Sujala Kapur

Multiple analytical approaches reveal distinct gene-environment interactions in smokers and non smokers in lung cancer.

Complex disease such as cancer results from interactions of multiple genetic and environmental factors. Studying these factors singularly cannot explain the underlying pathogenetic mechanism of the disease. Multi-analytical approach, including logistic regression (LR), classification and regression tree (CART) and multifactor dimensionality reduction (MDR), was applied in 188 lung cancer cases and 290 controls to explore high order interactions among xenobiotic metabolizing genes and environmental risk factors. Smoking was identified as the predominant risk factor by all three analytical approaches. Individually, CYP1A1*2A polymorphism was significantly associated with increased lung cancer risk (OR = 1.69;95%CI = 1.11–2.59,p = 0.01), whereas EPHX1 Tyr113His and SULT1A1 Arg213His conferred reduced risk (OR = 0.40;95%CI = 0.25–0.65,p<0.001 and OR = 0.51;95%CI = 0.33–0.78,p = 0.002 respectively). In smokers, EPHX1 Tyr113His and SULT1A1 Arg213His polymorphisms reduced the risk of lung cancer, whereas CYP1A1*2A, CYP1A1*2C and GSTP1 Ile105Val imparted increased risk in non-smokers only. While exploring non-linear interactions through CART analysis, smokers carrying the combination of EPHX1 113TC (Tyr/His), SULT1A1 213GG (Arg/Arg) or AA (His/His) and GSTM1 null genotypes showed the highest risk for lung cancer (OR = 3.73;95%CI = 1.33–10.55,p = 0.006), whereas combined effect of CYP1A1*2A 6235CC or TC, SULT1A1 213GG (Arg/Arg) and betel quid chewing showed maximum risk in non-smokers (OR = 2.93;95%CI = 1.15–7.51,p = 0.01). MDR analysis identified two distinct predictor models for the risk of lung cancer in smokers (tobacco chewing, EPHX1 Tyr113His, and SULT1A1 Arg213His) and non-smokers (CYP1A1*2A, GSTP1 Ile105Val and SULT1A1 Arg213His) with testing balance accuracy (TBA) of 0.6436 and 0.6677 respectively. Interaction entropy interpretations of MDR results showed non-additive interactions of tobacco chewing with SULT1A1 Arg213His and EPHX1 Tyr113His in smokers and SULT1A1 Arg213His with GSTP1 Ile105Val and CYP1A1*2C in nonsmokers. These results identified distinct gene-gene and gene environment interactions in smokers and non-smokers, which confirms the importance of multifactorial interaction in risk assessment of lung cancer.

Genome-wide analysis of chromosomal alterations in patients with esophageal squamous cell carcinoma exposed to tobacco and betel quid from high-risk area in India

Genomic alterations such as chromosomal amplifications, deletions and loss of heterozygosity play an important role in the pathogenesis and progression of cancer. Environmental risk factors contribute to the development and progression of tumors by facilitating the loss of tumor suppressor genes and amplification of oncogenes. In this current study, Affymetrix 10K single nucleotide polymorphism (SNP) arrays were used to evaluate genomic alterations in 20 pairs of matched germ-line and tumor DNA obtained from patients with esophageal squamous cell carcinoma (ESCC) from high-risk area of India where tobacco, betel quid and alcohol use are widespread. Twenty-two amplified regions and 16 deleted regions identified across chromosomal arms were biologically relevant. The candidate genes located at amplified regions of chromosomes or low-level gain regions such as PLA2G5 (1p36-p34), COL11A1 (1p21), KCNK2 (1q41), S100A3 (1q21), ENAH (1q42.12), RGS1 (1q31), KCNH1 (1q32-q41), INSIG2 (2q14.1), FGF12 (3q28), TRIO (5p15.2), RNASEN (5p15.2), FGF10 (5p13-p12), EDN1(6p24.1-p22.3), SULF1 (8q13.2-13.3), TLR4 (9q32-q33), TNC (9q33), NTRK2 (9q22.1), CD44 (11p13), NCAM1 (11q23.1), TRIM29 (11q22-q23), PAK1 (11q13-q14) and RAB27A (15q15-q21.1), are found to be associated with cellular migration and proliferation, tumor cell metastasis and invasion, anchorage independent growth and inhibition of apoptosis. The candidate genes located at deleted regions of chromosomes, such as FBLN2 (3p25.1), WNT7A (3p25), DLC1 (8p22), LZTS1 (8p22), CDKN2A (9p21), COL4A1 (13q34), CDK8 (13q12) and DCC (18q21.3), are found to be associated with the suppression of tumor. The suggested candidate genes were mostly involved in potential signaling pathways such as focal adhesion (COL4A1), tight junction (CLDN10), MAPK signaling pathway (FGF12) and neuroactive ligand receptor interaction pathway (CCKAR). Expression of FGF12 and COL4A1 was validated by tissue microarray. These unique copy number alteration profiles should be taken into consideration when developing biomarkers for the early detection of ESCC in high-risk areas of India in association with tobacco and betel quid use.

Polymorphisms of glutathione-S-transferase genes and the risk of aerodigestive tract cancers in the Northeast Indian population.

BACKGROUND Widespread use of tobacco and betel quid consumption and a high incidence of tobacco-associated aerodigestive tract cancers have been reported in different ethnic groups from several regions of Northeast (NE) India. This study was done to explore the possibility of phase II metabolic enzymes being responsible for the high prevalence of cancers in this region of India. METHODS Samples from 370 cases with oral, gastric, and lung cancers and 270 controls were analyzed for polymorphism of glutathione-S-transferase (GST) genes using polymerase chain reaction-restriction fragment length polymorphism-based methods. RESULTS AND CONCLUSIONS Tobacco smoking and betel quid chewing were found to be high risk factors for oral and lung cancers but not for gastric cancer, whereas tobacco chewing was found to be a risk factor for oral cancer but not for gastric or lung cancer. The variant genotypes of GSTP1 were not associated with any of the aerodigestive tract cancers. GSTT1 and GSTM1 null genotypes appeared to play a protective role for lung cancer (odds ratio [OR] = 0.47, 95% confidence interval [95% CI]: 0.24-0.93, p = 0.03) and (OR = 0.52, 95% CI: 0.28-0.96, p = 0.04), but they were not associated with oral and gastric cancers. However, when data was analyzed in different geographic regions the GSTT1 null genotype was found to be a significant risk factor for oral (OR = 2.58, 95% CI 1.01-6.61, p = 0.05) as well as gastric cancer (OR = 3.08, 95% CI 1.32-7.19, p = 0.009) in samples obtained from the Assam region of NE India. This is the first study on the association of GST polymorphisms and aerodigestive tract cancers in the high-risk region of NE India.

Investigation on the Role of p53 Codon 72 Polymorphism and Interactions with Tobacco, Betel Quid, and Alcohol in Susceptibility to Cancers in a High-Risk Population from North East India

The association of TP53 codon 72 polymorphism with cancer susceptibility remains uncertain and varies with ethnicity. Northeast India represents a geographically, culturally, and ethnically isolated population. The area reports high rate of tobacco usage in a variety of ways of consumption, compared with the rest of Indian population. A total of 411 cancer patients (161 lung, 134 gastric, and 116 oral) and 282 normal controls from the ethnic population were analyzed for p53 codon 72 polymorphism by polymerase chain reaction-restriction fragment length polymorphism. No significant difference in genotypic distribution of p53 between cases and controls was observed. Results suggested betel quid chewing as a major risk factor for all the three cancers (odds ratio [OR]=3.54, confidence interval [CI]=2.01-6.25, p<0.001; OR=1.74, CI=1.04-2.92, p=0.03; and OR=1.85, CI=1.02-3.33, p=0.04 for lung, gastric, and oral cancers, respectively). Tobacco smoking was associated with risk of lung and oral cancers (OR=1.88, CI=1.11-3.19, p=0.01 and OR=1.68, CI=1.00-2.81, p=0.04). Interactions between p53 genotypes and risk factors were analyzed to look for gene-environment interactions. Interaction of smoking and p53 genotype was significant only for oral cancer. Interactions of betel quid with p53 genotypes in lung cancer showed significant increase for all the three genotypes, indicating a major role of betel quid (OR=5.90, CI=1.67-20.81, p=0.006; OR=5.44, CI=1.67-17.75, p=0.005; and OR=5.84, CI=1.70-19.97, p=0.005 for Arg/Arg, Arg/Pro, and Pro/Pro, respectively). In conclusion, high incidence of these cancers in northeast India might be an outcome of risk habits; further, tissue- and carcinogen-specific risk modification by p53 gene is probable.

Expression of genes related to multiple drug resistance and apoptosis in acute leukemia: response to induction chemotherapy

Resistance to chemotherapy is a major impediment to the successful treatment of acute leukemia (AL). Expression of genes involved in drug resistance and apoptosis may be responsible for this. This study aimed to investigate the expression of drug resistance (MDR1, MRP1, LRP, BCRP, GSTP1, DHFR) and apoptotic genes (p53, BCL-2, Survivin) in adult acute leukemias and compare them with clinical and hematological findings and response to induction chemotherapy. Eighty-five patients with AL [45 with acute myeloid leukemia (AML) and 40 with acute lymphoblastic leukemia (ALL)] were used as a study group. Real-time PCR results showed that expression level of MDR1 was significantly higher in AML whereas expression of DHFR, BCRP and Survivin was significantly higher in ALL patients. In AML, significant correlation was observed between LRP and MRP1 (r(s)=0.44, p=0.016), LRP and DHFR (r(s)=0.41, p=0.02), MDR1 and BCL-2 (r(s)=0.38, p=0.03). Expression of GSTP1 and LRP correlated with high white blood count (p=0.03 and p=0.03) and BCL-2 with high peripheral blast count (p=0.009). MDR1 expression was significantly associated with the expression of immature stem cell marker CD34 (p=0.002). In ALL, significant association was found between LRP gene and female sex (p<0.0001), LRP and B-ALL patients (p=0.04) and LRP and BCR/ABL positive patients (p=0.004). High expression of MDR1 and BCL-2 in AML and MRP1 gene in ALL was associated with response to induction chemotherapy (p=0.001, p=0.02 and p=0.007 respectively). These results showed the potential clinical relevance of MDR1, MRP1 and BCL-2 in adult patients with acute leukemia in the context of induction chemotherapy.

Polymerase Chain Reaction for Detection of Endocervical Chlamydia trachomatis Infection in Women Attending a Gynecology Outpatient Department in India

OBJECTIVES To detect Chlamydia trachomatis infection by polymerase chain reaction (PCR) in symptomatic women attending a gynecology clinic in a city hospital and in randomly selected slum dwellers. STUDY DESIGN Endocervical specimens were collected from 350 women with genitourinary complaints (group I) and 53 slum dwellers (group II). Samples were analyzed by PCR, direct fluorescence assay (DFA) and Giemsa stain cytology for detection of C trachomatis and compared for their sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV). RESULTS The prevalence of endocervical C trachomatis infection was 43.1% and 24.5% in groups I and II, respectively. The sensitivity, specificity, PPV and NPV of PCR were 80.0%, 75.0%, 66.6% and 85.7%, respectively, when DFA was considered true positive. The percent increment in detection of C trachomatis by PCR was 15.3%. CONCLUSION Giemsa stain cytology has low sensitivity and specificity; hence, it cannot be recommended for use as a diagnostic technique. It appears that PCR can be used routinely in Chlamydia diagnosis and in screening selected populations. The high positivity of C trachomatis infection in urban slum dwellers is cause for concern.

Role of epoxide hydrolase 1 gene polymorphisms in esophageal cancer in a high-risk area in India

Background and Aim:  Microsomal epoxide hydrolase 1 (EPHX1) is involved in the metabolism of environmental and tobacco carcinogens. Tobacco smoking, betel quid chewing, and alcohol consumption are the major known risk factors for esophageal cancer. The present case‐control study evaluated the influence of EPHX1 genetic variations on esophageal cancer susceptibility in 142 patients and 185 healthy controls from a high‐incidence region of India where tobacco use and alcohol consumption are widespread and the users of these two substances are also betel quid chewers.

Association of glutathione S-transferase, EPHX, and p53 codon 72 gene polymorphisms with adult acute myeloid leukemia.

Polymorphisms in genes encoding detoxification enzymes have been suggested as susceptibility factors for many solid tumors. However, their association with hematological malignancies is controversial. A case-control study was done to determine the association between glutathione S-transferase M1 (GSTM1), GSTT1, GSTP1, EPHX1, and p53 codon 72 polymorphisms as risk factors in 120 adult acute myeloid leukemia (AML) cases and 202 healthy controls by polymerase chain reaction-restriction fragment length polymorphism techniques. Data were analyzed using χ(2) and conditional logistic regression model. None of the polymorphisms studied alone was associated with increased risk for AML. However, the frequency of GSTT1 null genotype was higher among controls (28.7%) than AML cases (21.6%), which showed a protective effect of the null genotype (odds ratio = 0.58, 95% confidence interval: 0.33-1.05, p = 0.07). In a combined analysis, both EPHX1 (His113His) and GSTP1 (Ile/Val) genes imparted a fourfold risk for adult AML but did not reach statistical significance (odds ratio = 4.22, 95% confidence interval: 0.992-17.99, p = 0.05). These findings suggest that the etiology of adult AML cannot be explained by polymorphism at a single locus, perhaps because of complexity involved in the metabolism of diverse xenobiotic compounds, and therefore, multiple gene-gene interactions should be investigated to predict the risk of AML.

Cytokeratin immunoexpression in esophageal squamous cell carcinoma of high-risk population in Northeast India.

Esophageal cancer is a frequently fatal malignancy, and is described in certain regions in Northeast India with an incidence of esophageal squamous cell carcinoma many fold higher than the rest of the population. The population in Northeast India is at higher risk due to poor nutritional status, consumption of fermented betel quid and other oral tobacco products besides smoking and alcohol intake. Cytokeratins (CKs) are the major constituents of the esophageal epithelium and may show gain or loss of CKs as the cancer progresses from normal epithelium to invasive phenotype. In this study, we studied the immunohistochemical expression of 5 CKs (CK4, CK5, CK8, CK14, and CK17) in the normal esophageal epithelium and esophageal squamous cell carcinoma from both the general population and the high-risk population of Assam in Northeast India. The CK expression profile was similar to other published data in general. Further analysis demonstrated differences in CK expression between the general and the high-risk tumor samples. CK5 and CK8 expression was altered in the high-risk population. The significance of these differences is unclear, but suggests a connection to the etiologic factors.

High order interactions of xenobiotic metabolizing genes and P53 codon 72 polymorphisms in acute leukemia.

Polymorphisms in xenobiotic metabolizing genes are associated with altered metabolism of carcinogens in acute leukemia (AL). This study applied two data mining approaches to explore potential interactions among P53 and xenobiotic metabolizing genes in 230 AL patients [131 acute myeloid leukemia (AML) and 99 acute lymphoblastic leukemia (ALL)] and 199 controls. Individually, none of the genotypes showed significant associations with AML risk. However, in ALL the CYP1A12A TC genotype was associated with increased risk (OR = 2.02; 95% CI = 1.14–3.58; P = 0.01), whereas the GSTM1 null genotype imparted reduced risk (OR = 0.55; 95% CI = 0.31–0.96; P = 0.03). In classification and regression tree analysis, combinations of GSTM1 present, CYP1A12C AA or GG, EPHX1 exon3 TC, and EPHX1 exon4 AA or GG genotype strongly enhanced the risk of AML (OR = 5.89; 95% CI = 1.40–26.62; P = 0.01). In ALL, combinations of CYP1A12A TT, P53 GG or CC and GSTP1 AG genotypes conferred the highest risk (OR = 4.19; 95% CI = 1.45–12.25; P = 0.004). In multifactor dimensionality reduction analysis, a four locus model (GSTP1, P53, EPHX1 exon3, and CYP1A12A) was the best predictor model for ALL risk. The association between this model and ALL risk remained true even at low prior probabilities of 0.01% (false positive report probability = 0.05). Interaction entropy interpretations of the best model of ALL revealed that two‐way interactions were mostly synergistic. These results suggest that high order gene–gene interactions play an important role in AL risk. Environ. Mol. Mutagen., 2012.