Involvement of hedgehog pathway in early onset, aggressive molecular subtypes and metastatic potential of breast cancer

Background Dysregulation of hedgehog pathway is observed in numerous cancers. Relevance of hedgehog pathway genes in cancer cohort and inhibition of its downstream effector (GLI1) towards metastasis in cell lines are explored in the study. Method One hundred fifty fresh tumours of breast cancer patients were collected for the study. Based on differential expression, panel of 6 key regulators of the pathway (SHH, DHH, IHH, PTCH1, SMO and GLI1) in microarray datasets were identified. Expressional profiles of aforementioned genes were later correlated with clinico-pathological parameters in Pakistani breast cancer cohort at transcript and protein levels. In addition, GLI1 over expressing breast cancer cell lines (MDA-MB-231 and MCF-7) were treated with GANT61 to explore its probable effects on metastasis. Result SHH, DHH, PTCH1 and GLI1 were significantly over-expressed in tumours as compared with respective normal mammary tissues. A significant correlation of SHH, DHH and GLI1 expression with advanced tumour size, stages, grades, nodal involvement and distant metastasis was observed (p < 0.05). Over-expression of SHH, DHH and GLI1 was significantly related with patients having early onset and pre-menopausal status. Of note, hedgehog pathway was frequently up regulated in luminal B and triple negative breast cancer affected women. In addition, positive correlations were observed among aforementioned members of pathway and Ki67 (r-value: 0.63–0.78) emphasizing their role towards disease progression. Exposure of GANT61 (inhibitor for GLI1) significantly restricted cell proliferation, reduced cell motility and invasion. Conclusion Role of activated hedgehog pathway in breast cancer metastasis provides a novel target for cancer therapy against aggressive cancer subtypes

Role of Plexin B1 in a breast cancer cohort of Pakistani patients and its contribution towards cancer metastasis as indicated by an in vitro model

Background/Aim: In the current study, the role of plexin B1 in breast cancer metastasis was explored. Materials and Methods: Freshly-excised tumours along with background tissues of affected patients (n=121) were collected from Pakistani hospitals and processed for RNA isolation and cDNA synthesis. Using quantitative polymerase chain reaction, expression of plexin B1 was evaluated and correlated with clinicopathological parameters. Furthermore, involvement of plexin B1 in metastasis was explored by generating gene knockdown in MDA-MB-231 and MCF-7 breast cancer cells. Results: Poorly-differentiated tumours showed low plexin B1 expression in comparison to well-differentiated ones. Similarly, reduced plexin B1 expression correlated positively with advanced tumour stage and metastasis. Loss of plexin B1 significantly reduced cell adhesion in comparison with respective control cell lines (p<0.05). Knockdown of plexin B1 in MDA-MB-231 cells led to a remarkable increase in cell motility in contrast to the respective control. Conclusion: Loss of plexin B1 expression might play a pivotal role in enhancing the metastatic potential of breast cancer cell

The plexin-B family and its role in cancer progression

Plexins are transmembrane protein receptors for semaphorin molecules. These molecules are involved in numerous cellular activities related to cell proliferation, adhesion along with the basement membrane, cellular motility and invasive capability. All nine members of Plexins identified in vertebrates have been grouped into subclasses, termed Plexin-A, Plexin-B, Plexin-C and Plexin-D. Plexin-B consists of three members, namely Plexin-B1, Plexin-B2 and Plexin-B3. Plexin-B1 functionally interacts with Sema4A (Yukawa et al., 2010) and can also form heterodimer with Plexin-B2 for Sema4A binding (Nkyimbeng-Takwi and Chapoval, 2011). Plexin-B2 binds with Sema4C. Plexin-B3 mediates interaction with both Sema4G and Sema5A. Some semaphorines exist in a membrane-bound form only, whereas other family members can be found in tissues/fluids in both secreted and membrane-bound forms. This ligand-receptor interaction between sema4D and Plexin-B1 indicated in different signaling pathways results in many intriguing and interesting findings, highlighting its importance in both physiology and pathology. Apart from bidirectional signaling among these molecules, the involvement of Plexin-B1 in the processes described here directly involves a bidirectional singaling between Sema4Dand Plexin-B1. Being a high affinity receptor for both Sema4A and Sema4D, the role played by Plexin-B1 in cancer progression, metastasis and angiogenesis is still an area requiring further research. Activation of Sema4D mediated downstream effectors is largely influenced by cross talk of Plexin-B1 with other molecules, such as Her-2 and Met. In this review, all findings regarding Plexin-B1 upstream and downstream regulation and its putative involvement in relation to the ultimate fate of cancer cells are discussed

SUPRA-MVC: A novel algorithmic approach to handle large scale bioinformatics applications

Abstract: The challenge to manage massive applications arises from weak application design strategy. The objective role of this article is to propose an algorithmic approach to engineer high performance bioinformatics softwares. The technique proposed in this article aids in building efficient and effective large applications which become difficult to manage otherwise. This framework methodology is designed to implement application development using codeigniter 2.1.0 framework

Capillary morphogenesis gene 2 inhibits growth of breast cancer cells and is inversely correlated with the disease progression and prognosis

Background Capillary morphogenesis gene 2 (CMG2) also known as anthrax toxin receptor 2 was identified as a gene being up-regulated in capillary morphogenesis. It has been shown to be involved in cell adhesion and motility which are critical functions for cancerous cells to disseminate. The present study aimed to examine the expression of CMG2 in breast cancer and its implication in the disease progression. Materials and methods Breast primary tumours and background tissues were collected immediately after surgery and stored at −80 °C with approval by the local ethics committee, and written consent obtained from patients. The expression of CMG2 in 127 breast cancer tumour samples and 34 normal mammary tissues was determined using real-time PCR. Knockdown and over-expression in breast cancer cells were established using constructed plasmid vectors carrying either anti-CMG2 ribozyme or full-coding sequence of human CMG2. The effect on growth of breast cancer cells was examined using in vitro and in vivo models. Results The CMG2 transcript levels were reduced in advanced tumours compared with its expression in tumours of early stage according to their TNM staging. The reduced expression was associated with shorter overall survival, p = 0.004 compared with patients who had higher expression. The knockdown of CMG2 resulted in an increased in vitro growth of MDA-MB-231 cells which express this gene at relatively higher levels. This is consistent with the finding from MCF-7 cells which express lower levels of CMG2 and exhibited reduced growth following over-expression of CMG2. The over-expression of CMG2 also demonstrated an inhibitory effect on in vivo growth of MCF-7 cells. Conclusion Reduced expression of CMG2 is associated with disease progression and poor prognosis of breast cancer. CMG2 has an inhibitory effect on growth of breast cancer cells. Further investigation is required to shed light on the prognostic and therapeutic potential of targeting this molecule

Genomic Relevance of FGFR2 on the Prognosis of HCV-Induced Hepatocellular Carcinoma Patients

The Fibroblast Growth Factor Receptors (FGFRs) are known to regulate cancer metabolism in different tumor types, including hepatocellular carcinoma (HCC). Several risk factors are associated with HCC, of which viral infections (Hepatitis B and C) and cirrhosis are prominent. In Pakistan as well as in highly developed countries like the United States, hepatitis C virus HCV infections are most commonly reported in HCC. Here, we aimed to investigate the clinical relevance of FGFR receptors in HCC and their role in HCV-positive HCC cases. 264 HCC samples along with their clinical information and 96 normal liver samples were collected. qPCR was done to estimate the expression of FGFR1, FGFR2, FGFR3 and FGFR4. Three independent HCV-induced HCC cohorts (containing 293 HCC samples) were used for validation. According to in vitro results, FGFR1 was upregulated in HCV+ HCC patients. However, in all three independent cohorts of HCC, significant a down-regulation of FGFR1 was observed. FGFR2 overexpression was observed in the in vitro cohort as well as in three independent HCC cohorts. Interestingly, a strong correlation of FGFR2 expression was observed between cirrhosis and HCV in all four HCC cohorts. Our study suggested that FGFR2 expression can be used to classify HCC patients based on HCV infection. This FGFR2-based classification may lead to new therapeutic strategies against HCV-positive HCC subtypes

Effect of MHC Linked 7-Gene Signature on Delayed Hepatocellular Carcinoma Recurrence

Dysregulated immune response significantly affects hepatocellular carcinoma’s (HCC) prognosis. Human Leukocyte Antigens are key in devising immune responses against HCC. Here, we investigated how HLAs modulate HCC development at the transcriptomic level. RNA-seq data of 576 patients from two independent cohorts was retrieved. The clinicopathological relevance of all HLA genes was investigated using Fisher-Exact, correlation, and Kaplan–Meier and cox regression survival tests. Clustering of ~800 immune-related genes against HLAs was completed using a ward-agglomerative method. Networks were generated using 40 HLA associated unique genes and hub genes were investigated. HLAs including HLA-DMA, HLA-DMB, HLA-DOA and HLA-DRB6 were associated with delayed recurrence in both discovery (204 HCC cases) and validation (372 HCC cases) cohorts. Clustering analyses revealed 40 genes associated with these four HLAs in both cohorts. A set of seven genes (NCF4, TYROBP, LCP2, ZAP70, PTPRC, FYN and WAS) was found co-expressed at gene–gene interaction level in both cohorts. Furthermore, survival analysis revealed seven HLA-linked genes as predictors of delayed recurrence. Multivariate analysis also predicted that mean expression of 7-gene is an independent predictor of delayed recurrence in both cohorts. We conclude that the expression of 7-gene signature may lead to improved patient prognosis. Further studies are required for consideration in clinical practice

Promoter Methylation Status Modulate the Expression of Tumor Suppressor (RbL2/p130) Gene in Breast Cancer

<div><p>Background</p><p>Aberrant expression of tumor suppressor genes may correspond to the abnormal cell development and tumorigenesis. Rbl2/p130, a member of retinoblastoma family of proteins, has growth suppressive properties. Numerous studies reported de-regulation of Rbl2/p130 in various types of cancer as a consequence of a number of genetic alterations. However, role of epigenetic mechanisms like DNA methylation in Rbl2/p130 expression remains elusive.</p><p>Methods</p><p>In the current study, 76 breast cancer tumors along with normal tissues (n = 76), blood (n = 76) of respective individuals and control blood (n = 50) were analyzed. Rbl2/p130 expression was analyzed by quantitative real time PCR (syber green method). Promoter methylation status was studied through methylation specific PCR of bisulfite converted genomic DNA. Data was analyzed using various statistical tests.</p><p>Results</p><p>We report significantly reduced Rbl2/p130 expression (P = 0.001) in tumors tissues as compared to control samples. Similarly, Rbl2/p130 expression varies with age and disease stages (P = 0.022), which suggest its involvement in tumor progression. Aberrant promoter methylation (Δmeth) was found in almost all the diseased samples and that was significantly different (P<0.001) with control samples. Similarly, methylation status varies significantly with tumor progression stages (P = 0.022). Hyper-methylation was observed at -1, +3, +15 and +75 of Rbl2/p130 promoter flanking around the TSS. Statistical analysis revealed that Rbl2/p130 expression negatively correlates to its promoter methylation (r = -0.412) in tumor tissues. Our results reflect an epigenetic regulation of Rbl2/p130 expression in breast cancer. This highlights the importance of Rbl2/p130 promoter methylation in breast cancer pathogenesis.</p></div

Statistical relation of Rbl2/p130 expression and its promoter methylation.

<p>(<b>A)</b> Differences in Rbl2/p130 promoter methylation status of tumor and normal tissues in study cohort I (<45 years) and study cohort II (≥45 years); (<b>B)</b> Observed differences in Rbl2/p130 promoter methylation status of various tumor tissues at different disease stages; (<b>C & D)</b> Correlation of relative expression (ΔCT values) and promoter methylation status as observed. Unmeth (C panel) and Meth (D panel) values are plotted separately. Adjusted R-values are shown in respective panels.</p