhibiting Ras farnesylation. FTIs potently synergize with paclitaxel and other microtubule-stabilizing drugs; however, the mechanistic basis underlying this synergistic interaction remains elusive. Here we show that the FTI lonafarnib affects the microtubule cytoskeleton resulting in microtubule bundle formation, increased microtubule stabilization and acetylation, and suppression of microtubule dynamics. Notably, treatment with the combination of low doses of lonafarnib with paclitaxel markedly enhanced tubulin acetylation (a marker of microtubule stability) as compared with either drug alone. This synergistic effect correlated with FT inhibition and was accompanied by a synergistic increase in mitotic arrest and cell death. Mechanistically, we show that the combination of lonafarnib and paclitaxel inhibits the in vitro deacetylating activity of the only known tubulin deacetylase, histone deacetylase 6 (HDAC6). In addition, the lonafarnib/taxane combination is synergistic only in cells lines expressing the wild-type HDAC6, but not a catalytic-mutant HDAC6, revealing that functional HDAC6 is required for the synergy of lonafarnib with taxanes. Furthermore, tubacin, a specific HDAC6 inhibitor, synergistically enhanced tubulin acetylation in combination with paclitaxel, similar to the combination of lonafarnib and paclitaxel. Taken together, these data suggest a relationship between FT inhibition, HDAC6 function, and cell death, providing insight into the putative molecular basis of the lonafarnib/taxane synergistic antiproliferative combination.
Data from The c-Myc Oncogene Directly Induces the H19 Noncoding RNA by Allele-Specific Binding to Potentiate Tumorigenesis
Authors: Barsyte-Lovejoy Dalia , Lau Suzanne K. , Boutros Paul C. , Khosravi Fereshteh , Jurisica Igor , Andrulis Irene L. , Tsao Ming S. , Penn Linda Z.
Published: March 2023

Abstract
The product of the MYC oncogene is widely deregulated in cancer and functions as a regulator of gene transcription. Despite an extensive profile of regulated genes, the transcriptional targets of c-Myc essential for transformation remain unclear. In this study, we show that c-Myc significantly induces the expression of the H19 noncoding RNA in diverse cell types, including breast epithelial, glioblastoma, and fibroblast cells. c-Myc binds to evolutionarily conserved E-boxes near the imprinting control region to facilitate histone acetylation and transcriptional initiation of the H19 promoter. In addition, c-Myc down-regulates the expression of insulin-like growth factor 2 (IGF2), the reciprocally imprinted gene at the H19/IGF2 locus. We show that c-Myc regulates these two genes independently and does not affect H19 imprinting. Indeed, allele-specific chromatin immunoprecipitation and expression analyses indicate that c-Myc binds and drives the expression of only the maternal H19 allele. The role of H19 in transformation is addressed using a knockdown approach and shows that down-regulation of H19 significantly decreases breast and lung cancer cell clonogenicity and anchorage-independent growth. In addition, c-Myc and H19 expression shows strong association in primary breast and lung carcinomas. This work indicates that c-Myc induction of the H19 gene product holds an important role in transformation. (Cancer Res 2006; 66(10): 5330-7)
Data from Notch3 Gene Amplification in Ovarian Cancer
Authors: Park Joon T. , Li Mei , Nakayama Kentaro , Mao Tsui-Lien , Davidson Ben , Zhang Zhen , Kurman Robert J. , Eberhart Charles G. , Shih Ie-Ming , Wang Tian-Li
Published: March 2023

Abstract
Gene amplification is one of the common mechanisms that activate oncogenes. In this study, we used single nucleotide polymorphism array to analyze genome-wide DNA copy number alterations in 31 high-grade ovarian serous carcinomas, the most lethal gynecologic neoplastic disease in women. We identified an amplicon at 19p13.12 in 6 of 31 (19.5%) ovarian high-grade serous carcinomas. This amplification was validated by digital karyotyping, quantitative real-time PCR, and dual-color fluorescence in situ hybridization (FISH) analysis. Comprehensive mRNA expression analysis of all 34 genes within the minimal amplicon identified Notch3 as the gene that showed most significant overexpression in amplified tumors compared with nonamplified tumors. Furthermore, Notch3 DNA copy number is positively correlated with Notch3 protein expression based on parallel immunohistochemistry and FISH studies in 111 high-grade tumors. Inactivation of Notch3 by both γ-secretase inhibitor and Notch3-specific small interfering RNA suppressed cell proliferation and induced apoptosis in the cell lines that overexpressed Notch3 but not in those with minimal amount of Notch3 expression. These results indicate that Notch3 is required for proliferation and survival of Notch3-amplified tumors and inactivation of Notch3 can be a potential therapeutic approach for ovarian carcinomas. (Cancer Res 2006; 66(12): 6312-8)
Data from Mnt-Deficient Mammary Glands Exhibit Impaired Involution and Tumors with Characteristics of Myc Overexpression
Authors: Toyo-oka Kazuhito , Bowen Timothy J. , Hirotsune Shinji , Li Zirong , Jain Sonia , Ota Sara , Lozach Laure Escoubet , Bassett Ivan Garcia , Lozach Jean , Rosenfeld Michael G. , Glass Christopher K. , Eisenman Robert , Ren Bing , Hurlin Peter , Wynshaw-Boris Anthony
Published: March 2023

Abstract
The proto-oncogene c-Myc plays a central role in cell growth and the development of human tumors. c-Myc interacts with Max and Myc-Max complexes bind to E-box and related sequences to activate transcription. Max also interacts with Mnt but Mnt-Max complexes repress transcription when bound to these sequences. MNT maps to human chromosome 17p13.3, a region frequently deleted in various human tumors, including mammary gland tumors. Consistent with the possibility that Mnt functions as a Myc antagonist, Mnt-deficient fibroblasts exhibit many of the hallmark characteristics of cells that overexpress Myc, and conditional (Cre/Lox) inactivation of Mnt in mammary gland epithelium leads to adenocarcinomas. Here, we further characterize mammary gland tissue following conditional deletion of Mnt in the mammary gland. We show that loss of Mnt severely disrupts mammary gland involution and leads to hyperplastic ducts associated with reduced numbers of apoptotic cells. These findings suggest that loss of Mnt in mammary tissue has similarities to Myc overexpression. We tested this directly by using promoter array analysis and mRNA expression analysis by oligonucleotide arrays. We found that Mnt and c-Myc bound to similar promoters in tumors from MMTV-c-Myc transgenic mice, and mRNA expression patterns were similar between mammary tumors from MMTV-Cre/MntKO/CKO and MMTV-c-Myc transgenic mice. These results reveal an important role for Mnt in pregnancy-associated mammary gland development and suggest that mammary gland tumorigenesis in the absence of Mnt is analogous to that caused by Myc deregulation. (Cancer Res 2006; 66(11): 5565-73)
Data from Deficiency in the Repair of DNA Damage by Homologous Recombination and Sensitivity to Poly(ADP-Ribose) Polymerase Inhibition
Authors: McCabe Nuala , Turner Nicholas C. , Lord Christopher J. , Kluzek Katarzyna , Białkowska Aneta , Swift Sally , Giavara Sabrina , O'Connor Mark J. , Tutt Andrew N. , Zdzienicka Małgorzata Z. , Smith Graeme C.M. , Ashworth Alan
Published: March 2023

Abstract
Deficiency in either of the breast cancer susceptibility proteins BRCA1 or BRCA2 induces profound cellular sensitivity to the inhibition of poly(ADP-ribose) polymerase (PARP) activity. We hypothesized that the critical role of BRCA1 and BRCA2 in the repair of double-strand breaks by homologous recombination (HR) was the underlying reason for this sensitivity. Here, we examine the effects of deficiency of several proteins involved in HR on sensitivity to PARP inhibition. We show that deficiency of RAD51, RAD54, DSS1, RPA1, NBS1, ATR, ATM, CHK1, CHK2, FANCD2, FANCA, or FANCC induces such sensitivity. This suggests that BRCA-deficient cells are, at least in part, sensitive to PARP inhibition because of HR deficiency. These results indicate that PARP inhibition might be a useful therapeutic strategy not only for the treatment of BRCA mutation-associated tumors but also for the treatment of a wider range of tumors bearing a variety of deficiencies in the HR pathway or displaying properties of ‘BRCAness.’ (Cancer Res 2006; 66(16): 8109-15)
Data from The ERBB4/HER4 Intracellular Domain 4ICD Is a BH3-Only Protein Promoting Apoptosis of Breast Cancer Cells
Authors: Naresh Anjali , Long Weiwen , Vidal Gregory A. , Wimley William C. , Marrero Luis , Sartor Carolyn I. , Tovey Sian , Cooke Timothy G. , Bartlett John M.S. , Jones Frank E.
Published: March 2023

Abstract
ERBB4/HER4 (referred to here as ERBB4) is a unique member of the epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases. In contrast to the other three members of the EGFR family (i.e., EGFR, ERBB2/HER2/NEU, and ERBB3), which are associated with aggressive forms of human cancers, ERBB4 expression seems to be selectively lost in tumors with aggressive phenotypes. Consistent with this observation, we show that ERBB4 induces apoptosis when reintroduced into breast cancer cell lines or when endogenous ERBB4 is activated by a ligand. We further show that ligand activation and subsequent proteolytic processing of endogenous ERBB4 results in mitochondrial accumulation of the ERBB4 intracellular domain (4ICD) and cytochrome c efflux, the essential and committed step of mitochondrial regulated apoptosis. Our results indicate that 4ICD is functionally similar to BH3-only proteins, proapoptotic members of the BCL-2 family required for initiation of mitochondrial dysfunction through activation of the proapoptotic multi-BH domain proteins BAX/BAK. Similar to other BH3-only proteins, 4ICD cell-killing activity requires an intact BH3 domain and 4ICD interaction with the antiapoptotic protein BCL-2, suppressed 4ICD-induced apoptosis. Unique among BH3-only proteins, however, is the essential requirement of BAK but not BAX to transmit the 4ICD apoptotic signal. Clinically, cytosolic but not membrane ERBB4/4ICD expression in primary human breast tumors was associated with tumor apoptosis, providing a mechanistic explanation for the loss of ERBB4 expression during tumor progression. Thus, we propose that ligand-induced mitochondrial accumulation of 4ICD represents a unique mechanism of action for transmembrane receptors, directly coupling a cell surface signal to the tumor cell mitochondrial apoptotic pathway. (Cancer Res 2006; 66(12): 6412-20)
Data from Gene Expression Signature in Organized and Growth-Arrested Mammary Acini Predicts Good Outcome in Breast Cancer
Authors: Fournier Marcia V. , Martin Katherine J. , Kenny Paraic A. , Xhaja Kris , Bosch Irene , Yaswen Paul , Bissell Mina J.
Published: March 2023

Abstract
Nonmalignant human mammary epithelial cells (HMEC) seeded in laminin-rich extracellular matrix (lrECM) form polarized acini and, in doing so, transit from a disorganized proliferating state to an organized growth-arrested state. We hypothesized that the gene expression pattern of organized and growth-arrested HMECs would share similarities with breast tumors with good prognoses. Using Affymetrix HG-U133A microarrays, we analyzed the expression of 22,283 gene transcripts in 184 (finite life span) and HMT3522 S1 (immortal nonmalignant) HMECs on successive days after seeding in a lrECM assay. Both HMECs underwent growth arrest in G0-G1 and differentiated into polarized acini between days 5 and 7. We identified gene expression changes with the same temporal pattern in both lines and examined the expression of these genes in a previously published panel of microarray data for 295 breast cancer samples. We show that genes that are significantly lower in the organized, growth-arrested HMEC than in their proliferating counterparts can be used to classify breast cancer patients into poor and good prognosis groups with high accuracy. This study represents a novel unsupervised approach to identifying breast cancer markers that may be of use clinically. (Cancer Res 2006; 66(14): 7095-102)
Data from AIMP3 Haploinsufficiency Disrupts Oncogene-Induced p53 Activation and Genomic Stability
Authors: Park Bum-Joon , Oh Young Sun , Park Seung Yong , Choi So Jung , Rudolph Cornelia , Schlegelberger Brigitte , Kim Sunghoon
Published: March 2023

Abstract
AIMP3 (previously known as p18) was shown to up-regulate p53 in response to DNA damage. Here, we show that AIMP3 couples oncogenic stresses to p53 activation to prevent cell transformation. Growth factor– or Ras-dependent induction of p53 was blocked by single allelic loss of AIMP3 as well as by suppression of AIMP3. AIMP3 heterozygous cells became susceptible to cell transformation induced by oncogenes such as Ras or Myc alone. The transformed AIMP3+/− cells showed severe abnormality in cell division and chromosomal structure. Thus, AIMP3 plays crucial roles in p53-mediated tumor-suppressive response against oncogenic stresses via differential activation of ATM and ATR, and in the maintenance of genomic stability. (Cancer Res 2006; 66(14): 6913-8)
Data from Predominant Bcl-XL Knockdown Disables Antiapoptotic Mechanisms: Tumor Necrosis Factor–Related Apoptosis-Inducing Ligand–Based Triple Chemotherapy Overcomes Chemoresistance in Pancreatic Cancer Cells In vitro
Authors: Bai Jirong , Sui Jianhua , Demirjian Aram , Vollmer Charles M. , Marasco Wayne , Callery Mark P.
Published: March 2023

Abstract
Pancreatic cancer is lethal because of its invasiveness, rapid progression, and profound resistance to chemotherapy and radiation therapy. To identify the molecular mechanisms underlying this, we have examined the expression and potency of three major death receptors: tumor necrosis factor receptor (TNF-R), TNF-related apoptosis-inducing ligand receptor (TRAIL-R), and Fas in mediating cytotoxicity in four invasive pancreatic cancer cell lines. We have analyzed the expression of major antiapoptotic factors, cell cycle regulators and death receptor decoys (DcR) in comparison with normal pancreas tissues and five other human malignant tumor cell lines. We have found that different pancreatic cancer cell lines coexpress high-level TRAIL-R, Fas, and TNF-R1 but are strongly resistant to apoptosis triggered by the death receptors. DcR2 and DcR3 overexpression may partly contribute to the resistance of pancreatic cancer cells to TRAIL-R– and Fas-mediated cytotoxicity. Bcl-XL and Bcl-2 are predominantly overexpressed in pancreatic cancer cell lines, respectively. Bcl-XL is also predominantly overexpressed in prostate, colorectal, and intestinal cancer cells. The knockdown of the predominant Bcl-XL overexpression significantly reduces the viability of pancreatic cancer cells to TNFα- and TRAIL-mediated apoptosis by sublethal-dose single and combined antitumor drugs, including geldanamycin, PS-341, Trichostatin A, and doxorubicine. Geldanamyin and PS-341 synergistically block NFκB activation, suppress Akt/PKB pathway, and down-regulate Bcl-XL, Bcl-2, cIAP-1, and cyclin D1 expression. This combined regimen dramatically enhances TRAIL cytotoxic effects and breaks through chemoresistance. Bcl-XL plays a vital role in pancreatic cancer chemoresistance. Geldanamycin, PS-341, and TRAIL triple combination may be a novel therapeutic strategy for pancreatic cancer.
Data from p53 Is Preferentially Recruited to the Promoters of Growth Arrest Genes p21 and GADD45 during Replicative Senescence of Normal Human Fibroblasts
Authors: Jackson James G. , Pereira-Smith Olivia M.
Published: March 2023

Abstract
Replicative senescence is the terminal growth arrest that most normal human cells enter into after a fixed number of divisions in vitro, limiting the proliferative potential of a cell and preventing genomic instability caused by critically short telomeres. Thus, senescence presents a tumor-suppressive mechanism and a barrier to tumor formation. However, senescent cells are inherently resistant to apoptosis and, as they accumulate in aging tissues, may contribute to organ dysfunction and promote tumor progression as part of the stromal environment. Replicative life span in normal human cells can be extended by inactivation of the tumor suppressor gene p53 or its direct target, the cyclin-dependent kinase inhibitor p21, suggesting a direct role for this pathway in senescence. However, p53 recruitment to promoters of target genes during replicative senescence has not been shown in live cells. In this study, we used chromatin immunoprecipitation to determine that p53 preferentially occupied the promoters of growth arrest genes p21 and GADD45 in senescent normal human diploid fibroblasts but not the promoters of other target genes that recruited p53 following doxorubicin-induced DNA damage, such as apoptosis regulators TNFRSF10b, TNFRSF6, and PUMA. This differential recruitment of p53 in senescent versus doxorubicin-treated fibroblasts was accompanied by differences in post-translational modification of p53. These data provide mechanisms for both the growth arrest mediated by p53 and the resistant nature of senescent cells to apoptosis despite p53 activity. (Cancer Res 2006; 66(17): 8356-60)
Data from Development of Ewing's Sarcoma from Primary Bone Marrow–Derived Mesenchymal Progenitor Cells
Authors: Riggi Nicolò , Cironi Luisa , Provero Paolo , Suvà Mario-Luca , Kaloulis Konstantinos , Garcia-Echeverria Carlos , Hoffmann Francesco , Trumpp Andreas , Stamenkovic Ivan
Published: March 2023

Abstract
Ewing's sarcoma is a member of Ewing's family tumors (EFTs) and the second most common solid bone and soft tissue malignancy of children and young adults. It is associated in 85% of cases with the t(11;22)(q24:q12) chromosomal translocation that generates fusion of the 5′ segment of the EWS gene with the 3′ segment of the ETS family gene FLI-1. The EWS-FLI-1 fusion protein behaves as an aberrant transcriptional activator and is believed to contribute to EFT development. However, EWS-FLI-1 induces growth arrest and apoptosis in normal fibroblasts, and primary cells that are permissive for its putative oncogenic properties have not been discovered, hampering basic understanding of EFT biology. Here, we show that EWS-FLI-1 alone can transform primary bone marrow–derived mesenchymal progenitor cells and generate tumors that display hallmarks of Ewing's sarcoma, including a small round cell phenotype, expression of EFT-associated markers, insulin like growth factor-I dependence, and induction or repression of numerous EWS-FLI-1 target genes. These observations provide the first identification of candidate primary cells from which EFTs originate and suggest that EWS-FLI-1 expression may constitute the initiating event in EFT pathogenesis. (Cancer Res 2005; 65(24): 11459-68)
Data from Evidence for the Presence of Disease-Perturbed Networks in Prostate Cancer Cells by Genomic and Proteomic Analyses: A Systems Approach to Disease
Authors: Lin Biaoyang , White James T. , Lu Wei , Xie Tao , Utleg Angelita G. , Yan Xiaowei , Yi Eugene C. , Shannon Paul , Khrebtukova Irina , Lange Paul H. , Goodlett David R. , Zhou Daixing , Vasicek Thomas J. , Hood Leroy
Published: March 2023

Abstract
Prostate cancer is initially responsive to androgen ablation therapy and progresses to androgen-unresponsive states that are refractory to treatment. The mechanism of this transition is unknown. A systems approach to disease begins with the quantitative delineation of the informational elements (mRNAs and proteins) in various disease states. We employed two recently developed high-throughput technologies, massively parallel signature sequencing (MPSS) and isotope-coded affinity tag, to gain a comprehensive picture of the changes in mRNA levels and more restricted analysis of protein levels, respectively, during the transition from androgen-dependent LNCaP (model for early-stage prostate cancer) to androgen-independent CL1 cells (model for late-stage prostate cancer). We sequenced >5 million MPSS signatures, obtained >142,000 tandem mass spectra, and built comprehensive MPSS and proteomic databases. The integrated mRNA and protein expression data revealed underlying functional differences between androgen-dependent and androgen-independent prostate cancer cells. The high sensitivity of MPSS enabled us to identify virtually all of the expressed transcripts and to quantify the changes in gene expression between these two cell states, including functionally important low-abundance mRNAs, such as those encoding transcription factors and signal transduction molecules. These data enable us to map the differences onto extant physiologic networks, creating perturbation networks that reflect prostate cancer progression. We found 37 BioCarta and 14 Kyoto Encyclopedia of Genes and Genomes pathways that are up-regulated and 23 BioCarta and 22 Kyoto Encyclopedia of Genes and Genomes pathways that are down-regulated in LNCaP cells versus CL1 cells. Our efforts represent a significant step toward a systems approach to understanding prostate cancer progression.
Data from Cooperation between Cdk4 and p27kip1 in Tumor Development: A Preclinical Model to Evaluate Cell Cycle Inhibitors with Therapeutic Activity
Authors: Sotillo Rocío , Renner Oliver , Dubus Pierre , Ruiz-Cabello Jesús , Martín-Caballero Juan , Barbacid Mariano , Carnero Amancio , Malumbres Marcos
Published: March 2023

Abstract
Deregulation of the G1-S transition of the cell cycle is a common feature of human cancer. Tumor-associated alterations in this process frequently affect cyclin-dependent kinases (Cdk), their regulators (cyclins, INK4 inhibitors, or p27Kip1), and their substrates (retinoblastoma protein). Although these proteins are generally thought to act in a linear pathway, mutations in different components frequently cooperate in tumor development. Using gene-targeted mouse models, we report in this article that Cdk4 resistance to INK4 inhibitors, due to the Cdk4 R24C mutation, strongly cooperates with p27Kip1 deficiency in tumor development. No such cooperation is observed between Cdk4 R24C and p18INK4c absence, suggesting that the only function of p18INK4c is inhibiting Cdk4 in this model. Cdk4R/R knock in mice, which express the Cdk4 R24C mutant protein, develop pituitary tumors with complete penetrance and short latency in a p27Kip1−/− or p27Kip1+/− background. We have investigated whether this tumor model could be useful to assess the therapeutic activity of cell cycle inhibitors. We show here that exposure to flavopiridol, a wide-spectrum Cdk inhibitor, significantly delays tumor progression and leads to tumor-free survival in a significant percentage of treated mice. These data suggest that genetically engineered tumor models involving key cell cycle regulators are a valuable tool to evaluate drugs with potential therapeutic benefit in human cancer.
Data from Sensitization of Human Carcinoma Cells to Alkylating Agents by Small Interfering RNA Suppression of 3-Alkyladenine-DNA Glycosylase
Authors: Paik Johanna , Duncan Tod , Lindahl Tomas , Sedgwick Barbara
Published: March 2023

Abstract
One of the major cytotoxic lesions generated by alkylating agents is DNA 3-alkyladenine, which can be excised by 3-alkyladenine DNA glycosylase (AAG). Inhibition of AAG may therefore result in increased cellular sensitivity to chemotherapeutic alkylating agents. To investigate this possibility, we have examined the role of AAG in protecting human tumor cells against such agents. Plasmids that express small interfering RNAs targeted to two different regions of AAG mRNA were transfected into HeLa cervical carcinoma cells and A2780-SCA ovarian carcinoma cells. Stable derivatives of both cell types with low AAG protein levels were sensitized to alkylating agents. Two HeLa cell lines with AAG protein levels reduced by at least 80% to 90% displayed a 5- to 10-fold increase in sensitivity to methyl methanesulfonate, N-methyl-N-nitrosourea, and the chemotherapeutic drugs temozolomide and 1,3-bis(2-chloroethyl)-1-nitrosourea. These cells showed no increase in sensitivity to UV light or ionizing radiation. After treatment with methyl methanesulfonate, AAG knockdown HeLa cells were delayed in S phase but accumulated in G2-M. Our data support the hypothesis that ablation of AAG activity in human tumor cells may provide a useful strategy to enhance the efficacy of current chemotherapeutic regimens that include alkylating agents.
Data from A Gene Expression Signature of Genetic Instability in Colon Cancer
Authors: Giacomini Craig P. , Leung Suet Yi , Chen Xin , Yuen Siu Tsan , Kim Young H. , Bair Eric , Pollack Jonathan R.
Published: March 2023

Abstract
Genetic instability plays a central role in the development and progression of human cancer. Two major classes of genetic instability, microsatellite instability (MSI) and chromosome instability (microsatellite stable; MSS), are best understood in the context of colon cancer, where MSI tumors represent ∼15% of cases, and compared with MSS tumors, more often arise in the proximal colon and display favorable clinical outcome. To further explore molecular differences, we profiled gene expression in a set of 18 colon cancer cell lines using cDNA microarrays representing ∼21,000 different genes. Supervised analysis identified a robust expression signature distinguishing MSI and MSS samples. As few as eight genes predicted with high accuracy the underlying genetic instability in the original and in three independent sample sets, comprising 13 colon cancer cell lines, 61 colorectal tumors, and 87 gastric tumors. Notably, the MSI signature was retained despite genetically correcting the underlying instability, suggesting the signature reflects a legacy of the tumor having arisen from MSI, rather than sensing the ongoing state of MSI. Our findings support a model in which MSI and MSS preferentially target different genes and pathways in cancer. Further, among the MSI signature genes, our findings implicate a role of elevated metallothionein expression in the clinical behavior of MSI cancers.
Data from BRCA1 Promotes Induction of ssDNA by Ionizing Radiation
Authors: Schlegel Brian P. , Jodelka Francine M. , Nunez Rafael
Published: March 2023

Abstract
The BRCA1 tumor suppressor contributes to the repair of DNA double-strand breaks (DSB) through homologous recombination, but the mechanism is unknown. The rapid accumulation of BRCA1 into nuclear foci in response to induction of DNA breaks suggests that BRCA1 may function in an early step in the repair pathway. We examined the role of BRCA1 in one such early step, the resection of DSBs to generate ssDNA. The appearance of ssDNA in response to ionizing radiation is similar to that of BRCA1 foci formation, suggesting that the two processes are related. Furthermore, BRCA1 colocalizes to ssDNA sites induced by ionizing radiation. Overexpression of BRCA1 causes an increase in cells exhibiting ssDNA induced by ionizing radiation. Mutant BRCA1 that lacks the COOH-terminal BRCT domain also promotes ssDNA but fails to form nuclear foci. Knockdown of BRCA1 expression reduces ssDNA and Rad51 foci formation in response to ionizing radiation. These results indicate that BRCA1 promotes induction of ssDNA in response to ionizing radiation and accumulates at sites of ssDNA. (Cancer Res 2006; 66(10): 5181-9)
Data from Janus Kinase 2: A Critical Target in Chronic Myelogenous Leukemia
Authors: Samanta Ajoy K. , Lin Hui , Sun Tong , Kantarjian Hagop , Arlinghaus Ralph B.
Published: March 2023

Abstract
The Bcr-Abl tyrosine kinase is the causative factor in most chronic myelogenous leukemia (CML) patients. We have shown that Bcr-Abl is associated with a cluster of signaling proteins, including Janus kinase (Jak) 2, growth factor receptor binding protein 2–associated binder (Gab) 2, Akt, and glycogen synthase kinase (GSK)-3β. Treatment of CML cell lines and mouse Bcr-Abl+ 32D cells with either Jak2 short interfering RNA or Jak2 kinase inhibitor AG490 inhibited pTyr Gab2 and pSer Akt formation, inhibited the activation of nuclear factor-κB, and caused the activation of GSK-3β, leading to the reduction of c-Myc. Importantly, BaF3 cells expressing T315I and E255K imatinib-resistant mutants of Bcr-Abl underwent apoptosis on exposure to AG490 yet were resistant to imatinib. Similar to wild-type Bcr-Abl+ cells, inhibition of Jak2 by Ag490 treatment resulted in decrease of pSer Akt and c-Myc in imatinib-resistant cells. These results identify Jak2 as a potentially important therapeutic target for CML. (Cancer Res 2006; 66(13): 6468-72)
Data from IQGAP1 Protein Specifies Amplifying Cancer Cells in Glioblastoma Multiforme
Authors: Balenci Laurent , Clarke Ian D. , Dirks Peter B. , Assard Nicole , Ducray François , Jouvet Anne , Belin Marie-Françoise , Honnorat Jerôme , Baudier Jacques
Published: March 2023

Abstract
The accurate identification and thorough characterization of tumorigenic cells in glioblastomas are essential to enhance our understanding of their malignant behavior and for the design of strategies that target this important cell population. We report here that, in rat brain, the scaffolding protein IQGAP1 is a marker of brain nestin+ amplifying neural progenitor cells. In a rat model of glioma, IQGAP1 also characterizes a subpopulation of nestin+ amplifying tumor cells in glioblastoma-like tumors but not in tumors with oligodendroglioma features. We next confirmed that IQGAP1 represents a new marker that may help to discriminate human glioblastoma from oligodendrogliomas. In human glioblastoma exclusively, IQGAP1 specifies a subpopulation of amplifying nestin+ cancer cells. Neoplastic IQGAP1+ cells from glioblastoma can be expanded in culture and possess all the characteristics of cancer stem-like progenitors. The similarities between amplifying neural progenitors and glioblastoma amplifying cancer cells may have significant implications for understanding the biology of glioblastoma. (Cancer Res 2006; 66(18): 9074-82)
Data from A Role for the WWOX Gene in Prostate Cancer
Authors: Qin Haiyan R. , Iliopoulos Dimitrios , Semba Shuho , Fabbri Muller , Druck Teresa , Volinia Stefano , Croce Carlo M. , Morrison Carl D. , Klein Russell D. , Huebner Kay
Published: March 2023

Abstract
Expression of the WWOX gene, encompassing the common chromosome fragile site FRA16D, is altered in a large fraction of cancers of various types, including prostate cancer. We have examined expression and biological functions of WWOX in prostate cancer. WWOX mRNA and protein expression were significantly reduced in prostate cancer-derived cells (LNCaP, DU145, and PC-3) compared with noncancer prostate cells (PWR-1E), and WWOX expression was reduced in 84% of prostate cancers, as assessed by immunohistochemical staining. Down-modulation of WWOX expression in the prostate cancer-derived cells is due to DNA hypermethylation in the WWOX regulatory region. Treatment with 5-aza-2′-deoxycytidine (AZA), a DNA methyltransferase inhibitor, and trichostatin A, a histone deacetylase inhibitor, led to increased WWOX mRNA and protein expression in prostate cancer-derived cells, most strikingly in DU145 cells. Transfection-mediated WWOX overexpression in DU145 cells suppressed colony growth (P = 0.0012), and WWOX overexpression by infection with Ad-WWOX virus induced apoptosis through a caspase-dependent mechanism and suppressed cell growth. Lastly, ectopic expression of WWOX by Ad-WWOX infection suppressed tumorigenicity of xenografts in nude mice, and intratumoral AZA treatment halted tumor growth. The data are consistent with a role for WWOX as a prostate cancer tumor suppressor and suggest that WWOX signal pathways should be further investigated in normal and cancerous prostate cells and tissues. (Cancer Res 2006; 66(13): 6477-81)
Data from Integrative Genomics Identifies Distinct Molecular Classes of Neuroblastoma and Shows That Multiple Genes Are Targeted by Regional Alterations in DNA Copy Number
Authors: Wang Qun , Diskin Sharon , Rappaport Eric , Attiyeh Edward , Mosse Yael , Shue Daniel , Seiser Eric , Jagannathan Jayanti , Shusterman Suzanne , Bansal Manisha , Khazi Deepa , Winter Cynthia , Okawa Erin , Grant Gregory , Cnaan Avital , Zhao Huaqing , Cheung Nai-Kong , Gerald William , London Wendy , Matthay Katherine K. , Brodeur Garrett M. , Maris John M.
Published: March 2023

Abstract
Neuroblastoma is remarkable for its clinical heterogeneity and is characterized by genomic alterations that are strongly correlated with tumor behavior. The specific genes that influence neuroblastoma biology and are targeted by genomic alterations remain largely unknown. We quantified mRNA expression in a highly annotated series of 101 prospectively collected diagnostic neuroblastoma primary tumors using an oligonucleotide-based microarray. Genomic copy number status at the prognostically relevant loci 1p36, 2p24 (MYCN), 11q23, and 17q23 was determined by PCR and was aberrant in 26, 20, 40, and 38 cases, respectively. In addition, 72 diagnostic neuroblastoma primary tumors assayed in a different laboratory were used as an independent validation set. Unsupervised hierarchical clustering showed that gene expression was highly correlated with genomic alterations and clinical markers of tumor behavior. The vast majority of samples with MYCN amplification and 1p36 loss of heterozygosity (LOH) clustered together on a terminal node of the sample dendrogram, whereas the majority of samples with 11q deletion clustered separately and both of these were largely distinct from the copy number neutral group of tumors. Genes involved in neurodevelopment were broadly overrepresented in the more benign tumors, whereas genes involved in RNA processing and cellular proliferation were highly represented in the most malignant cases. By combining transcriptomic and genomic data, we showed that LOH at 1p and 11q was associated with significantly decreased expression of 122 (61%) and 88 (27%) of the genes mapping to 1p35-36 and all of 11q, respectively, suggesting that multiple genes may be targeted by LOH events. A total of 71 of the 1p35-36 genes were also differentially expressed in the independent validation data set, providing a prioritized list of candidate neuroblastoma suppressor genes. Taken together, these data are consistent with the hypotheses that the neuroblastoma transcriptome is a sensitive marker of underlying tumor biology and that chromosomal deletion events in this cancer likely target multiple genes through alteration in mRNA dosage. Lead positional candidates for neuroblastoma suppressor genes can be inferred from these data, but the potential multiplicity of transcripts involved has significant implications for ongoing gene discovery strategies. (Cancer Res 2006; 66(12): 6050-62)
Data from Absence of the Full-Length Breast Cancer–Associated Gene-1 Leads to Increased Expression of Insulin-Like Growth Factor Signaling Axis Members
Authors: Shukla Vivek , Coumoul Xavier , Cao Liu , Wang Rui-Hong , Xiao Cuiying , Xu Xiaoling , Andò Sebastiano , Yakar Shoshana , LeRoith Derek , Deng Chuxia
Published: March 2023

Abstract
The breast cancer–associated gene-1 (BRCA1) plays many important functions in multiple biological processes/pathways. Mice homozygous for a targeted deletion of full-length BRCA1 (Brca1Δ11/Δ11) display both increased tumorigenesis and premature aging, yet molecular mechanisms underlying these defects remain elusive. Here, we show that Brca1 deficiency leads to increased expression of several insulin-like growth factor (IGF) signaling axis members in multiple experimental systems, including BRCA1-deficient mice, primary mammary tumors, and cultured human cells. Furthermore, we provide evidence that activation of IGF signaling by BRCA1 deficiency can also occur in a p53-independent fashion. Our data indicate that BRCA1 interacts with the IRS-1 promoter and inhibits its activity that is associated with epigenetic modification of histone H3 and histone H4 to a transcriptional repression chromatin configuration. We further show that BRCA1-deficient mammary tumor cells exhibit high levels of IRS-1, and acute suppression of Irs-1 using RNA interference significantly inhibits growth of these cells. Those observations provide a molecular insight in understanding both fundamental and therapeutic BRCA1-associated tumorigenesis and aging. (Cancer Res 2006; 66(14): 7151-7)
Data from Shared Epigenetic Mechanisms in Human and Mouse Gliomas Inactivate Expression of the Growth Suppressor SLC5A8
Authors: Hong Chibo , Maunakea Alika , Jun Peter , Bollen Andrew W. , Hodgson J. Graeme , Goldenberg David D. , Weiss William A. , Costello Joseph F.
Published: March 2023

Abstract
Tumors arise in part from the deleterious effects of genetic and epigenetic mechanisms on gene expression. In several mouse models of human tumors, the tumorigenic phenotype is reversible, suggesting that epigenetic mechanisms also contribute significantly to tumorigenesis in mice. It is not known whether these are the same epigenetic mechanisms in human and mouse tumors or whether they affect homologous genes. Using an integrated approach for genome-wide methylation and copy number analyses, we identified SLC5A8 on chromosome 12q23.1 that was affected frequently by aberrant methylation in human astrocytomas and oligodendrogliomas. SLC5A8 encodes a sodium monocarboxylate cotransporter that was highly expressed in normal brain but was significant down-regulated in primary gliomas. Bisulfite sequencing analysis showed that the CpG island was unmethylated in normal brain but frequently localized methylated in brain tumors, consistent with the tumor-specific loss of gene expression. In glioma cell lines, SLC5A8 expression was also suppressed but could be reactivated with a methylation inhibitor. Expression of exogenous SLC5A8 in LN229 and LN443 glioma cells inhibited colony formation, suggesting that it may function as a growth suppressor in normal brain cells. Remarkably, 9 of 10 murine oligodendroglial tumors (from p53+/− or ink4a/arf+/− animals transgenic for S100β-v-erbB) showed a similar tumor-specific down-regulation of mSLC5A8, the highly conserved mouse homologue. Taken together, these data suggest that SLC5A8 functions as a growth suppressor gene in vitro and that it is silenced frequently by epigenetic mechanisms in primary gliomas. The shared epigenetic inactivation of mSLC5A8 in mouse gliomas indicates an additional degree of commonality in the origin and/or pathway to tumorigenesis between primary human tumors and these mouse models of gliomas.
Data from Extracellular Matrix–Mediated Membrane-Type 1 Matrix Metalloproteinase Expression in Pancreatic Ductal Cells Is Regulated by Transforming Growth Factor-β1
Authors: Ottaviano Adam J. , Sun Limin , Ananthanarayanan Vijayalakshmi , Munshi Hidayatullah G.
Published: March 2023

Abstract
Pancreatic ductal adenocarcinoma (PDAC) is associated with an intense fibrotic reaction around the tumor known as desmoplastic reaction. This tissue is composed of interstitial matrix, predominantly type I collagen, together with proliferating fibroblastic cells. Despite the recognized importance of tumor-stromal interactions, very little is known about the interactions among pancreatic cells, myofibroblasts, and the interstitial matrix. The current study was undertaken to test the hypothesis that the desmoplastic reaction alters PDAC gene expression and cellular behavior. Evaluation of human pancreatic specimens showed increased fibrosis and enhanced membrane type 1-matrix metalloproteinase (MT1-MMP) expression in tumor specimens compared with normal pancreas. Using an in vitro model of tumor cell-stromal interactions, type I collagen and the extracellular matrix deposited by pancreatic fibroblasts and PDAC cells regulated motility of human papillomavirus–immortalized human pancreatic ductal epithelial (HPDE) cells. These “stromal” matrices also regulated MT1-MMP expression by HPDE cells, without affecting the expression of tissue inhibitor of metalloproteinase 2. Treatment with transforming growth factor-β1 (TGF-β1) type I receptor kinase inhibitors and function-blocking anti-TGF-β1 antibody abrogated matrix-mediated MT1-MMP induction. TGF-β1 also promoted MT1-MMP–dependent migration by HPDE cells. Moreover, compared with normal tissue, there was increased TGF-β1 signaling in grade 3 tumor specimens as shown by increased phospho-Smad2 staining. These data show that the crosstalk between cancer cells and stromal elements mediated by TGF-β1 influences cell surface– and pericellular matrix–degrading potential in vitro and may contribute to pancreatic cancer progression in vivo. (Cancer Res 2006; 66(14): 7032-40)
Data from Spontaneous Feline Mammary Carcinoma Is a Model of HER2 Overexpressing Poor Prognosis Human Breast Cancer
Authors: De Maria Raffaella , Olivero Martina , Iussich Selina , Nakaichi Munekazu , Murata Tomoaki , Biolatti Bartolomeo , Di Renzo Maria Flavia
Published: March 2023

Abstract
Companion animal spontaneous tumors are suitable models for human cancer, primarily because both animal population and the tumors are genetically heterogeneous. Feline mammary carcinoma (FMC) is a highly aggressive, mainly hormone receptor–negative cancer, which has been proposed as a model for poor prognosis human breast cancer. We have identified and studied the feline orthologue of the HER2 gene, which is both an important prognostic marker and therapeutic target in human cancer. Feline HER2 (f-HER2) gene kinase domain is 92% similar to the human HER2 kinase. F-HER2–specific mRNA was found 3- to 18-fold increased in 3 of 3 FMC cell lines, in 1 of 4 mammary adenomas and 6 of 11 FMC samples using quantitative reverse transcription-PCR. Western blot showed that an anti-human HER2 antibody recognized a protein comigrating with the human p185HER2 in FMC cell lines. The same antibodies strongly stained 13 of 36 FMC archival samples. These data show that feline HER2 overexpression qualifies FMC as homologous to the subset of HER2 overexpressing, poor prognosis human breast carcinomas and as a suitable model to test innovative approaches to therapy of aggressive tumors.