Mohammad Muzaffar Mir

p53 mutation profile of squamous cell carcinomas of the esophagus in Kashmir (India): a high-incidence area.

Esophageal squamous cell carcinoma (ESCC) has been reported to show geographical variation in its incidence, even within areas of ethnic homogeneity. Kashmir valley, in north of India, has been described as a high‐risk area for ESCC. Here, we make a preliminary attempt to study mutations in exons 5–8 (the DNA binding domain) of the tumor suppressor gene, p53, in 55 ESCC patients from Kashmir. Polymerase chain reaction followed by direct sequencing analysis revealed the presence of mutations in 36.36% (20/55) tumors, assessed for the extent of allelic instability. The 20 mutations, found in 20 patients, comprised of 17 single‐base substitutions (11 transitions + 6 transversions) and 3 deletions. The 17 single‐base variations represented 12 missense mutations, 2 nonsense mutations and 3 variations located in intron 6, 1 of which resulted in a splicing variant. The patients when compared for the incidence of p53 mutation with various demographic features revealed females to be at increased risk (p = 0.016; OR = 4.13; 95% CI = 1.26–13.46). Comparison of mutation profile with other high‐risk areas reflected both differences and similarities indicating coexposure to a unique set of risk factors. This might be due to the special dietary and cultural practices of Kashmir that needs validation, as does the gender‐based difference in the incidence of p53 mutation observed in this study.

Breast cancer and human papillomavirus infection: no evidence of HPV etiology of breast cancer in Indian women.

BackgroundTwo clinically relevant high-risk HPV (HR-HPV) types 16 and 18 are etiologically associated with the development of cervical carcinoma and are also reported to be present in many other carcinomas in extra-genital organ sites. Presence of HPV has been reported in breast carcinoma which is the second most common cancer in India and is showing a fast rising trend in urban population. The two early genes E6 and E7 of HPV type 16 have been shown to immortalize breast epithelial cells in vitro, but the role of HPV infection in breast carcinogenesis is highly controversial. Present study has therefore been undertaken to analyze the prevalence of HPV infection in both breast cancer tissues and blood samples from a large number of Indian women with breast cancer from different geographic regions.MethodsThe presence of all mucosal HPVs and the most common high-risk HPV types 16 and 18 DNA was detected by two different PCR methods - (i) conventional PCR assays using consensus primers (MY09/11, or GP5+/GP6+) or HPV16 E6/E7 primers and (ii) highly sensitive Real-Time PCR. A total of 228 biopsies and corresponding 142 blood samples collected prospectively from 252 patients from four different regions of India with significant socio-cultural, ethnic and demographic variations were tested.ResultsAll biopsies and blood samples of breast cancer patients tested by PCR methods did not show positivity for HPV DNA sequences in conventional PCRs either by MY09/11 or by GP5+/GP6+/HPV16 E6/E7 primers. Further testing of these samples by real time PCR also failed to detect HPV DNA sequences.ConclusionsLack of detection of HPV DNA either in the tumor or in the blood DNA of breast cancer patients by both conventional and real time PCR does not support a role of genital HPV in the pathogenesis of breast cancer in Indian women.

Transcription factor AP-1 in esophageal squamous cell carcinoma: Alterations in activity and expression during Human Papillomavirus infection

BackgroundEsophageal squamous cell carcinoma (ESCC) is a leading cause of cancer-related deaths in Jammu and Kashmir (J&K) region of India. A substantial proportion of esophageal carcinoma is associated with infection of high-risk HPV type 16 and HPV18, the oncogenic expression of which is controlled by host cell transcription factor Activator Protein-1 (AP-1). We, therefore, have investigated the role of DNA binding and expression pattern of AP-1 in esophageal cancer with or without HPV infection.MethodsSeventy five histopathologically-confirmed esophageal cancer and an equal number of corresponding adjacent normal tissue biopsies from Kashmir were analyzed for HPV infection, DNA binding activity and expression of AP-1 family of proteins by PCR, gel shift assay and immunoblotting respectively.ResultsA high DNA binding activity and elevated expression of AP-1 proteins were observed in esophageal cancer, which differed between HPV positive (19%) and HPV negative (81%) carcinomas. While JunB, c-Fos and Fra-1 were the major contributors to AP-1 binding activity in HPV negative cases, Fra-1 was completely absent in HPV16 positive cancers. Comparison of AP-1 family proteins demonstrated high expression of JunD and c-Fos in HPV positive tumors, but interestingly, Fra-1 expression was extremely low or nil in these tumor tissues.ConclusionDifferential AP-1 binding activity and expression of its specific proteins between HPV - positive and HPV - negative cases indicate that AP-1 may play an important role during HPV-induced esophageal carcinogenesis.

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.

Association of cyclin D1 gene polymorphisms with risk of esophageal squamous cell carcinoma in Kashmir Valley : a high risk area

Investigation of potential association of SNPs (G870A, rs9344; G1722C, rs678653) of cyclin D1 gene (CCND1) with susceptibility to esophageal squamous cell carcinoma (ESCC) in Kashmir valley (India). The study included 302 subjects comprising 151 ESCC cases and 151 controls. PCR‐RFLP and direct sequencing were employed for genotyping. The G870A polymorphism, the individuals carrying GA + AA genotype was having 2.80‐fold increased risk for development of ESCC (OR 2.8, 95% CI = 1.77–4.4; P = 0.0001) compared to GG genotype. Further a significantly higher risk was observed in individuals who consume >3 cups per day of salted tea (OR = 5.1; 95% CI = 1.6–16.7; P = 0.0016) and had smoking habits (OR = 6.3; 95% CI = 2.9–13.9; P = 0.0005). We also demonstrate for the first time in CCND1 1722 locus, the CC genotype was strongly associated with increased risk of developing ESCC (OR = 2.58; 95% CI = 1.61–4.15; P = 0.0001). In addition, the frequency of polymorphic C allele was also found to be higher in cases (OR = 1.92; 95% CI = 1.37–2.69; P = 0.0002). There appears to be an influence of CCND1 G870A/G1772C genotypes on genetic susceptibility to ESCC. Mol. Carcinog. ©2011 Wiley‐Liss, Inc.

Association Between Copper Excess, Zinc Deficiency, and TP53 Mutations in Esophageal Squamous Cell Carcinoma From Kashmir Valley, India—A High Risk Area

Trace element deficiency or excess is implicated in the development or progression in some cancers. Here we report the elevated level of copper and low level of zinc in the plasma of esophageal cancer patients in Kashmir India—a high incidence area. The average level of copper was significantly higher (P < 0.0001) for patients than for controls, with a mean concentration of 169 μg/dl and 149 μg/dl for patients and controls, respectively. The control group consisted of 55 healthy individuals matched for age, sex, and place of residence of the patients. In contrast, the average level of zinc in patients was significantly lower than in controls (P < 0.0001), with a mean concentration of 86.8 μg/dl and 96.1 μg/dl for patients and controls, respectively. The levels of both copper and zinc showed significant differences based on gender and age in patients as compared to controls. Similarly, smokers depicted a significant increase in serum copper (N = 39, P = 0.002) and a decrease in serum zinc approaching level of significance in the patient group as compared to controls. The copper and zinc levels were significantly altered in patients (N = 40) when compared to controls as a function of snuff consumption. The differences in the levels of copper and zinc showed significant association with the consumption of local salted tea up to 1,500 ml per day, but the changes were insignificant beyond that. Patients with poorly differentiated tumors (N = 7) had a higher copper concentration than those with moderately or well-differentiated tumors (P < 0.0001). To validate the general notion that imbalance in copper and zinc levels may lead to higher prevalence of TP53 mutations, we compared the 3 variables, and no association was found between copper concentration and TP53 mutation status; but patients with TP53 mutant tumor had lower zinc levels than those with no mutation. In conclusion, our results point toward a role of the trace element imbalance in the esophageal tumorigenesis in high-risk Kashmiri population exposed to a range of nitroso compounds or their precursors. Further prospective cohort studies are warranted to determine whether change in the plasma zinc and copper homeostasis may represent an independent risk factor for this malignancy as well as a possible target for preventive intervention.

Abstract 2722: Transcription factor NF-kB in esophageal squamous cell carcinoma: Alterations in activity and expression during Human Papillomavirus infection

Background: Esophageal squamous cell carcinoma (ESCC) is a leading cause of cancer-related deaths in Jammu and Kashmir (JK 46/61). On the other hand, HPV16 infected tumor tissues (n=14) showed hetrodimerization between p50 and cRel in more than 85% (12/14) of esophageal tumors. Interestingly, 2 cases with HPV infection p65 was also found to participate along with p50 and cRel in well-differentiated state of esophageal cancer. It is quite likely that the functional NF-kB complex formation in advanced grade of esophageal cancer comprises of p50/p50 homodimerization whereas in HPV infected esophageal tumors p50/cRel hetrodimerization occurs along with p65 in well differentiated state of the disease. Conclusion: Differential NF-kB dimerization and expression of its specific proteins between HPV – positive and HPV – negative cases indicate that NF-kB may play an important role during HPV induced esophageal carcinogenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2722. doi:10.1158/1538-7445.AM2011-2722

Polymorphonuclear leukocyte mediated oxidative inactivation of alpha-1-proteinase inhibitor: Modulation by nitric oxide

Alpha-1-proteinase inhibitor activity was studied in presence of resting and activated polymorphonuclear leucocytes. Four different agonists; phorbol myristic acetate, N-formyl-methionyl-leucyl-phenylalanine, opsonised zymosan and arachidonic acid decreased the inhibitor activity by 23.3%, 20%, 12% and 16.6^ respectively. The inhibitor activity was protected by using various free radical scavengers. Catalase and superoxide dismutase both restored activity by about 18%, mannitol by 13% and sodium azide by 17.3%. The inhibitor activity was also protected significantly by pretreatment of polymorphs with L-Arg, a precursor of nitric oxide, before activation. L-Arg was also observed to suppress the generation of superoxide and hydroxyl radical appreciably. The nitric oxide synthase inhibitor, aminoguanidine drastically inhibited the nitrite release and reversed the protection offered by L-Arg to the inhibitor activity. Our results indicate a multifactorial nature of the inactivation process, the culprit species being superoxide, hydrogen peroxide, hydroxyl radical and hypohalides. Nitric oxide seems to scavenge the superoxide radical directly after its formation rather than inhibiting its generation by NADPH oxidase as was believed earlier.