bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Receptor-Mediated DNA-Targeted Photoimmunotherapy
Authors: Karagiannis Tom C. , Lobachevsky Pavel N. , Leung Brenda K.Y. , White Jonathan M. , Martin Roger F.
Published: March 2023

Abstract
We show the efficacy of a therapeutic strategy that combines the potency of a DNA-binding photosensitizer, UVASens, with the tumor-targeting potential of receptor-mediated endocytosis. The photosensitizer is an iodinated bibenzimidazole, which, when bound in the minor groove of DNA and excited by UVA irradiation, induces cytotoxic lesions attributed to a radical species resulting from photodehalogenation. Although reminiscent of photochemotherapy using psoralens and UVA irradiation, an established treatment modality in dermatology particularly for the treatment of psoriasis and cutaneous T-cell lymphoma, a critical difference is the extreme photopotency of the iodinated bibenzimidazole, ∼1,000-fold that of psoralens. This feature prompted consideration of combination with the specificity of receptor-mediated targeting. Using two in vitro model systems, we show the UVA cytotoxicity of iodo ligand/protein conjugates, implying binding of the conjugate to cell receptors, internalization, and degradation of the conjugate-receptor complex, with release and translocation of the ligand to nuclear DNA. For ligand-transferrin conjugates, phototoxicity was inhibited by coincubation with excess native transferrin. Receptor-mediated UVA-induced cytotoxicity was also shown with the iodo ligand conjugate of an anti-human epidermal growth factor receptor monoclonal antibody, exemplifying the potential application of the strategy to other cancer-specific targets to thus improve the specificity of phototherapy of superficial lesions and for extracorporeal treatments. (Cancer Res 2006; 66(21): 10548-52)
Data from Hematopoietic Prostaglandin D Synthase Suppresses Intestinal Adenomas in ApcMin/+ Mice
Authors: Park Jae Man , Kanaoka Yoshihide , Eguchi Naomi , Aritake Kosuke , Grujic Sava , Materi Alicia M. , Buslon Virgilio S. , Tippin Brigette L. , Kwong Alan M. , Salido Eduardo , French Samuel W. , Urade Yoshihiro , Lin Henry J.
Published: March 2023

Abstract
Aspirin and other nonsteroidal anti-inflammatory drugs prevent some cases of colon cancer by inhibiting prostaglandin (PG) synthesis. PGE2 promotes colon neoplasia, as shown by knockout mouse studies on enzymes and receptors in the PG cascade. A few experiments 20 to 30 years ago suggested that PGD2 may suppress tumors, but a role for biosynthetic enzymes for PGD2 in tumor development has not been studied. We report here that disruption of the gene for hematopoietic PGD synthase in ApcMin/+ mice led to ∼50% more intestinal adenomas compared with controls. Tumor size was not affected. By immunohistochemistry, we detected hematopoietic PGD synthase mainly in macrophages and monocytes of the gut mucosa. The mean number of tumors did not increase with knockout of the gene for the lipocalin type of the enzyme, which is not produced in the intestine. On the other hand, ApcMin/+ mice with transgenic human hematopoietic PGD synthase tended to have 80% fewer intestinal adenomas. The transgene produced high mRNA levels (375-fold over endogenous). There was a suggestion of higher urinary excretion of 11β-PGF2α and a lower excretion of a PGE2 metabolite in transgenic mice, but differences (30–40%) were not statistically significant. The results support an interpretation that hematopoietic PGD synthase controls an inhibitory effect on intestinal tumors. Further studies will be needed to prove possible mechanisms, such as routing of PG production away from protumorigenic PGE2 or inhibition of the nuclear factor-κB cascade by PGD2 metabolites. [Cancer Res 2007;67(3):881–9]
Data from Activation of Human Long Interspersed Nuclear Element 1 Retrotransposition by Benzo(a)pyrene, an Ubiquitous Environmental Carcinogen
Authors: Stribinskis Vilius , Ramos Kenneth S.
Published: March 2023

Abstract
Long interspersed nuclear elements [LINE-1 (L1)] are abundant retrotransposons in mammalian genomes that remain silent under most conditions. Cellular stress signals activate L1, but the molecular mechanisms controlling L1 activation remain unclear. Evidence is presented here that benzo(a)pyrene (BaP), an environmental hydrocarbon metabolized by mammalian cytochrome P450s to reactive carcinogenic intermediates, increases L1 retrotransposition in HeLa cells. Increased retrotransposition is mediated by up-regulation of L1 RNA levels, increased L1 cDNA synthesis, and stable genomic integration. Activation of L1 is dependent on the ability of BaP to cause DNA damage because it is absent in HeLa cells challenged with nongenotoxic hydrocarbon carcinogens. Thus, the mutations and genomic instability observed in human populations exposed to genotoxic environmental hydrocarbons may involve epigenetic activation of mobile elements dispersed throughout the human genome. (Cancer Res 2006; 66(5): 2616-20)
Data from Bone Morphogenetic Protein 7 Protects Prostate Cancer Cells from Stress-Induced Apoptosis via Both Smad and c-Jun NH2-Terminal Kinase Pathways
Authors: Yang Shangxin , Lim Minyoung , Pham Linda K. , Kendall Stephen E. , Reddi A. Hari , Altieri Dario C. , Roy-Burman Pradip
Published: March 2023

Abstract
We reported earlier that exposure to exogenous bone morphogenetic protein 7 (BMP7) could strongly inhibit serum starvation–induced apoptosis to C4-2B cell line, a variant of the LNCaP human prostate cancer cell line with propensity for bone metastasis. Whereas serum starvation suppressed the expression of survivin, a member of the inhibitor of apoptosis protein family, its expression was sustained in the presence of BMP7. In this study, we present evidence that BMP7 exposure up-regulated survivin promoter activity, an effect that was associated with activation of Smad, and could be repressed by dominant-negative Smad5. Additionally, serum starvation–induced suppression of c-jun NH2-terminal kinase (JNK) activity in C4-2B cells could be mostly restored by BMP7, and a JNK inhibitor could counteract the antiapoptotic effect of BMP7, without a significant effect on the level of survivin expression. Thus, we identified JNK pathway as another signaling mode for the antiapoptotic function of BMP7. To test the effect of endogenous up-regulation of BMP7, we genetically modulated the C4-2B cell line to overexpress BMP7 protein. Not only was this altered cell line resistant to serum starvation–induced apoptosis but it also exhibited patterns of Smad activation, survivin up-regulation, and JNK activation similar to those of the parental C4-2B cells exposed to exogenous BMP7. Consistent with these in vitro findings of BMP7 action, we acquired correlative results of Smad activation, survivin expression, and JNK activation in the progression of prostate cancer in the conditional Pten deletion mouse model, in which we first obtained the evidence of BMP7 overexpression. (Cancer Res 2006; 66(8): 4285-90)
Data from Epidermal Growth Factor Receptor Signaling Is Required for Microadenoma Formation in the Mouse Azoxymethane Model of Colonic Carcinogenesis
Authors: Fichera Alessandro , Little Nathaniel , Jagadeeswaran Sujatha , Dougherty Urszula , Sehdev Amikar , Mustafi Reba , Cerda Sonia , Yuan Weihua , Khare Sharad , Tretiakova Maria , Gong Can , Tallerico Michael , Cohen Greg , Joseph Loren , Hart John , Turner Jerrold R. , Bissonnette Marc
Published: March 2023

Abstract
Colonic carcinogenesis involves the progressive dysregulation of homeostatic mechanisms that control growth. The epidermal growth factor (EGF) receptor (EGFR) regulates colonocyte growth and differentiation and is overexpressed in many human colon cancers. A requirement for EGFR in colonic premalignancy, however, has not been shown. In the current study, we used a specific EGFR antagonist, gefitinib, to investigate this role of the receptor in azoxymethane colonic premalignancy. The azoxymethane model shares many clinical, histologic, and molecular features of human colon cancer. Mice received azoxymethane i.p. (5 mg/kg/wk) or saline for 6 weeks. Animals were also gavaged with gefitinib (10 mg/kg body weight) or vehicle (DMSO) thrice weekly for 18 weeks, a dose schedule that inhibited normal receptor activation by exogenous EGF. Compared with control colonocytes [bromodeoxyuridine (BrdUrd), 2.2 ± 1.2%], azoxymethane significantly increased proliferation (BrdUrd, 12.6 ± 2.8%), whereas gefitinib inhibited this hyperproliferation (BrdUrd, 6.2 ± 4.0%; <0.005). Azoxymethane significantly induced pro-transforming growth factor-α (6.4 ± 1.3–fold) and increased phospho-(active) EGFR (5.9 ± 1.1–fold), phospho-(active) ErbB2 (2.3 ± 0.2–fold), and phospho-(active) extracellular signal-regulated kinase (3.3 ± 0.4–fold) in premalignant colonocytes. Gefitinib inhibited activations of these kinases by >75% (P < 0.05). Gefitinib also significantly reduced the number of large aberrant crypt foci and decreased the incidence of colonic microadenomas from 75% to 33% (P < 0.05). Gefitinib concomitantly decreased cell cycle–regulating cyclin D1 and prostanoid biosynthetic enzyme cyclooxygenase-2 in microadenomas, suggesting that these regulators are key targets of EGFR in colonic carcinogenesis. These results show for the first time that EGFR signaling is required for early stages of colonic carcinogenesis. Our findings suggest, moreover, that inhibitors of EGFR might be useful in chemopreventive strategies in individuals at increased risk for colonic malignancies. [Cancer Res 2007;67(2):827–35]
Data from Activation of TLX3 and NKX2-5 in t(5;14)(q35;q32) T-Cell Acute Lymphoblastic Leukemia by Remote 3′-BCL11B Enhancers and Coregulation by PU.1 and HMGA1
Authors: Nagel Stefan , Scherr Michaela , Kel Alexander , Hornischer Klaus , Crawford Gregory E. , Kaufmann Maren , Meyer Corinna , Drexler Hans G. , MacLeod Roderick A.F.
Published: March 2023

Abstract
In T-cell acute lymphoblastic leukemia, alternative t(5;14)(q35;q32.2) forms effect dysregulation of either TLX3 or NKX2-5 homeobox genes at 5q35 by juxtaposition with 14q32.2 breakpoints dispersed across the BCL11B downstream genomic desert. Leukemic gene dysregulation by t(5;14) was investigated by DNA inhibitory treatments with 26-mer double-stranded DNA oligonucleotides directed against candidate enhancers at, or near, orphan T-cell DNase I hypersensitive sites located between 3′-BCL11B and VRK1. NKX2-5 down-regulation in t(5;14) PEER cells was almost entirely restricted to DNA inhibitory treatment targeting enhancers within the distal breakpoint cluster region and was dose and sequence dependent, whereas enhancers near 3′-BCL11B regulated that gene only. Chromatin immunoprecipitation assays showed that the four most effectual NKX2-5 ectopic enhancers were hyperacetylated. These enhancers clustered ∼1 Mbp downstream of BCL11B, within a region displaying multiple regulatory stigmata, including a TCRA enhancer motif, deep sequence conservation, and tight nuclear matrix attachment relaxed by trichostatin A treatment. Intriguingly, although TLX3/NKX2-5 promoter/exon 1 regions were hypoacetylated, their expression was trichostatin A sensitive, implying extrinsic regulation by factor(s) under acetylation control. Knockdown of PU.1, known to be trichostatin A responsive and which potentially binds TLX3/NKX2-5 promoters, effected down-regulation of both homeobox genes. Moreover, genomic analysis showed preferential enrichment near ectopic enhancers of binding sites for the PU.1 cofactor HMGA1, the knockdown of which also inhibited NKX2-5. We suggest that HMGA1 and PU.1 coregulate ectopic homeobox gene expression in t(5;14) T-cell acute lymphoblastic leukemia by interactions mediated at the nuclear matrix. Our data document homeobox gene dysregulation by a novel regulatory region at 3′-BCL11B responsive to histone deacetylase inhibition and highlight a novel class of potential therapeutic target amid noncoding DNA. [Cancer Res 2007;67(4):1461–71]
Data from An Organometallic Protein Kinase Inhibitor Pharmacologically Activates p53 and Induces Apoptosis in Human Melanoma Cells
Authors: Smalley Keiran S.M. , Contractor Rooha , Haass Nikolas K. , Kulp Angela N. , Atilla-Gokcumen G. Ekin , Williams Douglas S. , Bregman Howard , Flaherty Keith T. , Soengas Maria S. , Meggers Eric , Herlyn Meenhard
Published: March 2023

Abstract
Unlike other tumors, melanomas harbor wild-type (WT) p53 but exhibit impaired p53-dependent apoptosis. The mechanisms for the impaired p53 activation are poorly understood but may be linked to the high expression of the p53 suppressor Mdm2, which is found in >50% of melanoma lesions. Here, we describe an organometallic glycogen synthase kinase 3β (GSK3β) inhibitor (DW1/2) as a potent activator of p53 and inducer of cell death in otherwise highly chemoresistant melanoma cells. Using RNA interference and pharmacologic approaches, we show that p53 is required for the cytotoxic effects of this organometallic inhibitor. The DW1/2 compound was barely able to induce cell death in melanoma cells with p53 mutations, further confirming the requirement for p53-WT in the cytotoxic effects of the GSK3β inhibition. Mechanistic analysis of the p53-dependent cell death indicated an apoptotic mechanism involving depolarization of mitochondrial membrane potential, caspase cleavage, and elevated NOXA expression. The effect of p53 was not simply due to passive up-regulation of protein expression as adenoviral-mediated overexpression of p53 was not able to induce cell death. Treatment of melanoma cells with DW1/2 was instead found to decrease levels of Mdm2 and Mdm4. The importance of Mdm2 down-regulation in DW1/2-induced apoptosis was confirmed by treating the p53-WT cells with the p53/Mdm2 antagonist Nutlin-3. Taken together, our data provide a new strategy for the pharmacologic activation of p53 in melanoma, which may be a viable approach for overcoming apoptotic resistance in melanoma and offer new hope for rational melanoma therapy. [Cancer Res 2007;67(1):209–17]
Data from Direct Visualization of Macrophage-Assisted Tumor Cell Intravasation in Mammary Tumors
Authors: Wyckoff Jeffrey B. , Wang Yarong , Lin Elaine Y. , Li Jiu-feng , Goswami Sumanta , Stanley E. Richard , Segall Jeffrey E. , Pollard Jeffrey W. , Condeelis John
Published: March 2023

Abstract
Although the presence of macrophages in tumors has been correlated with poor prognosis, until now there was no direct observation of how macrophages are involved in hematogenous metastasis. In this study, we use multiphoton microscopy to show, for the first time, that tumor cell intravasation occurs in association with perivascular macrophages in mammary tumors. Furthermore, we show that perivascular macrophages of the mammary tumor are associated with tumor cell intravasation in the absence of local angiogenesis. These results show that the interaction between macrophages and tumor cells lying in close proximity defines a microenvironment that is directly involved in the intravasation of cancer cells in mammary tumors. [Cancer Res 2007;67(6):2649–56]
Data from Peroxisome Proliferator-Activated Receptor γ Is a Zac Target Gene Mediating Zac Antiproliferation
Authors: Barz Thomas , Hoffmann Anke , Panhuysen Markus , Spengler Dietmar
Published: March 2023

Abstract
Zac is a C2H2 zinc finger protein, which regulates apoptosis and cell cycle arrest through DNA binding and transactivation. During tumorigenesis and in response to mitogenic activation, Zac gene expression is down-regulated in a methylation-sensitive manner. As yet, no target genes have been identified that could explain the potent antiproliferative function of Zac. Here, applying genome-wide expression analysis, we identify peroxisome proliferator-activated receptor γ (PPARγ) as a new bona fide Zac target gene, which is induced by direct Zac binding to the proximal PPARγ1 promoter. We show that in human colon carcinoma cells, ZAC activates expression of PPARγ target genes in a PPARγ-dependent manner. Moreover, we show that treatment of pituitary tumor cells with octreotide, a somatostatin analogue, leads to Zac induction and subsequent Zac-dependent up-regulation of PPARγ, which thereupon mediates part of the antiproliferative activity of Zac. Our work provides a first step toward elucidating a functional relationship between Zac and PPARγ that could be relevant to the understanding of tumorigenesis and diabetes as well. (Cancer Res 2006; 66(24): 11975-82)
Data from Recognition of Prostate and Melanoma Tumor Cells by Six-Transmembrane Epithelial Antigen of Prostate–Specific Helper T Lymphocytes in a Human Leukocyte Antigen Class II–Restricted Manner
Authors: Kobayashi Hiroya , Nagato Toshihiro , Sato Keisuke , Aoki Naoko , Kimura Shoji , Murakami Masamoto , Iizuka Hajime , Azumi Makoto , Kakizaki Hidehiro , Tateno Masatoshi , Celis Esteban
Published: March 2023

Abstract
The six-transmembrane epithelial antigen of prostate (STEAP) protein is an attractive candidate for T cell–based immunotherapy because it is overexpressed in prostate cancer and various other tumor types. Several peptide epitopes capable of stimulating CTLs that killed STEAP-expressing tumor cells have been described. Our goal was the identification of helper T lymphocyte (HTL) epitopes of STEAP for the optimization of T cell–based immunotherapies against STEAP-expressing malignancies. Candidate HTL epitopes for STEAP were predicted using in silico algorithms for HLA class II–binding peptides and were tested for their ability to elicit HTL responses by in vitro peptide vaccination of CD4 T lymphocytes from healthy individuals and prostate cancer patients. Two peptides (STEAP102–116 and STEAP192–206) were effective in stimulating in vitro antitumor HTL responses in both normal individuals and prostate cancer patients. Notably, both STEAP HTL peptides behaved as promiscuous T-cell epitopes because they stimulated T cells in the context of more than one MHC class II allele. These newly described STEAP HTL epitopes could be of value for the design and optimization of T cell–based immunotherapy against STEAP-expressing tumors. [Cancer Res 2007;67(11):5498–504]
Data from Enhanced Peritoneal Ovarian Tumor Dissemination by Tissue Transglutaminase
Authors: Satpathy Minati , Cao Liyun , Pincheira Roxana , Emerson Robert , Bigsby Robert , Nakshatri Harikrishna , Matei Daniela
Published: March 2023

Abstract
Tissue transglutaminase (TG2) is involved in Ca2+-dependent aggregation and polymerization of proteins. We previously reported that TG2 mRNA is up-regulated in epithelial ovarian cancer (EOC) cells compared with normal ovarian epithelium. Here, we show overexpression of the TG2 protein in ovarian cancer cells and tumors and its secretion in ascites fluid and define its role in EOC. By stable knockdown and overexpression, we show that TG2 enhances EOC cell adhesion to fibronectin and directional cell migration. This phenotype is preserved in vivo, where the pattern of tumor dissemination in the peritoneal space is dependent on TG2 expression levels. TG2 knockdown diminishes dissemination of tumors on the peritoneal surface and mesentery in an i.p. ovarian xenograft model. This phenotype is associated with deficient β1 integrin-fibronectin interaction, leading to weaker anchorage of cancer cells to the peritoneal matrix. Highly expressed in ovarian tumors, TG2 facilitates i.p. tumor dissemination by enhancing cell adhesion to the extracellular matrix and modulating β1 integrin subunit expression. [Cancer Res 2007;67(15):7194–202]
Data from Induction of p53-Dependent Senescence by the MDM2 Antagonist Nutlin-3a in Mouse Cells of Fibroblast Origin
Authors: Efeyan Alejo , Ortega-Molina Ana , Velasco-Miguel Susana , Herranz Daniel , Vassilev Lyubomir T. , Serrano Manuel
Published: March 2023

Abstract
Cellular senescence is emerging as an important in vivo anticancer response elicited by multiple stresses, including currently used chemotherapeutic drugs. Nutlin-3a is a recently discovered small-molecule antagonist of the p53-destabilizing protein murine double minute-2 (MDM2) that induces cell cycle arrest and apoptosis in cancer cells with functional p53. Here, we report that nutlin-3a induces cellular senescence in murine primary fibroblasts, oncogenically transformed fibroblasts, and fibrosarcoma cell lines. No evidence of drug-induced apoptosis was observed in any case. Nutlin-induced senescence was strictly dependent on the presence of functional p53 as revealed by the fact that cells lacking p53 were completely insensitive to the drug, whereas cells lacking the tumor suppressor alternative reading frame product of the CDKN2A locus underwent irreversible cell cycle arrest. Interestingly, irreversibility was achieved in neoplastic cells faster than in their corresponding parental primary cells, suggesting that nutlin-3a and oncogenic signaling cooperate in activating p53. Our current results suggest that senescence could be a major cellular outcome of cancer therapy by antagonists of the p53-MDM2 interaction, such as nutlin-3a. [Cancer Res 2007;67(15):7350–7]
Data from A Peptide Selected by Biopanning Identifies the Integrin αvβ6 as a Prognostic Biomarker for Nonsmall Cell Lung Cancer
Authors: Elayadi Anissa N. , Samli Kausar N. , Prudkin Ludmila , Liu Ying-Horng , Bian Aihua , Xie Xian-Jin , Wistuba Ignacio I. , Roth Jack A. , McGuire Michael J. , Brown Kathlynn C.
Published: March 2023

Abstract
The development of new modes of diagnosis and targeted therapy for lung cancer is dependent on the identification of unique cell surface features on cancer cells and isolation of reagents that bind with high affinity and specificity to these biomarkers. We recently isolated a 20-mer peptide which binds to the lung adenocarcinoma cell line, H2009, from a phage-displayed peptide library. We show here that the cellular receptor for this peptide, TP H2009.1, is the uniquely expressed integrin, αvβ6, and the peptide binding to lung cancer cell lines correlates to integrin expression. The peptide is able to mediate cell-specific uptake of a fluorescent nanoparticle via this receptor. Expression of αvβ6 was assessed on 311 human lung cancer samples. The expression of this integrin is widespread in early-stage nonsmall cell lung carcinoma (NSCLC). Log-rank test and Cox regression analyses show that expression of this integrin is significantly associated with poor patient outcome. Preferential expression is observed in the tumors compared with the surrounding normal lung tissue. Our data indicate that αvβ6 is a prognostic biomarker for NSCLC and may serve as a receptor for targeted therapies. Thus, cell-specific peptides isolated from phage biopanning can be used for the discovery of cell surface biomarkers, emphasizing the utility of peptide libraries to probe the surface of a cell. [Cancer Res 2007;67(12):5889–95]
Data from Akt1 Ablation Inhibits, whereas Akt2 Ablation Accelerates, the Development of Mammary Adenocarcinomas in Mouse Mammary Tumor Virus (MMTV)-ErbB2/Neu and MMTV-Polyoma Middle T Transgenic Mice
Authors: Maroulakou Ioanna G. , Oemler William , Naber Stephen P. , Tsichlis Philip N.
Published: March 2023

Abstract
Ample evidence to date links the phosphatidylinositol 3-kinase–regulated protein kinase Akt with the induction and progression of human cancer, including breast cancer. However, there are three Akt isoforms with limited information about their specificity during oncogenesis. This study addresses the role of the three isoforms in polyoma middle T (PyMT) and ErbB2/Neu-driven mammary adenocarcinomas in mice. The effects of ablation of Akt1, Akt2, and Akt3 on the induction and the biology of these tumors were dramatically different, with ablation of Akt1 inhibiting, ablation of Akt2 accelerating, and ablation of Akt3 having a small, not statistically significant, inhibitory effect on tumor induction by both transgenes. Whereas PyMT-induced tumors are all invasive, Akt1−/−Neu–induced tumors are more invasive than Akt2−/−Neu–induced tumors. Invasiveness, however, does not always correlate with metastasis. Ablation of individual Akt isoforms does not affect the development of the mammary gland during puberty or the expression of the transgenes. Akt ablation, therefore, influences tumor induction by modulating transgene-induced oncogenic signaling. Immunostaining for Ki-67 and cyclin D1 and terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling assays on tissue sections revealed that the delay of tumor induction in Akt1 knockout mice is due to the inhibitory effects of Akt1 ablation on cell proliferation and survival. Given that these animal models exhibit significant similarities to human breast cancer, the results of the present study may have significant translational implications because they may influence how Akt inhibitors will be used in the treatment of human cancer. [Cancer Res 2007;67(1):167–77]
Data from Platelet Factor-4 Variant Chemokine CXCL4L1 Inhibits Melanoma and Lung Carcinoma Growth and Metastasis by Preventing Angiogenesis
Authors: Struyf Sofie , Burdick Marie D. , Peeters Elke , Van den Broeck Karolien , Dillen Chris , Proost Paul , Van Damme Jo , Strieter Robert M.
Published: March 2023

Abstract
The platelet factor-4 variant, designated PF-4var/CXCL4L1, is a recently described natural non-allelic gene variant of the CXC chemokine platelet factor-4/CXCL4. PF-4var/CXCL4L1 was cloned, and the purified recombinant protein strongly inhibited angiogenesis. Recombinant PF-4var/CXCL4L1 was angiostatically more active (at nanomolar concentration) than PF-4/CXCL4 in various test systems, including wound-healing and migration assays for microvascular endothelial cells and the rat cornea micropocket assay for angiogenesis. Furthermore, PF-4var/CXCL4L1 more efficiently inhibited tumor growth in animal models of melanoma and lung carcinoma than PF-4/CXCL4 at an equimolar concentration. For B16 melanoma in nude mice, a significant reduction in tumor size and the number of small i.t. blood vessels was obtained with i.t. applied PF-4var/CXCL4L1. For A549 adenocarcinoma in severe combined immunodeficient mice, i.t. PF-4var/CXCL4L1 reduced tumor growth and microvasculature more efficiently than PF-4/CXCL4 and prevented metastasis to various organs better than the angiostatic IFN-inducible protein 10/CXCL10. Finally, in the syngeneic model of Lewis lung carcinoma, PF-4var/CXCL4L1 inhibited tumor growth equally well as monokine induced by IFN-γ (Mig)/CXCL9, also known to attract effector T lymphocytes. Taken together, PF-4var/CXCL4L1 is a highly potent antitumoral chemokine preventing development and metastasis of various tumors by inhibition of angiogenesis. These data confirm the clinical potential of locally released chemokines in cancer therapy. [Cancer Res 2007;67(12):5940–8]
Data from Cellular Transformation and Activation of the Phosphoinositide-3-Kinase–Akt Cascade by the ETV6-NTRK3 Chimeric Tyrosine Kinase Requires c-Src
Authors: Jin Wook , Yun Chohee , Hobbie Amy , Martin Matthew J. , Sorensen Poul H.B. , Kim Seong-Jin
Published: March 2023

Abstract
The ETV6-NTRK3 (EN) chimeric tyrosine kinase, a potent oncoprotein expressed in tumors derived from multiple cell lineages, functions as a constitutively active protein-tyrosine kinase. ETV6-NTRK expression leads to the constitutive activation of two major effector pathways of wild-type NTRK3, namely, the Ras–mitogen-activated protein kinase (MAPK) mitogenic pathway and the phosphoinositide-3-kinase (PI3K)-Akt pathway mediating cell survival, and both are required for EN transformation. However, it remains unclear how ETV6-NTRK3 activates Ras-Erk1/2 and/or PI3K-Akt cascades. Here, we define some aspects of the molecular mechanisms regulating ETV6-NTRK–dependent Ras-Erk1/2 and PI3K-Akt activation. We show that ETV6-NTRK3 associates with c-Src, and that treatment with SU6656, a c-Src inhibitor, completely blocks ETV6-NTRK-transforming activity. Treatment of NIH3T3 cells expressing ETV6-NTRK3 with SU6656 attenuated the activation of Ras-Erk1/2 and PI3K-Akt. Suppression of c-Src by RNA interference in NIH3T3-ETV6-NTRK3 cells resulted in markedly decreased expression of cyclin D1 and suppression of activation of Ras-Erk1/2 and PI3K-Akt. However, in Src-deficient cells, the ETV6-NTRK3 failed to activate the PI3K-Atk pathway, but not the Ras-Erk1/2 pathway. Therefore, these data indicate that ETV6-NTRK3 induces the PI3K-Akt cascade through the activation of c-Src. [Cancer Res 2007;67(7):3192–200]
Data from Ribosomal S6 Kinase 2 Is a Key Regulator in Tumor Promoter–Induced Cell Transformation
Authors: Cho Yong-Yeon , Yao Ke , Kim Hong-Gyum , Kang Bong Seok , Zheng Duo , Bode Ann M. , Dong Zigang
Published: March 2023

Abstract
The ribosomal S6 kinase 2 (RSK2), a member of the p90RSK (RSK) family of proteins, is a widely expressed serine/threonine kinase that is activated by extracellular signal-regulated kinase 1/2 and phosphoinositide-dependent kinase 1 in response to many growth factors and peptide hormones. Its activation signaling enhances cell survival. However, the roles of RSK2 in cell transformation have not yet been elucidated. Here, we found that RSK2 is a critical serine/threonine kinase for the regulation of cell transformation. When cells were stimulated with tumor promoters, such as epidermal growth factor (EGF) or 12-O-tetradecanoylphorbol-13-acetate (TPA), phosphorylation of RSK was increased within 5 min. Cell proliferation was suppressed in RSK2−/− mouse embryonic fibroblasts (MEFs) compared with RSK2+/+ MEFs. Moreover, RSK2−/− MEFs accumulated at the G1 phase of the cell cycle under normal cell culture conditions as well as after stimulation with EGF or TPA. In the anchorage-independent cell transformation assay (soft agar), stable expression of RSK2 in JB6 cells significantly enhanced colony formation in either the presence or absence of tumor promoters. Furthermore, knockdown of RSK2 with small interfering RNA-RSK2 suppressed constitutively active Ras (RasG12V)-induced foci formation in NIH3T3 cells. In addition, kaempferol, an inhibitor of RSK2, suppressed EGF-induced colony formation of JB6 Cl41 cells in soft agar, which was associated with inhibition of histone H3 phosphorylation (Ser10). These results showed that RSK2 is a key regulator for cell transformation induced by tumor promoters such as EGF and TPA. [Cancer Res 2007;67(17):8104–12]
Data from High Frequencies of Functional Tumor-Reactive T Cells in Bone Marrow and Blood of Pancreatic Cancer Patients
Authors: Schmitz-Winnenthal Friedrich H. , Volk Christine , Z'graggen Kaspar , Galindo Luis , Nummer Daniel , Ziouta Yvonne , Bucur Marianna , Weitz Jürgen , Schirrmacher Volker , Büchler Markus W. , Beckhove Philipp
Published: March 2023

Abstract
Pancreatic cancer is characterized by aggressive growth and treatment resistance. New approaches include immunotherapeutic strategies but the type and extent of spontaneous immune responses against tumor antigens remains unclear. A dominance of TH2 cytokines in patients' sera reported previously suggests systemic tumor-induced immunosuppression, potentially inhibiting the induction of tumor-reactive T cells. We characterized the localization, frequencies, and functional potential of spontaneously induced memory T cells specific for individual tumor antigens or the tumor-associated antigen mucin-1 in the peripheral blood and bone marrow of 41 pancreatic cancer patients. We found high numbers of tumor-reactive T cells in all bone marrow samples and in 50% of the blood samples. These cells secreted the TH1 cytokine IFN-γ rather than TH2 cytokines upon stimulation with tumor antigens. Although consistently induced during pancreatic cancer, T cells specific for pancreatic antigens were not detected during chronic pancreatitis, suggesting that their evaluation may be of diagnostic use in both diseases. Freshly isolated T cells from cancer patients recognized autologous tumor cells and rejected them in vitro and in a xenotransplant model in vivo, suggesting their therapeutic potential. Thus, tumor antigen–specific T cell responses occur regularly during pancreatic cancer disease and lead to enrichment of tumor cell–reactive memory T cells in the bone marrow. The bone marrow can therefore be considered an important organ for antitumor immune responses in pancreatic cancer.
Data from Spontaneous Mammary Tumors Differ Widely in Their Inherent Sensitivity to Adoptively Transferred T Cells
Authors: Wall Erika M. , Milne Katy , Martin Michele L. , Watson Peter H. , Theiss Patty , Nelson Brad H.
Published: March 2023

Abstract
Immunotherapy of cancer can lead to the selection of antigen loss variants, which provides strong rationale to target oncogenes that are essential for tumor growth or viability. To investigate this concept, we tagged the HER2/neu oncogene with epitopes from ovalbumin to confer recognition by T-cell receptor transgenic CD8+ (OT-I) and CD4+ (OT-II) T cells. Transgenic mice expressing neuOT-I/OT-II developed mammary adenocarcinomas at 6 to 10 months of age. Adoptively transferred naive OT-I cells (with or without OT-II cells) proliferated vigorously on encountering neuOT-I/OT-II-expressing tumors. This was followed by the complete regression of 37% of tumors, whereas others showed partial/stable responses (40%) or progressive disease (23%). Those tumors undergoing complete regression never recurred. In mice with multiple primary tumors, simultaneous regressions and nonregressions were often seen, indicating that immune evasion occurred at a local rather than systemic level. The majority of nonregressing tumors expressed NeuOT-I/OT-II and MHC class I, and many avoided rejection through a profound block to T-cell infiltration. Thus, T cells directed against an essential oncogene can permanently eradicate a subset of spontaneous, established mammary tumors. However, in other tumors, local barriers severely limit the therapeutic response. To maximize the efficacy of immunotherapy against spontaneous cancers, predictive strategies that take into account the heterogeneity of the tumor microenvironment will be required. [Cancer Res 2007;67(13):6442–50]
Data from Nm23-H1 Suppresses Tumor Cell Motility by Down-regulating the Lysophosphatidic Acid Receptor EDG2
Authors: Horak Christine E. , Lee Jong Heun , Elkahloun Abdel G. , Boissan Mathieu , Dumont Sylvie , Maga Tara K. , Arnaud-Dabernat Sandrine , Palmieri Diane , Stetler-Stevenson William G. , Lacombe Marie-Lise , Meltzer Paul S. , Steeg Patricia S.
Published: March 2023

Abstract
Exogenous overexpression of the metastasis suppressor gene Nm23-H1 reduces the metastatic potential of multiple types of cancer cells and suppresses in vitro tumor cell motility and invasion. Mutational analysis of Nm23-H1 revealed that substitution mutants P96S and S120G did not inhibit motility and invasion. To elucidate the molecular mechanism of Nm23-H1 motility suppression, expression microarray analysis of an MDA-MB-435 cancer cell line overexpressing wild-type Nm23-H1 was done and cross-compared with expression profiles from lines expressing the P96S and S120G mutants. Nine genes, MET, PTN, SMO, FZD1, L1CAM, MMP2, NETO2, CTGF, and EDG2, were down-regulated by wild-type but not by mutant Nm23-H1 expression. Reduced expression of these genes coincident with elevated Nm23-H1 expression was observed in human breast tumor cohorts, a panel of breast carcinoma cell lines, and hepatocellular carcinomas from control versus Nm23-M1 knockout mice. The functional significance of the down-regulated genes was assessed by transfection and in vitro motility assays. Only EDG2 overexpression significantly restored motility to Nm23-H1–suppressed cancer cells, enhancing motility by 60-fold in these cells. In addition, silencing EDG2 expression with small interfering RNA reduced the motile phenotype of metastatic breast cancer cells. These data suggest that Nm23-H1 suppresses metastasis, at least in part, through down-regulation of EDG2 expression. [Cancer Res 2007;67(15):7238–46]
Data from Prostate Cancer Cells with Stem Cell Characteristics Reconstitute the Original Human Tumor In vivo
Authors: Gu Guangyu , Yuan Jialing , Wills Marcia , Kasper Susan
Published: March 2023

Abstract
Cancer may arise from a cancer stem/progenitor cell that shares characteristics with its normal counterpart. We report the reconstitution of the original human prostate cancer specimen from epithelial cell lines (termed HPET for human prostate epithelial/hTERT) derived from this sample. These tumors can be described in terms of Gleason score, a classification not applied to any of the transgenic mouse models currently developed to mimic human disease. Immunohistochemical and Western blot analyses indicate that they do not express androgen receptor or p63, similar to that reported for prostate stem cells. These cell lines also express embryonic stem markers (Oct4, Nanog, and Sox2) as well as early progenitor cell markers (CD44 and Nestin) in vitro. Clonally derived HPET cells reconstitute the original human tumor in vivo and differentiate into the three prostate epithelial cell lineages, indicating that they arise from a common stem/progenitor cell. Serial transplantation experiments reconstitute the tumors, suggesting that a fraction of parental or clonally derived HPET cells have self-renewal potential. Thus, this model may enhance our understanding of human tumor development and provide a mechanism for studying cancer stem/progenitor cells in differentiation, tumorigenesis, preclinical testing, and the development of drug resistance. [Cancer Res 2007;67(10):4807–15]
Data from Overexpression of the Low Molecular Weight Cyclin E in Transgenic Mice Induces Metastatic Mammary Carcinomas through the Disruption of the ARF-p53 Pathway
Authors: Akli Said , Van Pelt Carolyn S. , Bui Tuyen , Multani Asha S. , Chang Sandy , Johnson David , Tucker Susan , Keyomarsi Khandan
Published: March 2023

Abstract
In tumor cells, cyclin E deregulation results in the appearance of five low molecular weight (LMW) isoforms. When overexpressed in breast cancer cells, these forms of cyclin E induce genomic instability, resistance to inhibition by p21 and p27, and resistance to antiestrogen therapy. Additionally, the LMW forms of cyclin E strongly correlate with decreased survival in patients with breast cancer. However, the oncologic role of the LMW forms of cyclin E in breast cancer tumorigenesis is yet to be determined. To this end, we generated transgenic mice expressing full-length cyclin E alone (M46A), full-length and the EL4 isoforms (EL1/EL4), or the EL2/3 isoforms of cyclin E (T1) under the control of the mouse mammary tumor virus promoter. Compared with full-length cyclin E, LMW cyclin E overexpression induces delayed mammary growth during the pubertal phase and abnormal cell morphology during lactation. Both primary mammary tumor formation and metastasis were markedly enhanced in LMW cyclin E transgenic mice. LMW cyclin E overexpression in mammary epithelial cells of mice is sufficient by itself to induce mammary adenocarcinomas in 34 of 124 (27%) animals compared with 7 of 67 (10.4%) mice expressing only the full-length cyclin E (P < 0.05). In addition, metastasis was seen in 25% of LMW cyclin E tumor–bearing animals compared with only 8.3% of tumors in the full-length cyclin E background (P < 0.05). Moreover, LMW cyclin E overexpression selects for inactivation of p53 by loss of heterozygosity and spontaneous and frequent inactivation of ARF. Therefore, LMW cyclin E overexpression strongly selects for spontaneous inactivation of the ARF-p53 pathway in vivo, canceling its protective checkpoint function and accelerating progression to malignancy. [Cancer Res 2007;67(15):7212–22]
Data from DR5 Receptor Mediates Anoikis in Human Colorectal Carcinoma Cell Lines
Authors: Laguinge Luciana M. , Samara Raed N. , Wang Wenge , El-Deiry Wafik S. , Corner Georgia , Augenlicht Leonard , Mishra Lopa , Jessup J. Milburn
Published: March 2023

Abstract
As human colorectal cancer (CRC) cells metastasize to distant sites, they are susceptible to detachment-induced cell death or anoikis — a form of apoptosis that occurs when anchorage-dependent CRC cells go into suspension. Our goal was to identify whether tumor necrosis factor receptor apoptosis-inducing ligand (TRAIL) receptors mediate anoikis in human CRC cells. First, we assessed whether caspases of the extrinsic (caspase-8) or intrinsic (caspase-9) death pathways were involved. Caspase-8 was cleaved during exposure to suspension culture in four CRC lines, and cell death was inhibited by caspase-3 and caspase-8 inhibitors but not by a caspase-9 inhibitor. Gene transcripts in macrophage inflammatory protein-101 (MIP-110), a weakly metastatic human CRC, were increased at least 2-fold for TRAIL-R2 (DR5) and TRAIL after 24 h of suspension culture compared with cells in monolayer culture. The increased expression of DR5 was confirmed at the protein level at 24 h, and exposure of MIP-101 cells to an antagonistic antibody to DR5 decreased caspase-8 activation. The antagonistic antibody to DR5 inhibited anoikis in four human CRC lines. Treatment with an antagonistic DR4 antibody or a neutralizing antibody to TRAIL ligand did not reduce anoikis consistently. Knockdown of DR5 or TRAIL also inhibited anoikis, whereas exogenous TRAIL or FasL did not consistently increase anoikis. In summary, DR5 receptor mediates death signals for anoikis in human CRC cells through the extrinsic apoptotic pathway. [Cancer Res 2008;68(3):909–17]
Data from Inhibition of Heat Shock Protein 27–Mediated Resistance to DNA Damaging Agents by a Novel PKCδ-V5 Heptapeptide
Authors: Kim Eun-Ho , Lee Hae-June , Lee Dae-Hoon , Bae Sangwoo , Soh Jae-Won , Jeoung Dooil , Kim Joon , Cho Chul-Koo , Lee Yoon-Jin , Lee Yun-Sil
Published: March 2023

Abstract
Heat shock protein 27 (HSP27), which is highly expressed in human lung and breast cancer tissues, induced resistance to cell death against various stimuli. Treatment of NCI-H1299 cells, which express a high level of HSP27, with small interference RNA specifically targeting HSP27 resulted in inhibition of their resistance to radiation or cisplatin, suggesting that HSP27 contributed to cellular resistance in these lung cancer cells. Furthermore, because HSP27 interacts directly with the COOH terminus of the protein kinase Cδ (PKCδ)-V5 region with ensuing inhibition of PKCδ activity and PKCδ-mediated cell death, we wished to determine amino acid residues in the V5 region that mediate its interaction with HSP27. Investigation with various deletion mutants of the region revealed that amino acid residues 668 to 674 of the V5 region mediate its interaction with HSP27. When NCI-H1299 cells were treated with biotin or with FITC-tagged heptapeptide of the residues 668 to 674 (E-F-Q-F-L-D-I), the cells exhibited dramatically increased cisplatin or radiation-induced cell death with the heptapeptide having efficient interaction with HSP27, which in turn restored the PKCδ activity that had been inhibited by HSP27. In vivo nude mice grafting data also suggested that NCI-H1299 cells were sensitized by this heptapeptide. The above data strongly show that the heptapeptide of the PKCδ-V5 region sensitized human cancer cells through its interaction with HSP27, thereby sequestering HSP27. The heptapeptide may provide a novel strategy for selective neutralization of HSP27. [Cancer Res 2007;67(13):6333–9]
Data from Seizure 6-Like (SEZ6L) Gene and Risk for Lung Cancer
Authors: Gorlov Ivan P. , Meyer Peter , Liloglou Triantafillos , Myles Jonathan , Boettger Melanie Barbara , Cassidy Adrian , Girard Luc , Minna John D. , Fischer Reiner , Duffy Stephen , Spitz Margaret R. , Haeussinger Karl , Kammerer Stefan , Cantor Charles , Dierkesmann Rainer , Field John K. , Amos Christopher I.
Published: March 2023

Abstract
DNA pooling in combination with high-throughput sequencing was done as a part of the Sequenom-Genefinder project. In the pilot study, we tested 83,715 single nucleotide polymorphisms (SNP), located primarily in gene-based regions, to identify polymorphic susceptibility variants for lung cancer. For this pilot study, 369 male cases and 287 controls of both sexes (white Europeans of Southern German origin) were analyzed. The study identified a candidate region in 22q12.2 that contained numerous SNPs showing significant case-control differences and that coincides with a region that was shown previously to be frequently deleted in lung cancer cell lines. The candidate region overlies the seizure 6-like (SEZ6L) gene. The pilot study identified a polymorphic Met430Ile substitution in the SEZ6L gene (SNP rs663048) as the top candidate for a variant modulating risk of lung cancer. Two replication studies were conducted to assess the association of SNP rs663048 with lung cancer risk. The M. D. Anderson Cancer Center study included 289 cases and 291 controls matched for gender, age, and smoking status. The Liverpool Lung Project (a United Kingdom study) included 248 cases and 233 controls. Both replication studies showed an association of the rs663048 with lung cancer risk. The homozygotes for the variant allele had more than a 3-fold risk compared with the wild-type homozygotes [combined odds ratio (OR), 3.32; 95% confidence interval (95% CI), 1.81–7.21]. Heterozygotes also had a significantly elevated risk of lung cancer from the combined replication studies with an OR of 1.15 (95% CI, 1.04–1.59). The effect remained significant after adjusting for age, gender, and pack-years of tobacco smoke. We also compared expression of SEZ6L in normal human bronchial epithelial cells (n = 7), non–small cell lung cancer (NSCLC; n = 52), and small cell lung cancer (SCLC; n = 22) cell lines by using Affymetrix HG-U133A and HG-U133B GeneChips. We found that the average expression level of SEZ6L in NSCLC cell lines was almost two times higher and in SCLC cell lines more than six times higher when compared with normal lung epithelial cell lines. Using the National Center for Biotechnology Information Gene Expression Omnibus database, we found a ∼2-fold elevated and statistically significant (P = 0.004) level of SEZ6L expression in tumor samples compared with normal lung tissues. In conclusion, the results of these studies representing 906 cases compared with 811 controls indicate a role of the SEZ6L Met430Ile polymorphic variant in increasing lung cancer risk. [Cancer Res 2007;67(17):8406–11]
Data from Statin-Induced Breast Cancer Cell Death: Role of Inducible Nitric Oxide and Arginase-Dependent Pathways
Authors: Kotamraju Srigiridhar , Willams Carol L. , Kalyanaraman Balaraman
Published: March 2023

Abstract
Statins are widely used cholesterol-lowering drugs that selectively inhibit the enzyme 3-hydroxy-3-methylglutaryl CoA reductase, leading to decreased cholesterol biosynthesis. Emerging data indicate that statins stimulate apoptotic cell death in several types of proliferating tumor cells, including breast cancer cells, which is independent of its cholesterol-lowering property. The objective here was to elucidate the molecular mechanism(s) by which statins induce breast cancer cell death. Fluvastatin and simvastatin (5–10 μmol/L) treatment enhanced the caspase-3–like activity and DNA fragmentation in MCF-7 cells, and significantly inhibited the proliferation of MCF-7 cells but not MCF-10 cells (noncancerous epithelial cells). Statin-induced cytotoxic effects were reversed by mevalonate, an immediate metabolic product of the acetyl CoA/3-hydroxy-3-methylglutaryl CoA reductase reaction. Both simvastatin and fluvastatin enhanced nitric oxide (·NO) levels which were inhibited by mevalonate. Statin-induced ·NO and tumor cell cytotoxicity were inhibited by 1400W, a more specific inhibitor of inducible nitric oxide synthase (iNOS or NOS II). Both fluvastatin and simvastatin increased iNOS mRNA and protein expression. Stimulation of iNOS by statins via inhibition of geranylgeranylation by GGTI-298, but not via inhibition of farnesylation by FTI-277, enhanced the proapoptotic effects of statins in MCF-7 cells. Statin-mediated antiproliferative and proapoptotic effects were exacerbated by sepiapterin, a precursor of tetrahydrobiopterin, an essential cofactor of ·NO biosynthesis by NOS. We conclude that iNOS-mediated ·NO is responsible in part for the proapoptotic, tumoricidal, and antiproliferative effects of statins in MCF-7 cells. [Cancer Res 2007;67(15):7386–94]