Syed Akhtar Husain

Lipid peroxidation, free radical production and antioxidant status in breast cancer

Reactive oxygen metabolites (ROMs), including superoxide anion (O2·−), hydrogen peroxide (H2O2) and hydroxyl radical (·OH), play an important role in carcinogenesis. There are some primary antioxidants such as superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) which protect against cellular and molecular damage caused by the ROMs. We conducted the present study to determine the rate of O2·− and H2O2 production, and concentration of malondialdehyde (MDA), as an index of lipid peroxidation, along with the SOD, GPx and CAT activities in 54 breast cancer (BC) patients. Forty-two age- and sex-matched patients with minor surgical problems, who had no history of any neoplastic or breast disorders, were taken as controls.The rate of O2·− production was significantly higher (p<0.001) in BC patients than controls, irrespective of clinical stages and menopausal status. Similarly, H2O2 production was significantly higher in BC patients, especially in stage III and postmenopausal groups, as compared to the respective controls. MDA concentration was also observed significantly elevated in stage II (p<0.001), stage III (p<0.01), postmenopausal (p<0.005), and premenopausal (p<0.02) group as compared to their corresponding controls. SOD and GPx activities were found significantly raised in all the groups (p<0.001), except the GPx activity was found a smaller alteration in stage IV (p<0.02). On the contrary, CAT activity was found significantly depressed in all the study groups. The maximum depression was observed in stage II (−61.8%). Lower CAT activity in our study may be the effect of higher production of ROMs, particularly O2·− and ·OH. SOD and GPx, however, were less effected by these higher ROMs production. The results of our study have shown a higher ROMs production and decreased CAT activity, which support the oxidative stress hypothesis in carcinogenesis. The relatively higher SOD and GPx may be due to the response of increased ROMs production in the blood. However, the higher SOD and GPx activities may be inadequate to detoxify high levels of H2O2 into H2O leading to the formation of the most dangerous ·OH radical followed by MDA. Therefore, administration of CAT may be helpful in the management of BC patients. However, further elaborate clinical studies are required to evaluate the role of such antioxidant enzymes in BC management.

Hepatitis E virus infection and fulminant hepatic failure during pregnancy.

Background and Aim:  Hepatitis E virus (HEV) infection leading to fulminant hepatic failure (FHF) and high mortality is a common feature in Indian women during the second and third trimesters of pregnancy. An altered status of hormones and immunity are observed during pregnancy but the actual cause of high mortality is still unknown. The present study was carried out to analyze CD3, CD4 and CD8 T cell counts and to assay the level of pregnancy‐related hormones such as estrogen, progesterone and β‐HCG in order to discover the role played by these factors.

Role of lipids, lipoproteins and vitamins in women with breast cancer.

OBJECTIVES Improper balance between the production of reactive oxygen metabolites (ROMs), and antioxidative defense system have been defined as oxidative stress in various pathologic conditions. Lipids, lipoproteins and antioxidative vitamins have been associated with the risk of breast cancer. The present case-control study was conducted to investigate the status of antioxidative vitamins (A, C and E), lipids (total cholesterol; TC and triglycerides; TG), lipoproteins (high-density lipoprotein cholesterol; HDL-C and low-density lipoprotein cholesterol; LDL-C) and retinol-binding protein (RBP) in breast cancer patients. The aim of the study was to find out oxidative stress in breast cancer. DESIGN AND METHODS Plasma lipids, lipoproteins and vitamins were estimated in 54 untreated breast cancer patients of different clinical stages and in 42 age- and sex-matched controls. RESULTS Plasma TC (p < 0.05), and LDL-C and TG (p < 0.01) were found to be significantly elevated among breast cancer patients as compared to the controls. On the other hand, plasma HDL-C concentration (p < 0.001) and vitamin C and E (p < 0.01) were observed significantly decreased in breast cancer patients than in the controls. The maximum changes in plasma TC, and vitamin C and E concentrations were observed in breast cancer patients with stage IV when compared with controls. CONCLUSION The study suggests that higher levels of TC and TG may play important role in carcinogenesis. Furthermore, the elevated plasma LDL-C concentration, which is more susceptible to oxidation, may result in higher lipid peroxidation in breast cancer patients. However, decreased concentrations of HDL-C and vitamin C and E are not likely to be sufficient enough to counter higher ROMs production reported earlier in breast cancer patients that may cause oxidative stress leading to cellular and molecular damage thereby resulting in cell proliferation and malignant conversions.

Berberine modulates AP-1 activity to suppress HPV transcription and downstream signaling to induce growth arrest and apoptosis in cervical cancer cells

Background-Specific types of high risk Human papillomaviruses (HR-HPVs) particularly, HPV types 16 and 18 cause cervical cancer and while the two recently developed vaccines against these HPV types are prophylactic in nature, therapeutic options for treatment and management of already existing HPV infection are not available as yet. Because transcription factor, Activator Protein-1 (AP-1) plays a central role in HPV-mediated cervical carcinogenesis, we explored the possibility of its therapeutic targeting by berberine, a natural alkaloid derived from a medicinal plant species, Berberis which has been shown to possess anti-inflammatory and anti-cancer properties with no known toxicity; however, the effect of berberine against HPV has not been elucidated.Results-We studied the effect of berberine on HPV16-positive cervical cancer cell line, SiHa and HPV18-positive cervical cancer cell line, HeLa using electrophoretic mobility gel shift assays, western and northern blotting which showed that berberine could selectively inhibit constitutively activated AP-1 in a dose- and time-dependent manner and downregulates HPV oncogenes expression. Inhibition of AP-1 was also accompanied by changes in the composition of their DNA-binding complex. Berberine specifically downregulated expression of oncogenic c-Fos which was also absent in the AP-1 binding complex. Treatment with berberine resulted in repression of E6 and E7 levels and concomitant increase in p53 and Rb expression in both cell types. Berberine also suppressed expression of telomerase protein, hTERT, which translated into growth inhibition of cervical cancer cells. Interestingly, a higher concentration of berberine was found to reduce the cell viability through mitochondria-mediated pathway and induce apoptosis by activating caspase-3.Conclusion-These results indicate that berberine can effectively target both the host and viral factors responsible for development of cervical cancer through inhibition of AP-1 and blocking viral oncoproteins E6 and E7 expression. Inhibition of AP-1 activity by berberine may be one of the mechanisms responsible for the anti-HPV effect of berberine. We propose that berberine is a potentially promising compound for the treatment of cervical cancer infected with HPV.

Does Hepatitis E Viral Load and Genotypes Influence the Final Outcome of Acute Liver Failure During Pregnancy

BACKGROUND:Hepatitis E is a major health problem in developing countries including India. The incidence and mortality rate in pregnant women with fulminant hepatic failure (FHF) due to hepatitis E virus (HEV) has been reported to be significantly higher, specifically in Asian women. Pregnancy is usually associated with an altered status of sex steroid hormones and immunity. Steroid hormones directly influence the replication through their effects on viral regulatory elements. Moreover, pregnant women in Asia generally suffer from folate deficiency, which is known to cause reduced immunocompetence leading to greater risk of multiple viral infections and higher viral load.OBJECTIVES:To correlate and analyze the viral load and genotypes of HEV in acute liver failure with that of acute viral hepatitis among pregnant and nonpregnant women.MATERIALS AND METHODS: A total of 100 FHF and 150 acute viral hepatitis (AVH) patients (50, 75 pregnant and 50, 75 nonpregnant, respectively), were included in the study. These cases were evaluated on the basis of history, clinical examination, liver function profile, and serological test of hepatitis A, B, C, and E using commercially available ELISA kits. Quantification of HEV RNA-positive samples was carried out.RESULTS:Out of 100 FHF and 150 acute viral hepatitis (AVH) patients, 28 (56%) and 22 (29.3%) pregnant and 7 (14%) and 8 (16%) nonpregnant, respectively, were HEV RNA-positive. HEV viral load in FHF pregnant women was 5.87 × 104± 1.5 × 105μ L/mL as compared to AVH pregnant women 343.29 ± 216.44 μL/mL and FHF and AVH nonpregnant 199.2 ± 225.5 μL/mL and 13.83 ± 7.8 μL/mL, respectively. Sequencing data of all the positive samples of FHF and AVH pregnant and nonpregnant women showed genotype 1.CONCLUSION:HEV viral load was found to be significantly higher (P < 0.05) in pregnant patients compared to the nonpregnant. Pregnancy appears to be a risk factor for viral replication. The viral copies of HEV in FHF pregnant women were comparatively higher when compared to AVH pregnant women, which may be related to the severity of the disease in these patients. We could detect only one genotype (genotype 1) in our study population. Thus in the absence of other genotypes in this population, the impact of genotype could not be adequately assessed in this study.

CpG methylation of the FHIT, FANCF, cyclin-D2, BRCA2 and RUNX3 genes in Granulosa cell tumors (GCTs) of ovarian origin

BackgroundGranulosa cell tumors (GCTs) are relatively rare and are subtypes of the sex-cord stromal neoplasms. Methylation induced silencing in the promoters of genes such as tumor suppressor genes, DNA repair genes and pro-apoptotic genes is recognised as a critical factor in cancer development.MethodsWe examined the role of promoter hypermethylation, an epigenetic alteration that is associated with the silencing tumor suppressor genes in human cancer, by studying 5 gene promoters in 25 GCTs cases by methylation specific PCR and RT-PCR. In addition, the compatible tissues (normal tissues distant from lesion) from three non-astrocytoma patients were also included as the control.ResultsFrequencies of methylation in GCTs were 7/25 (28 % for FHIT), 6/25 (24% for FNACF), 3/25 (12% for Cyclin D2), 1/25 (4% for BRCA2) and 14/25 (56%) in RUNX3 genes. Correlation of promoter methylation with clinical characteristics and other genetic changes revealed that overall promoter methylation was higher in more advanced stage of the disease. Promoter methylation was associated with gene silencing in GCT cell lines. Treatment with methylation or histone deacetylation-inhibiting agents resulted in profound reactivation of gene expression.ConclusionsThese results may have implications in better understanding the underlying epigenetic mechanisms in GCT development, provide prognostic indicators, and identify important gene targets for treatment.

Role of polymorphic N-acetyl transferase2 and cytochrome P4502E1 gene in antituberculosis treatment-induced hepatitis.

Background and Aim:  Antituberculosis drugs, isoniazid and rifampicin, in combination, are known to develop drug‐induced hepatotoxicity (DIH). A higher risk of DIH during antituberculosis treatment (ATT) has been reported in the Indian subcontinent compared to its Western counterparts. The role of genetic factors in a higher incidence of ATT hepatotoxicity in the Indian population is still unclear. The present study was aimed at investigating the role of the N‐acetyltransferase2 (NAT2) and cytochrome P4502E1 (CYP2E1) gene polymorphisms in ATT hepatotoxicity.

Novel germline mutations in breast cancer susceptibility genes BRCA1, BRCA2 and p53 gene in breast cancer patients from India

Mutations in breast cancer susceptibility genes, BRCA1 and BRCA2 account for more than 80% of hereditary breast and ovarian cancers. p53 tumor suppressor gene that controls cellular growth and differentiation is also known to be mutated in more than 50% of human cancers including breast cancer. We have carried out a study on BRCA1 and BRCA2 along with p53 gene mutations in both sporadic as well as familial breast cancer patients from India where breast cancer is fast emerging as a major cancer among premenopausal urban women. We examined 124 untreated primary breast cancer patients comprising 100 sporadic and 24 familial cases including 56 age-matched healthy controls for the presence of BRCA1, BRCA2 and the p53 gene mutations using PCR-SSCP and direct nucleotide sequencing. Certain frequently mutated exons such as 2, 5, 11, 13 and 20 of BRCA1, exons 2, 9, 11 (for 6174delT), 18 and 20 of BRCA2 and 4–9 exons of p53 gene were analyzed in sporadic breast cancer while all 22 coding exons of BRCA1 including its flanking intronic regions along with above mentioned exons of BRCA2 and p53 gene were analyzed in familial breast cancer patients. We identified six patients (25%) with BRCA1 mutation of which three were found to be of novel type one in exon 16 (4956insG) and two in exon 7 (Lys110Thr) (Ser114Pro) out of 24 familial breast cancer patients studied from two different geographic regions/populations of India. Two sisters from a single family (12.5%) out of eight families from Goa with Portuguese colonial origin showed presence of founder Ashkenazi Jewish BRCA1 mutation (185delAG) along with (IVS7 561−34T>C; IVS18 527166G>A). While from New Delhi, four (25%) of 16 breast cancer families showed BRCA1 mutations; a frame shift protein truncating (4956insG), a transition nonsense (Gln1395Stop) and two amino acid substitutions (Lys110Thr) and (Ser114Pro). Only one (4%) p53 mutation (Val97Ile) in its exon 4 along with BRCA1 mutation (4956insG) could be detected. No major sequence variation in BRCA2 gene was observed except for G203A at 5′ UTR of exon 2, a common population polymorphism in two Goan patients who also showed silent nucleotide change for amino acid serine at codon 1436 of BRCA1 gene. None of the 100 sporadic breast cancer patients revealed any protein truncating or deleterious BRCA1 or BRCA2 gene mutation. Interestingly, three (3%) p53 mutations in its exon 5 were detected in sporadic breast cancer patients. Although three novel BRCA1 mutations including a founder Ashkenazi Jewish BRCA1 mutation were recorded in Indian women with familial breast cancer, the overall prevalence of BRCA gene mutations in Indian women with a family history of breast cancer appears to be low.

Polymorphism of the p53 codon 72 arg/pro and the risk of HPV type 16/18-associated cervical and oral cancer in India

Infection of high risk human papillomaviruses (HPVs) specifically the types 16 and 18 has been strongly implicated in the development of cervical cancer. The E6 oncoproteins of these high risk HPVs are known to bind and induce degradation of p53 tumour suppressor protein through the ubiquitin pathways. This degradation is controlled by a common polymorphism of the p53 gene encoding either a proline or an arginine at its codon 72 in exon 4. Recently, it has been demonstrated that the presence of homozygous arginine at codon 72 renders p53 about seven times more susceptible to E6-mediated proteolytic degradation as well as to cervical cancer than those with proline homozygotes or proline/arginine heterozygotes. In India, prevalence of HPV as well as cancers of the uterine cervix and the oral cavity are highest in the world. We have examined this allele-specific predisposition in cervical and oral cancer which is associated with HPV as well as in a non-HPV-linked cancer of the breast. We have carried out investigation in women comprising whole spectrum of cervical lesions with 128 HPV 16/18 positive and 35 HPV negative invasive cervical carcinomas and 34 cases of HPV (16/18) positive and 16 HPV negative cervical dysplasias (mild, moderate and severe) and 104 age-group-matched healthy women as controls. Additionally, we have analysed p53Arg-Pro polymorphism in 13 high risk HPV positive and 31 HPV negative oral cancers along with 20 normal controls and 77 breast cancers with 41 age-matched healthy controls.We observed more than 2 fold higher risk for homozygous arginine (χ2 = 6.3, df = 2, p = 0.04; OR = 2.3; 95% CI: 1.08–5.16) for HPV 16/18-positive cervical carcinomas when comparison was made only between HPV positive cervical cancers and normal controls but most interestingly, no significant association either in the frequency of homozygous arginine or proline alleles or their heterozygotes could be observed when all the three groups i.e. HPV-positive, HPV-negative cervical cancers and controls were considered simultaneously. No difference was also observed for either arginine or proline polymorphism between women with precancerous lesions of the uterine cervix carrying HPV 16/18 infection and controls. Similarly, increased risk of oral or breast cancer could not be correlated with the polymorphism of arginine/proline allele.Thus the interaction between HPV oncoproteins and the p53 gene polymorphism specifically, homozygous arginine at codon 72 appears to play no role in the development of either cervical or oral cancer and also it can not serve as a biomarker for early identification of cervical, oral or breast cancer.

Increasing trend of acute hepatitis A in north India: Need for identification of high-risk population for vaccination

Background and Aims:  Hepatitis A (HAV) is endemic in India and most of the population is infected asymptomatically in early childhood with lifelong immunity. Because of altered epidemiology and decreasing endemicity, the pattern of acute HAV infection is changing from asymptomatic childhood infection to an increased incidence of symptomatic disease in the 18–40 age group. The aims of the present study were to assess whether the proportion of adults with acute HAV infection has been increasing over the years and to analyze the seroprevalence of immunoglobulin G (IgG) anti‐HAV antibodies in young adults above the age of 15 years as well as in cases of chronic liver disease.