Association of mutation and low expression of the CTCF gene with breast cancer progression.

BACKGROUND: CTCF encodes 11-zinc finger protein which is implicated in multiple tumors including the carcinoma of the breast. The Present study investigates the association of CTCF mutations and their expression in breast cancer cases. METHODS: A total of 155 breast cancer and an equal number of adjacent normal tissue samples from 155 breast cancer patients were examined for CTCF mutation(s) by PCR-SSCP and automated DNA sequencing. Immunohistochemistry (IHC) method was used to analyze CTCF expression. Molecular findings were statistically analyzed with various clinicopathological features to identify associations of clinical relevance. RESULTS: Of the total, 16.1% (25/155) cases exhibited mutation in the CTCF gene. Missense mutations Gln > His (G > T) in exon 1 and silent mutations Ser > Ser (C > T) in exon 4 of CTCF gene were analyzed. A significant association was observed between CTCF mutations and some clinicopathological parameters namely menopausal status (p = 0.02) tumor stage (p = 0.03) nodal status (p = 0.03) and ER expression (p = 0.04). Protein expression analysis showed 42.58% samples having low or no expression (+), 38.0% with moderate (++) expression and 19.35% having high (+++) expression for CTCF. A significant association was found between CTCF protein expression and clinicopathological parameters include histological grade (p = 0.04), tumor stage (p = 0.04), nodal status (p = 0.03) and ER status (p = 0.04). CONCLUSIONS: The data suggest that CTCF mutations leading to its inactivation significantly contribute to the progression of breast cancer.

Analysis of predicted proteasomal cleavages in the methyltransferase domain from JEV.

The methyltransferase (MTase, a 265 amino acid residues long region at the N-terminal end of the viral nonfunctional supermolecule NS5 domain) is key for viral replication in Japanese Encephalitis Virus (JEV). Sequence to structure to functional information with adequate knowledge on MTase from JEV is currently limited. Therefore, it is of interest to document a report on the comprehensive analysis of predicted proteasomal cleavage data in the methyltransferase domain from JEV. This data is relevant in the design and development of vaccine and other therapeutic candidates for further consideration.

Evaluation of anticancer potential of Bacopa monnieri L. against MCF-7 and MDA-MB 231 cell line.

BACKGROUND: The ethanolic extract of Bacopa monnieri contains bacoside A and B, brahmin, cucurbitacins, and betulinic acid. Currently, cucurbitacins have also been reported for their strong anti-tumorigenic and anti-proliferative activity by inducing cell cycle arrest at the G2/M phase and formation of multiplied cells. The present study was carried out to evaluate the in vitro cytotoxic activity of ethanolic extract of dichloromethane (DCM) fraction of B. monnieri on two different cell lines. MATERIALS AND METHODS: The ethanolic extract of B. monnieri was prepared using soxhlet extraction method and different fractions (hexane, DCM, methanol, acetone, and water) of ethanolic extracts were prepared. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay of ethanolic extract and of all fractions was carried out on MCF-7 and MDA-MB 231 cell lines. The presence of cucurbitacins and betulinic acid in these fractions was confirmed by high-performance thin layer chromatography. RESULTS: The IC50 values of ethanolic extract of B. monnieri in MCF-7 and MDA-MB 231 cell lines were 72.0 µg/mL and 75.0 µg/mL, respectively. The DCM fraction of B. monnieri showed maximum cytotoxic activity among all fraction upto 72 h and was found to be 57.0 µg/mL and 42.0 µg/mL, respectively. CONCLUSION: The results showed good cytotoxic activity in DCM fraction in both the cell lines may be due to the presence of cucurbitacins and betulinic acid in DCM fraction.

A study on genetic variants of Fibroblast growth factor receptor 2 (FGFR2) and the risk of breast cancer from North India.

Genome-Wide Association Studies (GWAS) have identified Fibroblast growth factor receptor 2 (FGFR2) as a candidate gene for breast cancer with single nucleotide polymorphisms (SNPs) located in intron 2 region as the susceptibility loci strongly associated with the risk. However, replicate studies have often failed to extrapolate the association to diverse ethnic regions. This hints towards the existing heterogeneity among different populations, arising due to differential linkage disequilibrium (LD) structures and frequencies of SNPs within the associated regions of the genome. It is therefore important to revisit the previously linked candidates in varied population groups to unravel the extent of heterogeneity. In an attempt to investigate the role of FGFR2 polymorphisms in susceptibility to the risk of breast cancer among North Indian women, we genotyped rs2981582, rs1219648, rs2981578 and rs7895676 polymorphisms in 368 breast cancer patients and 484 healthy controls by Polymerase chain reaction-Restriction fragment length polymorphism (PCR-RFLP) assay. We observed a statistically significant association with breast cancer risk for all the four genetic variants (P<0.05). In per-allele model for rs2981582, rs1219648, rs7895676 and in dominant model for rs2981578, association remained significant after bonferroni correction (P<0.0125). On performing stratified analysis, significant correlations with various clinicopathological as well as environmental and lifestyle characteristics were observed. It was evident that rs1219648 and rs2981578 interacted with exogenous hormone use and advanced clinical stage III (after Bonferroni correction, P<0.000694), respectively. Furthermore, combined analysis on these four loci revealed that compared to women with 0-1 risk loci, those with 2-4 risk loci had increased risk (OR = 1.645, 95%CI = 1.152-2.347, P = 0.006). In haplotype analysis, for rs2981578, rs2981582 and rs1219648, risk haplotype (GTG) was associated with a significantly increased risk compared to the common (ACA) haplotype (OR = 1.365, 95% CI = 1.086-1.717, P = 0.008). Our results suggest that intron 2 SNPs of FGFR2 may contribute to genetic susceptibility of breast cancer in North India population.

Association of mutation and hypermethylation of p21 gene with susceptibility to breast cancer: a study from north India.

p21 gene located at chromosome 6p21.2 is a possible tumour suppressor gene involved in the pathogenesis of breast cancer. Both genetic and epigenetic alterations in p21 have been implicated in breast carcinoma. In the present study, our main aim was to study the impact of these two kinds of alterations of p21 gene in Indian female breast cancer patients. A total of 150 female breast cancer patients of north India were screened by PCR-SSCP followed by direct sequencing and methylation specific PCR. Mutational screening of p21 gene revealed significant amount of mutations [32.66% (49/150)] in exon 2, whereas p21 promoter was found hypermethylated in 42 of 150 (28%) breast cancer patients in our population. The intriguing feature of the study was the G>T transition (GAG>TAG) at codon 107 and the A>C transition (AGC>CGC) at codon 146 possibly rendering p21 completely ineffective in its anti- proliferative activity. Our results suggest a significant association between the mutational and hypermethylation profile of p21 gene. Therefore, we show for the first time that the significant association of p21 mutation and hypermethylation leads to the complete inactivation of p21 gene in Indian female breast cancer patients. Complete silencing of the p21 gene seems to be the result not only of genetic alterations but also of epigenetic modification.

Genetic polymorphisms of ESR1, ESR2, CYP17A1, and CYP19A1 and the risk of breast cancer: a case control study from North India.

Estrogen is a key driver of breast cancer and genes involved in its signaling and biosynthesis are crucial in breast cancer progression. In this study, we investigated the role of estrogen signaling and synthesis related genes polymorphism in susceptibility to breast cancer risk in North India population in a case-control approach. We examined the association of single nucleotide polymorphism (SNP) in estrogen receptors, ESR1 (rs2234693) and ESR2 (rs2987983); estrogen biosynthesis enzymes, CYP17A1 (rs743572); and aromatase, CYP19A1 (rs700519) with breast cancer risk. Cases (n = 360) were matched to controls (n = 360) by age, sex, ethnicity, and geographical location. Results provided evidence that all the genetic variants were significantly associated with breast cancer risk among North Indian women. Furthermore, on performing stratified analysis between breast cancer risk and different clinicopathological characteristics, we observed strong associations for menopausal status, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) status, clinical stage, and histological grade. Our results suggest that these genes could be used as molecular markers to assess breast cancer susceptibility and predicting prognosis in North India population.

Mutational analysis of prohibitin--a highly conserved gene in Indian female breast cancer cases.

Prohibitin (PHB) is a chaperone protein which is highly conserved evolutionarily. It shows significant homology with the Drosophila cc gene which is considered important for development and differentiation of Drosophila melanogaster. Investigations have revealed an involvement of PHB in cellular proliferation and development, apoptosis, signal transduction, mitochondrial function and regulation of the estrogen and androgen receptors. Therefore, we conducted the present study to analyze mutations in the highly conserved region in Indian female breast cancer patients. Conventional PCR-SSCP and Automated DNA sequencing were performed with a total of 105 breast cancer samples along with adjacent normal tissue. Of the total, 14.2% (15/105) demonstrated a mutation status of prohibitin observed in our study population. We identified a novel missense mutation (Thr>Ser), a novel deletion of T nucleotide in an intron adjacent to intron-exon boundary and a previously determined missense mutation (Val>Ala). A statistically significant correlation was obtained which suggested that prohibitin may be associated with tumor development and/or progression of at least some proportion of breast cancers.