Defining the pathway to insulin-like growth factor system targeting in cancer

The insulin-like growth factors (IGEs; IGF-1 and IGF-2) play central roles in cell growth, differentiation, survival, transformation and metastasis. The biologic effects of the IGFs are mediated by the IGF-1 receptor (IGF-1R), a receptor tyrosine kinase with homology to the insulin receptor (IR). Dysregulation of the ICE system is well recognized as a key contributor to the progression of multiple cancers, with IGF-1R activation increasing the tumorigenic potential of breast, prostate, lung, colon and head and neck squamous cell carcinoma (HNSCC). Despite this relationship, targeting the IGF-1R has only recently undergone development as a molecular cancer therapeutic. As it has taken hold, we are witnessing a robust increase and interest in targeting the inhibition of IGF-1R signaling. This is accentuated by the list of over 30 drugs, including monoclonal antibodies (mAbs) and tyrosine kinase inhibitors (TKIs) that are under evaluation as single agents or in combination therapies 1]. The ICE-binding proteins (IGFBPs) represent the third component of the ICE system consisting of a class of six soluble secretory proteins. They represent a unique class of naturally occurring ICE-antagonists that bind to and sequester IGF-1 and IGF-2, inhibiting their access to the IGF-1R. Due to their dual targeting of the IGFs without affecting insulin action, the IGFBPs are an untapped ``third'' class of IGF-1R inhibitors. in this commentary, we highlight some of the significant aspects of and prospects for targeting the IGF-1R and describe what the future may hold. (C) 2010 Elsevier Inc. All rights reserved.

DNA targeting polyaza macrobicyclic dizinc(II) complexes promoting high in vitro caspase dependent anti-proliferative activity against human carcinoma cancer cells

A series of macrobicyclic dizinc(II) complexes Zn2L1-2B](ClO4)(4) (1-6) have been synthesized and characterized (L1-2 are polyaza macrobicyclic binucleating ligands, and B is the N,N-donor heterocyclic base (viz. 2,2'-bipyridine (bipy) and 1,10-phenanthroline (phen)). The DNA and protein binding, DNA hydrolysis and anticancer activity of these complexes were investigated. The interactions of complexes 1-6 with calf thymus DNA were studied by spectroscopic techniques, including absorption, fluorescence and CD spectroscopy. The DNA binding constant values of the complexes were found to range from 2.80 x 10(5) to 5.25 x 10(5) M-1, and the binding affinities are in the following order: 3 > 6 > 2 > 5 > 1 > 4. All the dizinc(II) complexes 1-6 are found to effectively promote the hydrolytic cleavage of plasmid pBR322 DNA under anaerobic and aerobic conditions. Kinetic data for DNA hydrolysis promoted by 3 and 6 under physiological conditions give observed rate constants (k(obs)) of 5.56 +/- 0.1 and 5.12 +/- 0.2 h(-1), respectively, showing a 10(7)-fold rate acceleration over the uncatalyzed reaction of dsDNA. Remarkably, the macrobicyclic dizinc(II) complexes 1-6 bind and cleave bovine serum albumin (BSA), and effectively promote the caspase-3 and caspase-9 dependent deaths of HeLa and BeWo cancer cells. The cytotoxicity of the complexes was further confirmed by lactate dehydrogenase enzyme levels in cancer cell lysate and content media.

Oncogenic MicroRNA-155 Down-regulates Tumor Suppressor CDC73 and Promotes Oral Squamous Cell Carcinoma Cell Proliferation IMPLICATIONS FOR CANCER THERAPEUTICS

The CDC73 gene is mutationally inactivated in hereditary and sporadic parathyroid tumors. It negatively regulates beta-catenin, cyclin D1, and c-MYC. Down-regulation of CDC73 has been reported in breast, renal, and gastric carcinomas. However, the reports regarding the role of CDC73 in oral squamous cell carcinoma (OSCC) are lacking. In this study we show that CDC73 is down-regulated in a majority of OSCC samples. We further show that oncogenic microRNA-155 (miR-155) negatively regulates CDC73 expression. Our experiments show that the dramatic up-regulation of miR-155 is an exclusive mechanism for down-regulation of CDC73 in a panel of human cell lines and a subset of OSCC patient samples in the absence of loss of heterozygosity, mutations, and promoter methylation. Ectopic expression of miR-155 in HEK293 cells dramatically reduced CDC73 levels, enhanced cell viability, and decreased apoptosis. Conversely, the delivery of a miR-155 antagonist (antagomir-155) to KB cells overexpressing miR-155 resulted in increased CDC73 levels, decreased cell viability, increased apoptosis, and marked regression of xenografts in nude mice. Cotransfection of miR-155 with CDC73 in HEK293 cells abrogated its pro-oncogenic effect. Reduced cell proliferation and increased apoptosis of KB cells were dependent on the presence or absence of the 3'-UTR in CDC73. In summary, knockdown of CDC73 expression due to overexpression of miR-155 not only adds a novelty to the list of mechanisms responsible for its down-regulation in different tumors, but the restoration of CDC73 levels by the use of antagomir-155 may also have an important role in therapeutic intervention of cancers, including OSCC.

Primary Microcephaly Gene MCPH1 Shows Signatures of Tumor Suppressors and Is Regulated by miR-27a in Oral Squamous Cell Carcinoma

Mutations in the MCPH1 (microcephalin 1) gene, located at chromosome 8p23.1, result in two autosomal recessive disorders: primary microcephaly and premature chromosome condensation syndrome. MCPH1 has also been shown to be downregulated in breast, prostate and ovarian cancers, and mutated in 1/10 breast and 5/41 endometrial tumors, suggesting that it could also function as a tumor suppressor (TS) gene. To test the possibility of MCPH1 as a TS gene, we first performed LOH study in a panel of 81 matched normal oral tissues and oral squamous cell carcinoma (OSCC) samples, and observed that 14/71 (19.72%) informative samples showed LOH, a hallmark of TS genes. Three protein truncating mutations were identified in 1/15 OSCC samples and 2/5 cancer cell lines. MCPH1 was downregulated at both the transcript and protein levels in 21/41 (51.22%) and 19/25 (76%) OSCC samples respectively. A low level of MCPH1 promoter methylation was also observed in 4/40 (10%) tumor samples. We further observed that overexpression of MCPH1 decreased cellular proliferation, anchorage-independent growth in soft agar, cell invasion and tumor size in nude mice, indicating its tumor suppressive function. Using bioinformatic approaches and luciferase assay, we showed that the 3'-UTR of MCPH1 harbors two non-overlapping functional seed regions for miR-27a which negatively regulated its level. The expression level of miR-27a negatively correlated with the MCPH1 protein level in OSCC. Our study indicates for the first time that, in addition to its role in brain development, MCPH1 also functions as a tumor suppressor gene and is regulated by miR-27a.

Antitumor and antioxidant status of Terminalia catappa against Ehrlich ascites carcinoma in Swiss albino mice

Objective: The present study was undertaken to evaluate the antitumor and antioxidant status of ethanol extract of Terminalia catappa leaves against Ehrlich ascites carcinoma (EAC) in Swiss albino mice. Materials and Methods: The leaves powder was extracted with Soxhlet apparatus and subjected to hot continuous percolation using ethanol (95% v/v). Tumor bearing animals was treated with 50 and 200 mg/kg of ethanol extract. EAC induced in mice by intraperitoneal injection of EAC cells 1 x 10(6) cells/mice. The study was assed using life span of EAC-bearing hosts, hematological parameters, volume of solid tumor mass and status of antioxidant enzymes such as lipid peroxidation (LPO), reduced glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT) activities. Total phenolics and flavonoids contents from the leaves extract were also determined. Results: Total phenolics and flavonoids contents from the leaves extract were found 354.02 and 51.67 mg/g extract. Oral administration of ethanol extract of T. catappa (50 and 200 mg/kg) increased the life span (27.82% and 60.59%), increased peritoneal cell count (8.85 +/- 0.20 and 10.37 +/- 0.26) and significantly decreased solid tumor mass (1.16 +/- 0.14 cm(2)) at 200 mg/kg as compared with EAC-tumor bearing mice (P < 0.01). Hematological profile including red blood cell count, white blood cell count, hemoglobin (11.91 +/- 0.47 % g) and protein estimation were found to be nearly normal levels in extract-treated mice compared with tumor bearing control mice. Treatment with T. catappa significantly decreased levels of LPO and GSH, and increased levels of SOD and CAT activity (P < 0.01). Conclusion: T. catappa exhibited antitumor effect by modulating LPO and augmenting antioxidant defense systems in EAC bearing mice. The phenolic and flavonoid components in this extract may be responsible for antitumor activity.

MicroRNA-125a Reduces Proliferation and Invasion of Oral Squamous Cell Carcinoma Cells by Targeting Estrogen-related Receptor alpha IMPLICATIONS FOR CANCER THERAPEUTICS

Estrogen-related receptor (ESRRA) functions as a transcription factor and regulates the expression of several genes, such as WNT11 and OPN. Up-regulation of ESRRA has been reported in several cancers. However, the mechanism underlying its up-regulation is unclear. Furthermore, the reports regarding the role and regulation of ESRRA in oral squamous cell carcinoma (OSCC) are completely lacking. Here, we show that tumor suppressor miR-125a directly binds to the 3UTR of ESRRA and represses its expression. Overexpression of miR-125a in OSCC cells drastically reduced the level of ESRRA, decreased cell proliferation, and increased apoptosis. Conversely, the delivery of an miR-125a inhibitor to these cells drastically increased the level of ESRRA, increased cell proliferation, and decreased apoptosis. miR-125a-mediated down-regulation of ESRRA impaired anchorage-independent colony formation and invasion of OSCC cells. Reduced cell proliferation and increased apoptosis of OSCC cells were dependent on the presence of the 3UTR in ESRRA. The delivery of an miR-125a mimic to OSCC cells resulted in marked regression of xenografts in nude mice, whereas the delivery of an miR-125a inhibitor to OSCC cells resulted in a significant increase of xenografts and abrogated the tumor suppressor function of miR-125a. We observed an inverse correlation between the expression levels of miR-125a and ESRRA in OSCC samples. In summary, up-regulation of ESRRA due to down-regulation of miR-125a is not only a novel mechanism for its up-regulation in OSCC, but decreasing the level of ESRRA by using a synthetic miR-125a mimic may have an important role in therapeutic intervention of OSCC and other cancers.

Genomic amplification upregulates estrogen-related receptor alpha and its depletion inhibits oral squamous cell carcinoma tumors in vivo

The ESRRA gene encodes a transcription factor and regulates several genes, such as WNT11 and OPN, involved in tumorigenesis. It is upregulated in several cancers, including OSCC. We have previously shown that the tumor suppressor miR-125a targets ESRRA, and its downregulation causes upregulation of ESRRA in OSCC. Upregulation of ESRRA in the absence of downregulation of miR-125a in a subset of OSCC samples suggests the involvement of an alternative mechanism. Using TaqMan (R) copy number assay, here we report for the first time that the genomic amplification of ESRRA causes its upregulation in a subset of OSCC samples. Ectopic overexpression of ESRRA led to accelerated cell proliferation, anchorage-independent cell growth and invasion, and inhibited apoptosis. Whereas, knockdown of ESRRA expression by siRNA led to reduced cell proliferation, anchorage-independent cell growth and invasion, and accelerated apoptosis. Furthermore, the delivery of a synthetic biostable ESRRA siRNA to OSCC cells resulted in regression of xenografts in nude mice. Thus, the genomic amplification of ESRRA is another novel mechanism for its upregulation in OSCC. Based on our in vitro and in vivo experiments, we suggest that targeting ESRRA by siRNA could be a novel therapeutic strategy for OSCC and other cancers.

Inhibition of p300 lysine acetyltransferase activity by luteolin reduces tumor growth in head and neck squamous cell carcinoma (HNSCC) xenograft mouse model

Chromatin acetylation is attributed with distinct functional relevance with respect to gene expression in normal and diseased conditions thereby leading to a topical interest in the concept of epigenetic modulators and therapy. We report here the identification and characterization of the acetylation inhibitory potential of an important dietary flavonoid, luteolin. Luteolin was found to inhibit p300 acetyltransferase with competitive binding to the acetyl CoA binding site. Luteolin treatment in a xenografted tumor model of head and neck squamous cell carcinoma (HNSCC), led to a dramatic reduction in tumor growth within 4 weeks corresponding to a decrease in histone acetylation. Cells treated with luteolin exhibit cell cycle arrest and decreased cell migration. Luteolin treatment led to an alteration in gene expression and miRNA profile including up-regulation of p53 induced miR-195/215, let7C; potentially translating into a tumor suppressor function. It also led to down regulation of oncomiRNAs such as miR-135a, thereby reflecting global changes in the microRNA network. Furthermore, a direct correlation between the inhibition of histone acetylation and gene expression was established using chromatin immunoprecipitation on promoters of differentially expressed genes. A network of dysregulated genes and miRNAs was mapped along with the gene ontology categories, and the effects of luteolin were observed to be potentially at multiple levels: at the level of gene expression, miRNA expression and miRNA processing.

Prevalencia del carcinoma escamoso ocular y sus precursores en un hato lechero nicaraguense, relacionado con raza, edad y pigmentación

En el presente trabajo se estudió la prevalencia del Carcinoma escamoso ocular bovino y la relación que tiene esta enfermedad con la pigmentación ocular, la raza y la edad en la Empresa Genética “Roberto Alvarado” (Chiltepe). Para esto se estudiaron 4108 animales de los grupos raciales Holstein Friesian (Canadiense), Holstein Friesian (Criollo), Holstein x Pardo, 75% Holstein y Pardo Suizo. Se determinó que la prevalencia total del cáncer en la población analizada es de 2.6%. Además que existe presencia de lesión únicamente cuando existe pigmentación. Se concluyó que hay claras diferencias entre los grupos raciales estudiados siendo el más afectado el Holstein Friesian(Canadiense), y que hay una tendencia a incrementarse la presencia del cáncer a medida la edad.

Vascular endothelial growth factor regulates melanoma cell adhesion and growth in the bone marrow microenvironment via tumor cyclooxygenase-2

Background: Human melanoma frequently colonizes bone marrow (BM) since its earliest stage of systemic dissemination, prior to clinical metastasis occurrence. However, how melanoma cell adhesion and proliferation mechanisms are regulated within bone marrow stromal cell (BMSC) microenvironment remain unclear. Consistent with the prometastatic role of inflammatory and angiogenic factors, several studies have reported elevated levels of cyclooxygenase-2 (COX-2) in melanoma although its pathogenic role in bone marrow melanoma metastasis is unknown. Methods: Herein we analyzed the effect of cyclooxygenase-2 (COX-2) inhibitor celecoxib in a model of generalized BM dissemination of left cardiac ventricle-injected B16 melanoma (B16M) cells into healthy and bacterial endotoxin lipopolysaccharide (LPS)-pretreated mice to induce inflammation. In addition, B16M and human A375 melanoma (A375M) cells were exposed to conditioned media from basal and LPS-treated primary cultured murine and human BMSCs, and the contribution of COX-2 to the adhesion and proliferation of melanoma cells was also studied. Results: Mice given one single intravenous injection of LPS 6 hour prior to cancer cells significantly increased B16M metastasis in BM compared to untreated mice; however, administration of oral celecoxib reduced BM metastasis incidence and volume in healthy mice, and almost completely abrogated LPS-dependent melanoma metastases. In vitro, untreated and LPS-treated murine and human BMSC-conditioned medium (CM) increased VCAM-1-dependent BMSC adherence and proliferation of B16M and A375M cells, respectively, as compared to basal medium-treated melanoma cells. Addition of celecoxib to both B16M and A375M cells abolished adhesion and proliferation increments induced by BMSC-CM. TNF alpha and VEGF secretion increased in the supernatant of LPS-treated BMSCs; however, anti-VEGF neutralizing antibodies added to B16M and A375M cells prior to LPS-treated BMSC-CM resulted in a complete abrogation of both adhesion-and proliferation-stimulating effect of BMSC on melanoma cells. Conversely, recombinant VEGF increased adherence to BMSC and proliferation of both B16M and A375M cells, compared to basal medium-treated cells, while addition of celecoxib neutralized VEGF effects on melanoma. Recombinant TNFa induced B16M production of VEGF via COX-2-dependent mechanism. Moreover, exogenous PGE2 also increased B16M cell adhesion to immobilized recombinant VCAM-1. Conclusions: We demonstrate the contribution of VEGF-induced tumor COX-2 to the regulation of adhesion-and proliferation-stimulating effects of TNFa, from endotoxin-activated bone marrow stromal cells, on VLA-4-expressing

Three-dimensional growth as multicellular spheroid activates the proangiogenic phenotype of colorectal carcinoma cells via LFA-1-dependent VEGF: implications on hepatic micrometastasis

Background: The recruitment of vascular stromal and endothelial cells is an early event occurring during cancer cell growth at premetastatic niches, but how the microenvironment created by the initial three-dimensional (3D) growth of cancer cells affects their angiogenesis-stimulating potential is unclear. Methods: The proangiogenic profile of CT26 murine colorectal carcinoma cells was studied in seven-day cultured 3D-spheroids of <300 mu m in diameter, produced by the hanging-drop method to mimic the microenvironment of avascular micrometastases prior to hypoxia occurrence. Results: Spheroid-derived CT26 cells increased vascular endothelial growth factor (VEGF) secretion by 70%, which in turn increased the in vitro migration of primary cultured hepatic sinusoidal endothelium (HSE) cells by 2-fold. More importantly, spheroid-derived CT26 cells increased lymphocyte function associated antigen (LFA)-1-expressing cell fraction by 3-fold; and soluble intercellular adhesion molecule (ICAM)-1, given to spheroid-cultured CT26 cells, further increased VEGF secretion by 90%, via cyclooxygenase (COX)-2-dependent mechanism. Consistent with these findings, CT26 cancer cells significantly increased LFA-1 expression in non-hypoxic avascular micrometastases at their earliest inception within hepatic lobules in vivo; and angiogenesis also markedly increased in both subcutaneous tumors and hepatic metastases produced by spheroid-derived CT26 cells. Conclusion: 3D-growth per se enriched the proangiogenic phenotype of cancer cells growing as multicellular spheroids or as subclinical hepatic micrometastases. The contribution of integrin LFA-1 to VEGF secretion via COX-2 was a micro environmental-related mechanism leading to the pro-angiogenic activation of soluble ICAM-1-activated colorectal carcinoma cells. This mechanism may represent a new target for specific therapeutic strategies designed to block colorectal cancer cell growth at a subclinical micrometastatic stage within the liver.

Ejercicio físico y prevención del cáncer de colon

Proyecto desarrollado en castellano. Trata sobre los efectos de la actividad física en la prevención del cáncer de colon y cómo su práctica puede reducir el riesgo de padecerlo.

Interacciones entre las células madre pluripotentes del carcinoma embrionario y las células mioides peritubulares del testículo

112 p.+ anexos

O papel da proteína supressora de tumor p53 na indução de ciclooxigenase-2 em Carcinoma de Pulmão de Células Não Pequenas: aspectos moleculares e clínicos

O Carcinoma de Pulmão de Células Não Pequenas (NSCLC) é uma doença freqüentemente letal e altamente resistente à terapia oncológica convencional, como por exemplo, o tratamento quimioterápico com cisplatina e paclitaxel. A superexpressão de Ciclooxigenase-2 (COX-2) é constantemente observada em pacientes com NSCLC, estando associada ao prognóstico ruim destes pacientes. Acredita-se que a alta expressão de COX-2 produz efeitos anti-apoptóticos, porém pouco é conhecido sobre os mecanismos de regulação desta enzima. Muitos sinais capazes de ativar COX-2 também induzem a proteína supressora de tumor p53, conhecida pelo seu papel fundamental no controle da proliferação celular e apoptose. Dados recentes indicam que a proteína p53 é um importante regulador da expressão de COX-2. O objetivo desta dissertação foi avaliar os efeitos da quimioterapia na expressão da enzima COX-2 em linhagens celulares com diferente status do gene TP53, e ainda, correlacionar a expressão de COX-2 e o status mutacional de TP53, com as características clínico-patológicas de pacientes com NSCLC. Como ferramentas experimentais foram usadas técnicas de biologia celular e molecular como interferência de RNA, PCR em tempo real, análise mutacional e imuno-histoquímica. Com os resultados obtidos, observamos que as linhagens celulares de câncer de pulmão que apresentam p53 na sua forma selvagem, quando expostas ao tratamento com cisplatina, apresentaram indução da expressão de COX-2 (RNAm e proteína), em adição ao aumento da síntese de Prostaglandina E2 (PGE2). Em contrapartida, a expressão de COX-2 não foi alterada após o tratamento com cisplatina nas linhagens celulares que apresentavam mutação no gene TP53. Ao avaliar o tratamento com paclitaxel, foi observado um aumento da expressão de COX-2 nas linhagens A549 e H460 (linhagens celulares do tipo selvagem para p53), entretanto não foi observada alteração nos níveis de PGE2. Em adição, o tratamento com paclitaxel induziu um aumento da expressão de COX-2 na linhagem com deleção em TP53, ACC LC-319. Em seguida, após silenciamento de p53 na linhagem celular A549, por interferência de RNA, a cisplatina passou a não ser mais capaz de induzir o aumento da expressão de COX-2. No tratamento com paclitaxel, o silenciamento de TP53 não mudou a expressão de COX-2, indicando assim um efeito independente de p53. Dessa maneira, sugerimos que a indução de COX-2, por cisplatina, em linhagens celulares NSCLC é dependente de p53. Na análise dos pacientes NSCLC, os resultados demonstram que 54% dos pacientes apresentam expressão positiva de COX-2. Mutações em TP53 foram observadas em 57% dos pacientes, incluindo 56% de fumantes correntes e 37% de ex-fumantes. Uma associação entre a expressão de COX-2 e o status selvagem de TP53 foi observada, entre os pacientes que apresentaram expressão positiva de COX-2, 80% apresentaram TP53 selvagem. Um número maior de pacientes é necessário para aumentar o poder estatístico e confirmar as tendências observadas nesse estudo