Mausumi Bharadwaj

TNFα–308G/A Polymorphism as a Risk Factor for HPV Associated Cervical Cancer in Indian Population

Background: Investigation of the potential association of single nucleotide polymorphisms (SNPs) at –308 G/A and –238 G/A of Tumor necrosis factor α (TNFα) with susceptibility to HPV-16 associated cervical cancer in Indian women. Methods: The study included 165 histologically confirmed cases with 45 precancer and 120 cancer patients and an equal number (165) of healthy controls with normal cervical cytology. PCR-RFLP was employed to analyze TNFα promoter polymorphisms, which were confirmed by direct sequencing. Both patients and controls were screened for Human Papillomavirus (HPV) infection. Results: The frequency of –308 A allele in TNFα was significantly higher in cases compared with control subjects (21% in cases vs. 9% in controls; p < 0.01), with an odds ratio of 2.7 (95% CI = 1.41–5.15). Also, women carrying A allele for this locus presented 3 times increased susceptibility to HPV 16 infection as evident from carrier genotype distribution between HPV positive cases and control subjects (24% in HPV positive cases vs. 9% in controls; p < 0.01; OR = 3.1; 95% CI = 1.60–6.03). No such association was found for TNFα–238 (G/A) polymorphism with the risk of development of cervical cancer. Conclusion: It suggests that SNP at –308 (G/A) of TNFα promoter may represent an increased risk for HPV infection and development of cervical cancer in Indian women.

Kras Gene Mutation and RASSF1A, FHIT and MGMT Gene Promoter Hypermethylation: Indicators of Tumor Staging and Metastasis in Adenocarcinomatous Sporadic Colorectal Cancer in Indian Population

Objective Colorectal cancer (CRC) development involves underlying modifications at genetic/epigenetic level. This study evaluated the role of Kras gene mutation and RASSF1A, FHIT and MGMT gene promoter hypermethylation together/independently in sporadic CRC in Indian population and correlation with clinicopathological variables of the disease. Methods One hundred and twenty four consecutive surgically resected tissues (62 tumor and equal number of normal adjacent controls) of primary sporadic CRC were included and patient details including demographic characteristics, lifestyle/food or drinking habits, clinical and histopathological profiles were recorded. Polymerase chain reaction - Restriction fragment length polymorphism and direct sequencing for Kras gene mutation and Methylation Specific-PCR for RASSF1A, FHIT and MGMT genes was performed. Results Kras gene mutation at codon 12 & 13 and methylated RASSF1A, FHIT and MGMT gene was observed in 47%, 19%, 47%, 37% and 47% cases, respectively. Alcohol intake and smoking were significantly associated with presence of Kras mutation (codon 12) and MGMT methylation (p-value <0.049). Tumor stage and metastasis correlated with presence of mutant Kras codon 12 (p-values 0.018, 0.044) and methylated RASSF1A (p-values 0.034, 0.044), FHIT (p-values 0.001, 0.047) and MGMT (p-values 0.018, 0.044) genes. Combinatorial effect of gene mutation/methylation was also observed (p-value <0.025). Overall, tumor stage 3, moderately differentiated tumors, presence of lymphatic invasion and absence of metastasis was more frequently observed in tumors with mutated Kras and/or methylated RASSF1A, FHIT and MGMT genes. Conclusion Synergistic interrelationship between these genes in sporadic CRC may be used as diagnostic/prognostic markers in assessing the overall pathological status of CRC.

High-sensitivity C-reactive protein levels and type 2 diabetes in urban North Indians.

CONTEXT Elevated high-sensitivity C-reactive protein (hsCRP) levels have frequently been shown to be associated with type 2 diabetes (T2D); however, very little is known about this in Asian Indians, a high-risk group. OBJECTIVE The aim of the study was to assess the association of hsCRP with T2D and to determine its correlates in North Indians of Indo-European origin. DESIGN AND PATIENTS A cross-sectional population-based study of 2520 urban subjects, comprising 1410 T2D patients and 1110 nondiabetic subjects, was carried out and 18 metabolic traits were assessed. RESULTS Median hsCRP levels were significantly higher in both diabetic men and women as compared to their nondiabetic counterparts (P < 0.0001). Elevated hsCRP was positively associated with T2D (odds ratio, 1.66; 95% confidence interval, 1.21-2.28; P = 0.002) even after adjusting for markers of obesity. After adjustments for age, sex, and BMI, HbA1c was the major correlate of hsCRP in nondiabetic subjects (beta = 0.28; P = 0.03). We observed that T2D patients were at higher risk for cardiovascular disease compared to nondiabetic subjects when classified into low-, intermediate-, and high-risk groups based on hsCRP levels (p(trend) = 3.8 x 10(-15)). CONCLUSIONS We demonstrate the association of low-grade systemic inflammation, as indicated by elevated hsCRP levels, with T2D in North Indian population. This association was independent of obesity. Obesity and glycemic control were the major correlates of hsCRP levels. Future studies are required to evaluate the influence of modulators including genetic variations on the elevation of hsCRP levels in this population.

Association between human leukocyte antigen class II alleles and human papillomavirus-mediated cervical cancer in Indian women.

We investigated the association of human leukocyte antigen (HLA) II (DRB1 and DQB1) alleles with susceptibility to human papillomavirus (HPV)-associated cervical precancer and cancer cases in a hospital-based case-control study in a northern Indian population. A total of 202 subjects, including 100 patients comprising 31 cervical precancer (cervical intraepithelial neoplasia [CIN] 2/3) and 69 invasive cervical cancer cases, and 102 healthy controls participated in the study. Both patients and controls were screened for HPV infection using a polymerase chain reaction (PCR-based approach. Low-resolution PCR-sequence specific priming (PCR-SSP) was used to genotype HLA II (DRB1 and DQB1). Our results demonstrate that the DRB1*15 allele/DRB1*15-DQB1*06 haplotype may have a predisposition for HPV infection (p(c) < 0.05) or cervical cancer/precancer (p(c) < 0.05) development, whereas the DRB1*04 allele/DRB1*04-DQB1*03 haplotype might exhibit susceptibility to cervical precancerous lesions (p(c) < 0.05). The DRB1*13 allele/DRB1*13-DQB1*06 haplotype was strongly protective against risk to HPV infection (p(c) < 0.002) as well as cervical cancer (p(c) 0.01). Therefore, we have demonstrated that HLA DR-DQ polymorphisms are involved in genetic susceptibility to cervical cancer or HPV infection in a northern Indian population.

Effect of aberrant promoter methylation of FHIT and RASSF1A genes on susceptibility to cervical cancer in a North Indian population

Abstract As current evidence suggests the involvement of epigenetic modification of tumour suppressor genes in human cancer, we investigated the aberrant promoter methylation of FHIT and RASSF1A genes in human papillomavirus (HPV)-mediated cervical cancer in Indian women. We analysed 60 cervical cancer tissue biopsies of different clinical stage and histological grading and 23 healthy control samples with normal cervical cytology. Methylation-specific polymerase chain reaction (MSP) was performed to analyse the methylation status of FHIT and RASSF1A genes and confirmed by sequencing. Both patients and controls were screened for HPV infection and 98% of the HPV-infected cases showed positivity for HPV type 16. Aberrant promoter methylation of the FHIT gene was found in 28.3% (17/60) of cases and of the RASSF1A gene in 35.0% (21/60) of cases; promoter methylation of both the genes was found in 13.3% (8/60) of cervical cancer cases. Methylation was significantly (p<0.01) associated with the cervical cancer cases compared with controls. None of the 23 controls was found to be methylated in either of these genes. This is the first study indicating a correlation between the promoter methylation of FHIT and RASSF1A genes and the clinical stage and histological grading of cervical carcinoma in Indian women. Future studies are underway to examine the practical implications of these findings for use as a biomarker.

Association of MDM2 and p53 Polymorphisms with the Advancement of Cervical Carcinoma.

Cervical cancer is one of the most common gynecological malignancies that causes a serious health problem worldwide. The aim of the present study was to analyze the association of p53 codon72 (arginine/proline) polymorphism (rs1042522) and Murine Double Minute 2 (MDM2) SNP309 T/G (rs2279744) with the advancement of cervical cancer by using polymerase chain reaction-restriction fragment length polymorphism method followed by direct sequencing. The frequencies of GG genotype at 309 position in the second promoter (P2) of MDM2 and Arginine in codon72 of p53 were found to be 3.5 (odds ratio [OR]=3.51; 95% confidence interval [CI]=1.93-6.4; p<0.0001) and 5 (OR=4.978; 95% CI=2.7-9.2; p<0.0001) fold higher, respectively, in cases than in the control. On gene-gene interactions between MDM2 and p53 polymorphisms, the frequency of MDM2 G/G and p53 Arg/Arg together was found to be 6.5-fold higher in cervical cancer patients compared with healthy controls (OR=6.497; 95% CI=2.987-14.13; p<0.0001). We found an association of p53 codon72 arginine and MDM2 SNP309 GG genotype with different clinical and histological grades, human papillomavirus (HPV) infection, and age at the time of diagnosis of cervical cancer. In conclusion, Arginine at codon72 of p53 and GG genotype at 309 in P2 of MDM2 together reveal a direct proportionality with the tumor grade of cervical cancer along with HPV infection in postmenopausal women.

Human papillomavirus infection among young adolescents in India: Impact of vaccination

High‐risk human papillomaviruses (HR‐HPVs) are the causative agents of cervical cancer and prophylactic HPV vaccination has been recommended for adolescents but no data are available on the prevalence of HPV infection among adolescents in India. Self‐collected midstream urine samples from 940 healthy school children, aged 8–17 years, from 12 different schools in and around Noida and Delhi, India, were collected for HPV detection by PCR. Of 458 girls, 15 (3.2%) were positive for HPV and 10 (66.6%) were positive for high‐risk human papillomavirus (HR‐HPV) type16 and 2 (13.3%) for HPV 18. Of 342 boys, 7 (2.1%) were HPV positive, of which 5 (71.4%) had HPV type 6 but interestingly, none were positive for HR‐HPV types 16 or 18. Among HPV positive girls, 13 (66.6%) were >13 years and the rest were <13 years (P = 0.004), while all seven HPV positive boys were >13 years (P = 0.007). The majority of HPV positive adolescents (80–86%) belonged to the Hindu and related communities, whereas only about 14–20% belonged to the Muslim community. A significant association (P < 0.001) was observed between the parents education and the awareness of cervical cancer, which was significantly higher among adolescent girls from India, thereby exerting an immense psychosocial impact on vaccination programs. A lower prevalence of HR‐HPV infection among adolescent girls will have significant positive effect on HPV vaccination and cancer control programs in India where education and awareness should go hand in hand. J. Med. Virol. 84:298–305, 2012.

Homocysteine levels are associated with cervical cancer independent of methylene tetrahydrofolate reductase gene (MTHFR) polymorphisms in Indian population

Human papillomavirus is considered to be a major aetiological factor but is not sufficient for the development of cervical cancer. Other host factors, including altered homocysteine levels, a functional marker of folate inadequacy, might contribute to the carcinogenic process. Herein we investigated the potential association of homocysteine levels and MTHFR polymorphisms with cervical cancer in 203 histologically confirmed cases including 39 precancer cases and 231 healthy controls with normal cervical cytology. Both patients and controls were screened for human papillomavirus infection. We found that homocysteine and consequently cysteine levels were significantly higher in cases, both cancer and precancer (p < 0.001) than controls. However, polymorphisms in the MTHFR gene (677C/T and 1298A/C) that are reported to modulate homocysteine levels were not associated with disease. Thus, our study establishes an association of total homocysteine levels with the risk of developing carcinoma of the uterine cervix.

Methylation-mediated gene silencing of suppressor of cytokine signaling-1 (SOCS-1) gene in esophageal squamous cell carcinoma patients of Kashmir valley.

Context: Esophageal squamous cell carcinoma (ESCC) is a leading cause of cancer-related deaths in Jammu and Kashmir. The negative regulation of tumor suppressor gene leading to change in signaling pathway is one of the major mechanisms responsible for tumorigenic transformation. Objective: In the present study, the role of silencing of suppressor of cytokine signaling-1 (SOCS-1) gene, a negative regulator of JAK/STAT pathway, was analyzed in ESCC. Methods: The expression pattern of SOCS-1 gene was analyzed in esophageal tumor biopsies although normal adjacent tissues that served as controls. Reverse transcriptase polymerase chain reaction (RT-PCR), immunohistochemistry, methylation-specific PCR (MSP), and human papillomavirus (HPV) detection were performed to assess the expression pattern and promoter methylation of SOCS-1 gene including HPV status in a total of 75 surgically resected tissue specimens. Results: Compared with the level of SOCS-1 expression in normal tissues, 53% (40/75) of the tumor tissues expressed either undetectable or reduced SOCS-1 expression (>50% loss of expression), which was significantly associated with advanced clinical stage or severe histopathological grade of the disease (P < 0.01). Aberrant promoter methylation of the SOCS-1 gene was found in 45% (34/75) of the esophageal tumor tissues, which was also found to be significantly associated with advanced stage of esophageal carcinoma (P < 0.01). The prevalence of HPV infection was found in 19% of tumor cases, whereas no HPV could be detected in any of the normal adjacent tissues. Conclusion: Transcriptional inactivation of SOCS-1 gene, primarily due to its promoter hypermethylation although HPV infection, may play an important role in esophageal carcinogenesis in Kashmir.

Deregulation of STAT-5 isoforms in the development of HPV-mediated cervical carcinogenesis

Background: Cervical cancer is the second most common cancer and is leading cause of cancer related deaths in women worldwide. High Risk-Human papillomavirus (HPV) types play an important role in cervical carcinogenesis. Considering the important role of signal transducer and activator of transcription-5 (STAT-5), an important member of JAK/STAT family which plays a crucial role in various cancers and HPV as a key mediator in the development of cervical carcinogenesis, the purpose of the current study was to examine the possible relationship between HPV infection and expression of STAT-5 gene isoforms in cervical cancer. Methods: A total of 120 fresh cervical tissue specimens comprising precancer (n = 12), cancer (n = 78) and normal controls (n = 30) were analyzed for HPV infection and expression pattern of STAT-5 mRNA (both isoforms STAT-5a and STAT-5b) and protein in different stages of cervical carcinoma biopsies by reverse-transcriptase-PCR, western blotting and immunohistochemistry. Results: A significantly increased expression of STAT-5 was detected in most of the cervical tumors (P < 0.001), whereas it was almost undetectable in normal controls. Also the study of relative contribution of STAT-5 isoforms revealed a higher expression pattern of STAT-5b and was associated with severity of the disease. On the contrary, STAT-5a was found to be significantly downregulated in cervical tumor tissues (P < 0.001). HPV infection was found in 90% of the cervical cancer cases and was significantly associated with STAT-5 overexpression (P = 0.001). Conclusions: We observed for the first time the differential expression pattern of STAT-5 isoforms in cervical cancer and that STAT-5 may play an important role in the progression of HPV-mediated cervical cancer.