Regulation of protumorigenic pathways by Insulin like growth factor binding protein2 and its association along with beta-catenin in breast cancer lymph node metastasis

Background: Insulin like growth factor binding proteins modulate the mitogenic and pro survival effects of IGF. Elevated expression of IGFBP2 is associated with progression of tumors that include prostate, ovarian, glioma among others. Though implicated in the progression of breast cancer, the molecular mechanisms involved in IGFBP2 actions are not well defined. This study investigates the molecular targets and biological pathways targeted by IGFBP2 in breast cancer. Methods: Transcriptome analysis of breast tumor cells (BT474) with stable knockdown of IGFBP2 and breast tumors having differential expression of IGFBP2 by immunohistochemistry was performed using microarray. Differential gene expression was established using R-Bioconductor package. For validation, gene expression was determined by qPCR. Inhibitors of IGF1R and integrin pathway were utilized to study the mechanism of regulation of beta-catenin. Immunohistochemical and immunocytochemical staining was performed on breast tumors and experimental cells, respectively for beta-catenin and IGFBP2 expression. Results: Knockdown of IGFBP2 resulted in differential expression of 2067 up regulated and 2002 down regulated genes in breast cancer cells. Down regulated genes principally belong to cell cycle, DNA replication, repair, p53 signaling, oxidative phosphorylation, Wnt signaling. Whole genome expression analysis of breast tumors with or without IGFBP2 expression indicated changes in genes belonging to Focal adhesion, Map kinase and Wnt signaling pathways. Interestingly, IGFBP2 knockdown clones showed reduced expression of beta-catenin compared to control cells which was restored upon IGFBP2 re-expression. The regulation of beta-catenin by IGFBP2 was found to be IGF1R and integrin pathway dependent. Furthermore, IGFBP2 and beta-catenin are co-ordinately overexpressed in breast tumors and correlate with lymph node metastasis. Conclusion: This study highlights regulation of beta-catenin by IGFBP2 in breast cancer cells and most importantly, combined expression of IGFBP2 and beta-catenin is associated with lymph node metastasis of breast tumors.

Enhanced Metastatic Potential in a 3D Tissue Scaffold toward a Comprehensive in Vitro Model for Breast Cancer Metastasis

Metastasis is clinically the most challenging and lethal aspect of breast cancer. While animal-based xenograft models are expensive and time-consuming, conventional two-dimensional (2D) cell culture systems fail to mimic in vivo signaling. In this study we have developed a three-dimensional (3D) scaffold system that better mimics the topography and mechanical properties of the breast tumor, thus recreating the tumor microenvironment in vitro to study breast cancer metastasis. Porous poly(e-caprolactone) (PCL) scaffolds of modulus 7.0 +/- 0.5 kPa, comparable to that of breast tumor tissue were fabricated, on which MDA-MB-231 cells proliferated forming tumoroids. A comparative gene expression analysis revealed that cells growing in the scaffolds expressed increased levels of genes implicated in the three major events of metastasis, viz., initiation, progression, and the site-specific colonization compared to cells grown in conventional 2D tissue culture polystyrene (TCPS) dishes. The cells cultured in scaffolds showed increased invasiveness and sphere efficiency in vitro and increased lung metastasis in vivo. A global gene expression analysis revealed a significant increase in the expression of genes involved in cell cell and cell matrix interactions and tissue remodeling, cancer inflammation, and the PI3K/Akt, Wnt, NF-kappaB, and HIFI signaling pathways all of which are implicated in metastasis. Thus, culturing breast cancer cells in 3D scaffolds that mimic the in vivo tumor-like microenvironment enhances their metastatic potential. This system could serve as a comprehensive in vitro model to investigate the manifold mechanisms of breast cancer metastasis.

CancerLectinDB: a database of lectins relevant to cancer

The role of lectins in mediating cancer metastasis, apoptosis as well as various other signaling events has been well established in the past few years. Data on various aspects of the role of lectins in cancer is being accumulated at a rapid pace. The data on lectins available in the literature is so diverse, that it becomes difficult and time-consuming, if not impossible to comprehend the advances in various areas and obtain the maximum benefit. Not only do the lectins vary significantly in their individual functional roles, but they are also diverse in their sequences, structures, binding site architectures, quaternary structures, carbohydrate affinities and specificities as well as their potential applications. An organization of these seemingly independent data into a common framework is essential in order to achieve effective use of all the data towards understanding the roles of different lectins in different aspects of cancer and any resulting applications. An integrated knowledge base (CancerLectinDB) together with appropriate analytical tools has therefore been developed for lectins relevant for any aspect of cancer, by collating and integrating diverse data. This database is unique in terms of providing sequence, structural, and functional annotations for lectins from all known sources in cancer and is expected to be a useful addition to the number of glycan related resources now available to the community. The database has been implemented using MySQL on a Linux platform and web-enabled using Perl-CGI and Java tools. Data for individual lectins pertain to taxonomic, biochemical, domain architecture, molecular sequence and structural details as well as carbohydrate specificities. Extensive links have also been provided for relevant bioinformatics resources and analytical tools. Availability of diverse data integrated into a common framework is expected to be of high value for various studies on lectin cancer biology.

ErbB-2 expression and its association with other biological parameters of breast cancer among Indian women

Overexpression of the epidermal growth factor receptor family genes, which include ErbB-1, 2, 3 and 4, has been implicated in a number of cancers. We have studied the extent of ErbB-2 overexpression among Indian women with sporadic breast cancer. Methods: Immmunohistochemistry and genomic polymerase chain reaction (PCR) were used to study the ErbB2 overexpression. ErbB2 status was correlated with other clinico-pathological parameters, including patient survival. Results: ErbB-2 overexpression was detected in 43.2% (159/368) of the cases by immunohistochemistry. For a sub-set of patients (n = 55) for whom total DNA was available, ErbB-2 gene amplification was detected in 25.5% (14/55) of the cases by genomic PCR. While the ErbB2 overexpression was significantly higher in patients with lymphnode (χ2 = 12.06, P≤ 0.001), larger tumor size (χ2 = 8.22, P = 0.042) and ductal carcinoma (χ2 = 15.42, P ≤ 0.001), it was lower in patients with disease-free survival (χ2 = 22.13, P ≤ 0.001). Survival analysis on a sub-set of patients for whom survival data were available (n = 179) revealed that ErbB-2 status (χ2 =25.94, P ≤ 0.001), lymphnode status (χ2 = 12.68, P ≤ 0.001), distant metastasis (χ2 = 19.49, P ≤ 0.001) and stage of the disease (χ2 = 28.04, P ≤0.001) were markers of poor prognosis. Conclusions: ErbB-2 overexpression was significantly greater compared with the Western literature, but comparable to other Indian studies. Significant correlation was found between ErbB-2 status and lymphnode status, tumor size and ductal carcinoma. ErbB-2 status, lymph node status, distant metastasis and stage of the disease were found to be prognostic indicators.

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.

Recapitulating tumour microenvironment in chitosan-gelatin three-dimensional scaffolds: an improved in vitro tumour model

Owing to the reduced co-relationship between conventional flat Petri dish culture (two-dimensional) and the tumour microenvironment, there has been a shift towards three-dimensional culture systems that show an improved analogy to the same. In this work, an extracellular matrix (ECM)-mimicking three-dimensional scaffold based on chitosan and gelatin was fabricated and explored for its potential as a tumour model for lung cancer. It was demonstrated that the chitosan-gelatin (CG) scaffolds supported the formation of tumoroids that were similar to tumours grown in vivo for factors involved in tumour-cell-ECM interaction, invasion and metastasis, and response to anti-cancer drugs. On the other hand, the two-dimensional Petri dish surfaces did not demonstrate gene-expression profiles similar to tumours grown in vivo. Further, the three-dimensional CG scaffolds supported the formation of tumoroids, using other types of cancer cells such as breast, cervix and bone, indicating a possible wider potential for in vitro tumoroid generation. Overall, the results demonstrated that CG scaffolds can be an improved in vitro tool to study cancer progression and drug screening for solid tumours.

Inflammation and cancer stem cells

Cancer stem cells are becoming recognised as being responsible for metastasis and treatment resistance. The complex cellular and molecular network that regulates cancer stem cells and the role that inflammation plays in cancer progression are slowly being elucidated. Cytokines, secreted by tumour associated immune cells, activate the necessary pathways required by cancer stem cells to facilitate cancer stem cells progressing through the epithelial-mesenchymal transition and migrating to distant sites. Once in situ, these cancer stem cells can secrete their own attractants, thus providing an environment whereby these cells can continue to propagate the tumour in a secondary niche. (C) 2013 Elsevier Ireland Ltd. All rights reserved.

Rrp1B gene polymorphism (1307T > C) in metastatic progression of breast cancer

Rrp1B (ribosomal RNA processing1 homolog B) is a novel candidate metastasis modifier gene in breast cancer. Functional gene assays demonstrated that a physical and functional interaction existing between Rrp1b and metastasis modifier gene SIPA1 causes reduction in the tumor growth and metastatic potential. Ectopic expression of Rrp1B modulates various metastasis predictive extra cellular matrix (ECM) genes associated with tumor suppression. The aim of this study is to determine the functional significance of single nucleotide polymorphism (SNP) in human Rrp1B gene (1307 T > C; rs9306160) with breast cancer development and progression. The study consists of 493 breast cancer cases recruited from Nizam's Institute of Medical Sciences, Hyderabad, and 558 age-matched healthy female controls from rural and urban areas. Genomic DNA was isolated by non-enzymatic method. Genotyping was done by amplification refractory mutation system (ARMS-PCR) method. Genotypes were reconfirmed by sequencing and results were analyzed statistically. We have performed Insilco analysis to know the RNA secondary structure by using online tool m fold. The TT genotype and T allele frequencies of Rrp1B1307 T > C polymorphism were significantly elevated in breast cancer (chi (2); p = < 0.008) cases compared to controls under different genetic models. The presence of T allele had conferred 1.75-fold risk for breast cancer development (OR = 1.75; 95 % CI = 1.15-2.67). The frequency of TT genotype of Rrp1b 1307T > C polymorphism was significantly elevated in obese patients (chi (2); p = 0.008) and patients with advanced disease (chi (2); p = 0.01) and with increased tumor size (chi (2); p = 0.01). Moreover, elevated frequency of T allele was also associated with positive lymph node status (chi (2); p = 0.04) and Her2 negative receptor status (chi (2); p = 0.006). Presence of Rrp1b1307TT genotype and T allele confer strong risk for breast cancer development and progression.