Association of Genetic Variants at the 4q25 Locus with Atrial Fibrillation in Indian Population.

INTRODUCTION: Atrial fibrillation (AF) is the most common cardiac arrhythmia in clinical settings known to impair cardiac function. Genome-wide association studies identified SNPs on chromosome 4q25 to be associated with AF. Till date no information is available from India related to the association of these variants with AF. MATERIALS AND METHODS: Two hundred and sixty-seven individuals comprising of 170 patients with Coronary Artery Disease (CAD), 41 patients with AF, and 56 healthy controls were genotyped for rs10033464 and rs2200733 at 4q25 locus. RESULTS: Strong association of rs10033464 with AF was observed on comparison with control groups (OR: 2.59; 95% CI: 1.08-6.21; P: 0.031) and between post-coronary artery bypass grafting (CABG) AF and control with dominant genetic model (OR: 4.73; 95% CI: 1.50-14.89; P: 0.0071). Comparison of post-CABG AF with CAD also indicated association (OR: 2.73; 95% CI: 0.9-7.56; P value: 0.05). In contrast, the rs2200733 C>T variant did not show any association with post-CABG AF, but with lone AF with the 'T' allele being associated with increased risk was seen (OR: 2.80; 95% CI: 1.08-7.24; P value: 0.042). CONCLUSION: In conclusion, the rs10033464 (T) allele is associated with the risk of post-CABG AF and the rs2200733 (T) with lone AF.

Hb F Levels in Indian Sickle Cell Patients and Association with the HBB Locus Variant rs10128556 (C>T), and the HBG XmnI (Arab-Indian) Variant.

The prevalence of sickle cell disease in India is very high. Hb F is one of the most powerful modulators of disease severity in sickle cell disease patients. It was traditionally thought that the disease is milder in Indian sickle cell disease patients predominantly due to the Arab-Indian haplotype characterized by the HBG XmnI [rs7482144 (G>A)] variant, which is associated with increased Hb F levels. In the current study, we investigated the Hb F levels in individuals with the rs10128556 (C>T) variant and also determined its linkage with the HBG XmnI variant. The present study was conducted on a cohort of 275 individuals, which consisted of 221 patients with sickle cell disease and 54 patients with sickle cell trait. Analysis of hemoglobin (Hb) fractions and variants was done on the high performance liquid chromatography (HPLC) system. Genotyping for rs10128556 was done by direct sequencing of the products. Mean Hb F levels in the sickle cell disease patients was 19.36 ± 6.79. The genotypic frequencies for rs10128556 were 82.0% (TT), 16.7% (CT) and 1.3% (CC) for sickle cell disease patients. The minor C allele resulted in 52.0% decrease in Hb F levels when homozygous and 7.0% decrease when heterozygous. The rs10128556 single nucleotide polymorphism (SNP) was in strong but not complete linkage with the HBG XmnI variant. In conclusion, the study determined for the first time the frequency and association of rs10128556 in Indian sickle cell disease patients with Hb F. It also established that it was not in complete linkage with the HBG XmnI variant in this high risk population.

Variant at 9p21 rs1333049 is associated with age of onset of coronary artery disease in a Western Indian population: a case control association study.

The 9p21 chromosomal region has been associated with coronary artery disease (CAD) in many genome wide association studies (GWAS). To date no information exists regarding the rs1333039 SNP which showed the strongest association in the WTCCC GWAS with CAD risk in the Indian population. The present study attempts to replicate the findings in the Indian population. Genotyping for rs1333049 was done in 229 cases and 151 controls by allele-specific real-time assay. A higher frequency of the risk allele rs1333049C was seen in cases (0·60) as compared with controls (0·49), which associated with CAD risk both in univariate (OR = 1·564, 95%CI = 1·154-2·119, P = 0·003) and multivariate analysis (OR = 2·460, 95%CI = 1·139-5·314, P = 0·022). Increased frequency of the risk allele was seen in younger individuals with CAD where 40% individuals in the age group 30-55 years had the CC genotype as compared with 29 and 24·5% in the age group 56-65 years and > 65 years, respectively (CC versus GG, P = 0·045). Higher incidence of the CC genotype was seen in MI patients, but missed significance when compared with controls (OR = 1·361, 95%CI = 0·954-1·942, P = 0·084). In conclusion, the rs1333049 variant is significantly associated with CAD risk and also with age of onset in the Western Indian population. However there are differences in the haplotype structure of this SNP with the neighbouring rs10757278 SNP, these differences emphasize the importance of genotyping all risk variants at this locus which could underlie the differences in risk susceptibility to CAD across populations.

Molecular analysis of genetic variation in angiotensin I-converting enzyme identifies no association with sporting ability: First report from Indian population.

INTRODUCTION: A polymorphism in the angiotensin-converting enzyme (ACE) gene was the first performance enhancing polymorphisms (PEPs) to be identified and correlated with athletic abilities. This polymorphism (rs. 5186) is the absence (deletion; D allele), rather than the presence (insertion, I allele) of 287bp Alu repeat element in intron 16. However, the association of ACE I/D polymorphism in sports abilities have been contradicted and debated. No study has evaluated the ACE gene polymorphism in Indian athletes so far. Hence, the genotype distribution and allelic frequency of ACE gene in selected Indian athletic and non-athletic population was studied. MATERIALS AND METHODS: A total of 147 athletes and 131 controls were genotyped for the ACE gene polymorphism using PCR. RESULTS: No significant association was observed between the allelic frequencies of ACE gene in controls and athletes on a whole, as well as after sub-categorizing the athletes based on the type of sport they played (P > 0.1). However, a higher representation of I allele was observed in the athletes. CONCLUSION: ACE genotyping studies need to focus on truly elite athletes of a single sporting discipline, to be able to find an association. The ACE I/D polymorphism may not be considered a marker for human performance, but can be further studied in combination with other potent performance enhancing polymorphisms.

Mutations in the HD and PEST domain of Notch-1 receptor in T-cell acute lymphoblastic leukemia: report of novel mutations from Indian population.

Notch-1 is a transmembrane receptor protein that directs T-cell differentiation. Gain-of-function mutations in Notch-1 have been reported in more than 50% of human T-cell acute lymphoblastic leukemia (T-ALL). The current study was undertaken to characterize mutations in the heterodimerization (HD) domain and proline, glutamic acid, serine, threonine-rich (PEST) domain of the Notch-1 receptor. RNA was isolated from peripheral blood/bone marrow of 15 de novo T-ALL subjects; the Notch-1 HD and PEST regions were amplified and sequenced. Overall six patients (40%) had at least one Notch-1 mutation, 2/15 (13%) in the HD and 4/15 (27%) in the PEST domain. None of the samples showed simultaneous mutations in HD and PEST domains. Mutations were seen in 4/10 adult patients (40%); in the pediatric cohort 2/5 (40%) had mutations both of which were in the PEST domain. Of the different mutations, two have been previously reported and the other four are novel. A high incidence of Notch-1 mutations has been seen; unlike other studies, a higher frequency of mutations was found in PEST domain. The current study also served to identify four novel mutants that add new insights into the genetic heterogeneity of T-ALL. More ongoing larger studies are warranted to elucidate the molecular pathogenesis of T-ALL that arises in this part of the world.

Spinocerebellar Ataxias in India: Three­year Molecular Data from a Central Reference Laboratory.

INTRODUCTION: There is a great deal of heterogeneity, both phenotypically and genotypically among the autosomal dominant cerebellar ataxias (ADCA). Their prevalence also varies in different populations. Trinucleotide repeat expansions (CTG/CAG) have been shown predominantly to cause a number of ADCAs. AIM: The present study describes the frequency of spinocerebellar ataxias (SCA) and the CAG repeat sizes among the different regions of India. SETTINGS AND DESIGN: Molecular data from our central reference laboratory were retrospectively analyzed for SCAs 1, 2, 3, 6, 7, 10, 12, 17 and DRPLA. Correlation between age at diagnosis and the CAG repeats of the expanded and the normal alleles were tested with the Spearman correlation test. RESULTS: The presence of SCAs vary according to geographical regions and ethnicities; SCA 12 was detected with the highest frequency (229/901), but was restricted to a specific ethnic population, followed by SCA 2 with a positivity of 12% (101/845). SCA 3 previously known as Machado-Joseph Disease had a prevalence of 4.05% (32/789), whereas SCA 1 was diagnosed in 30/773 (3.88%). No positivity was seen for SCA 10 from the 103 samples tested and for SCA 17 from the 131 samples tested either as a part of an extended panel or stand-alone. CONCLUSION: In this report, we are able to expand the portrait of SCAs in India by presenting the largest ever molecular data from a central reference laboratory.

Frequency of fokI and taqI polymorphism of vitamin D receptor gene in Indian population and its association with 25-hydroxyvitamin D levels.

BACKGROUND: The VDR protein is at the centre of the vitamin D endocrine system, a complex physiological system with substantial feedback regulatory mechanisms involved in maintaining serum calcium and 1, 25 dihydroxy vitamin D3. Variations in VDR gene are shown to have implications in several diseases and have also been implicated as an important genetic factor affecting bone mass. AIM: To determine the frequency of Fok I and Taq I variants in healthy Indian individuals and its association with 25-OH-Vitamin D levels. SETTINGS AND DESIGN: Blood samples were collected from 143 unrelated normal individuals (Male-84 and Female-59) and their genotypes determined. MATERIALS AND METHODS: After amplification by polymerase chain reaction, each polymorphism was genotyped by restriction fragment length polymorphism. For 100 normal healthy individuals 25-hydroxyvitamin D estimation was done using DiaSorin kit method. STATISTICAL ANALYSIS: Graph pad software was used to calculate the P values from the Chi-square. RESULTS: Out of 143 samples analyzed for FokI and TaqI polymorphisms the following genotypic frequency was obtained FF 59%, Ff 36%, ff 5% and TT 49%, Tt 43%, tt 8% respectively. CONCLUSIONS: Results indicate that the distribution of the polymorphic loci Fok I and Taq I vary considerably not only in different populations, but also within India. Furthermore, when the genotypes were analyzed with respect to 25-OH-Vitamin D levels, a significant association was seen for the Taq 1 SNP but not with the Fok I.

Reference ranges for lymphocyte subsets in adults from western India: influence of sex, age and method of enumeration.

BACKGROUND: The enumeration of absolute CD4 counts is of primary importance, since therapeutic protocols for HIV1 patients are based on these. AIMS: To establish reference ranges for the CD4 and CD8 T-lymphocytes in the Indian population. SETTINGS AND DESIGN: Enumeration of absolute numbers and percentages of lymphocyte subsets was performed in 252 healthy adult Indians. METHODS AND MATERIALS: The assays for SPT were carried out using the Beckman EPICS XL-MCL flow cytometer and the cytostat tetraCHROME reagent containing CD45/CD8/CD4/CD3 monoclonal antibodies. For comparison with DPT the absolute lymphocyte count was obtained using the Coulter STK-S fully automated hematology analyzer. STATISTICAL ANALYSIS: Regression analysis and Students t test were used for data analysis. RESULTS: Median values were as follows; absolute CD3 counts 1446 cells/mm3 (total), 1361 cells/mm3 (males) and 1511 cells/mm3 (females); absolute CD4 counts are 771 cells/mm3 (total), 705 cells/mm3 (males) and 839 cells/mm3 (females); absolute CD8 counts are 555 cells/mm3 (total), 552 cells/mm3 (males) and 561 cells/mm3 (females). The median CD4/CD8 ratio for the total samples was 1.34, for males 1.22 and for females 1.49. CONCLUSIONS: In this study we have established reference ranges for normal Indian adults using the fully automated Single Platform Technology. The lymphocyte subsets values of our population are closer to those of the population from Botswana and China rather than the Western population. The absolute CD3 and CD4 counts and the CD4:CD8 ratio are higher in females than in males. Consistently higher values are obtained by the DPT as compared to the SPT.

Frequency and association of disabled homolog 2-interacting protein (DAB2IP) variant rs7025486 G>A with coronary artery disease risk in Indian population.

Genome wide association study has identified rs7025486 G>A polymorphism within DAB2IP (Disabled homolog 2-interacting protein) gene with increased risk of coronary heart disease (CAD). In this study we have determined the frequency and association of rs7025486 with CAD in Indians. The study was performed on 214 patients with CAD and 125 controls. The 'AA' genotype was associated with an increased risk in the CAD age group <50 yrs as compared to CAD age group>50 yrs (OR 3.149; P 0.034) and controls >50 yrs (OR 3.430; P 0.080). The risk allele (A) was significantly associated with premature CAD.

Age, sex, and HIV seroprevalence among individuals from different parts of India tested for HIV infection in a nongovernmental setting.

India has a large population of HIV-infected individuals, but the amount of information available about this population is small and not commensurate with the population's size. Here, we report the age and sex for 11,925 individuals tested for HIV infection in a nongovernmental setting over a 36-month period. The samples were derived from 161 sites located in different parts of the country and the odds of HIV infection among males was 2.27 times that for females. Of the samples from males and females tested, 50 and 65%, respectively, were in the 16-35 year age group. Of the seropositive samples excluding less than 1 year of age, 92% were in the 16-50 year age group. Analysis of this testing data provides limited but valuable information on the HIV epidemic in India.

Homozygous c.1160C>T (P38L) in the MECP2 gene in a female Rett syndrome patient.

Rett syndrome is a severe X-linked dominant neurodevelopmental disorder. Mutations in the MECP2 gene on chromosome Xq28 have been shown to be the cause of Rett syndrome. Sequencing of the MECP2 gene in a patient with clinical suspicion of Rett syndrome revealed c.1160C>T (P387L) in exon 4 of the MECP2 gene homozygously. Females with Rett syndrome are usually heterozygous for a mutation in MECP2. Uniparental disomy as a probable cause for the homozygous presence of this mutation was ruled out by quantitative fluorescence-polymerase chain reaction. Moreover to our knowledge this mutation has only been reported in males with X-linked mental retardation (MRX). We hypothesize that the presence of this mutation c.1160C>T (P387L) in the homozygous form is responsible for the Rett syndrome-like phenotype seen in this patient. This novel report reveals for the first time the homozygous presence of a mutation which has hitherto only been reported in males with MRX.

Trends in antibiotic resistance among major bacterial pathogens isolated from blood cultures tested at a large private laboratory network in India, 2008-2014.

OBJECTIVE: There have been no long-term studies on trends in antibiotic resistance (ABR) on a national scale in India. Using a private laboratory network, the ABR patterns of organisms most commonly associated with bacteremia, obtained from patients across India between 2008 and 2014, were examined. METHODS: A retrospective study of patient blood cultures collected over a 7-year period (January 1, 2008-December 31, 2014) was conducted. Data on the microorganism(s) identified and their antimicrobial susceptibility were obtained from SRL Diagnostics (Mumbai, India). RESULTS: Of 135268 blood cultures, 18695 (14%) had at least one identified pathogen. In addition to continual high rates of methicillin-resistant Staphylococcus aureus (MRSA; approximately 44.2%), high resistance to nalidixic acid among Salmonella Typhi (98%) was observed, and carbapenem resistance increased in both Escherichia coli (7.8% to 11.5%; p=0.332) and Klebsiella pneumoniae (41.5% to 56.6%; p<0.001). Carbapenem resistance was also stable and high for both Acinetobacter species (approximately 69.6%) and Pseudomonas aeruginosa (approximately 49%). Resistance was also detected to colistin in the Gram-negatives and to vancomycin and linezolid in S. aureus. CONCLUSION: Increasing resistance to antibiotics of last-resort, particularly among Gram-negatives, suggests an urgent need for new antibiotics and improved antimicrobial stewardship programs in India.

Cartilage oligomeric matrix protein in monitoring and prognostication of osteoarthritis and its utility in drug development.

Osteoarthritis (OA) is a major public concern as it is one of the leading causes of morbidity and lays a huge medical and economic burden on health resources. Early detection of OA has been a clinical challenge as early signs of joint inflammation are often not evidently identifiable on routine radiographic images. This presents a dire unmet medical need for a biomarker, which could detect early signs of joint inflammation much before irreversible joint damage and radiographic changes set in. Besides, the treatment of OA has remained mainly symptomatic. A disease modifying OA drug (DMOAD), which can act as targeted anti-OA therapy has not been able to receive regulatory approval yet. The clinical development of a DMAOD too warrants the need of a biomarker; which can act as a surrogate clinical endpoint used to monitor therapeutic efficacy and to validate a clinically meaningful change within the restricted time frame of a clinical study. In this regard, the current review focuses on cartilage oligomeric matrix protein (COMP), a potential OA biomarker which has shown significant clinical promise as a tool for early detection, therapeutic monitoring, prognostication and drug development for OA. This brief review is pivoted around the findings of selected relevant publications from PubMed indexed journals.

Genetics of fetal hemoglobin in tribal Indian patients with sickle cell anemia.

India tops the list of countries with sickle cell disease (SCD) with an estimated 44,000 live births in 2010 and a prevalence of 10%-33%. In the present study, the first from India, we have investigated the effect of genetic variants in the BCL11A, the HMIP (HBS1L-MYB intergenic polymorphism) locus, in addition to the HBB locus, which are known to be associated with fetal hemoglobin (HbF) levels, a major modulator of the disease phenotype. The present study was conducted on 240 individuals with SCD and 60 with sickle cell trait. Genotyping was performed for the BCL11A rs11886868 and rs34211119; HMIP rs9399137, rs189600565, rs7776196, rs34778774, and rs53293029; HBG2 Xmn1 polymorphism rs7482144; and -68C > T HBD promoter polymorphism. All the 3 quantitative trait loci were associated with HbF levels in Indian patients with SCD. The highest difference was seen in the Xmn1 single-nucleotide polymorphism, which accounted for 11% of the trait variance, the BCL11A rs11886868 for 3.65%, whereas the HMIP rs9399137 for 3.8%. The present study indicates the BCL11A, HMIP, and ß-globin region to be associated with increased HbF levels in Indian patient. Further interrogation of these genotypes with respect to pain crisis is warranted in this population, which may help in prognostication, as also a genome-wide association study, which may help uncover new loci controlling HbF levels.

Oral cancer: reviewing the present understanding of its molecular mechanism and exploring the future directions for its effective management.

The present review aims to analyze the information available regarding the molecular mechanisms of Oral Carcinogenesis and explore the future directions where the field of Cancer Biology is venturing. Oncologists have excellently followed the proverb "Necessity is the mother of Invention". The desire to be more precise and comprehensive in their studies has led to the invention of some of the most innovative techniques like laser capture microdissection, comparative genomic hybridization, microarrays, and protein chips etc. Various Biotech companies and Cancer Institutes are on a hunt for anti-cancer drugs and molecular markers for cancers. These revolutionary approaches and the new breed of Oncologists have made the field very exciting and have generated the hope that finally the war against cancer would be won. In the end it is urged that the lead taken in other cancers like colon, breast, leukemia will be emulated in oral cancer. This is expected to provide a molecular blueprint for HNSCC, thus helping to identify suitable markers for the early detection of pre-neoplastic lesions, as well as novel targets for its pharmacological intervention.

Molecular characterization of complex chromosomal rearrangement: first report of novel t(7;12) (q11;q22) as part of a complex karyotype in de novo AML-M2 case.

The strong association of diagnostic karyotype with clinical outcome has made cytogenetics one of the most valuable diagnostic and prognostic tools for acute myeloid leukemia (AML) till today. Complex chromosomal findings are reported to be seen in nearly 10-15% of adult AMLs and are generally associated with poor outcome. In the current report, we present the results of hematologic, immunophenotypic, cytogenetic, chromosomal microarray and molecular analyses of a 60-year-old female patient diagnosed with AML-M2. Cytogenetic analysis revealed complex chromosomal findings involving seven different chromosomes. However, cytogenetic analyses were not able to precisely unveil all karyotypic changes, hence chromosomal microarray was used for further characterization. The most interesting observation was identification of a t(7;12) (q11;q22) as part of this complex karyotype. To the best of our knowledge, this is the first report of identification of novel t(7;12) (q11;q22) as part of a complex karyotype in de novo AML-M2.

Pedometer assessed physical activity in urban pubertal children: first report from India.

BACKGROUND: Inadequate physical activity is a risk factor for several lifestyle diseases. In the current study we have tried to evaluate the physical activity levels in urban Indian pubertal children as well as investigate the relationship between step counts and body composition. METHODS: A total of 1032 children aged 12 to 15 years wore pedometers for 2 weekdays and 2 weekend days, the final cohort included 910 subjects with 467 boys and 443 girls. RESULTS: Mean weekday steps were 11,062 ± 4741 for boys and 9619 ± 4144 for girls; weekend steps were 10,842 ± 5034 for boys and 9146 ± 5159 for girls, which were both significantly different. The weekend steps were consistently lower in both genders. Analysis of children not meeting a cut-off of 10,000 steps indicated that 45% of the boys aged 12; 54% aged 13; 43% to 48% aged 14 and 50% in the aged 15 did not meet the cut-off. In girls higher levels of inactivity were seen with 58% to 65% aged 12; 69% to 73% aged 13; 49% to 58% aged 14 and 50% to 100% in age-group 15 did not meet the cut-off on weekdays and weekends respectively. CONCLUSIONS: The high level of physical inactivity in the representative urban Indian children is a cause of grave concern and necessitates urgent intervention strategies to be formulated.

Combined mutational analysis of KRAS, NRAS and BRAF genes in Indian patients with colorectal carcinoma.

The epidermal growth factor receptor (EGFR) is an excellent candidate for targeted therapy in colorectal cancer. Recent studies have demonstrated that apart from wild-type KRAS, a wild-type BRAF and NRAS genotype is required for response to anti-EGFR therapy. This suggests that NRAS and BRAF genotype criteria should be used together with KRAS genotype to select patients who will likely benefit from anti-EGFR therapy. We investigated the prevalence of mutations in the KRAS, BRAF and NRAS genes and its correlation with demographic characteristics, tumor location and stage in 100 colorectal carcinoma patients from India. The frequency of KRAS, BRAF and NRAS mutations was found to be 23%, 17% and 2.0%, respectively. There was no significant difference in KRAS, NRAS and BRAF mutation with respect to gender, age, tumor location (colon vs rectum) and clinicopathological stage. In addition, we found a novel point variant (T20I) of unknown significance in NRAS exon 1 in addition to a KRAS codon 12 mutation in one of the rectal carcinoma patients. In the present study, combined evaluation of genetic biomarkers (KRAS, NRAS and BRAF) was able to classify 42% of colorectal cancer patients as likely non-responders to anti-EGFR therapy.

Balanced autosomal translocation and double Robertsonian translocation in cases of primary amenorrhea in an Indian population.

OBJECTIVE: To assess the frequency of balanced autosomal translocations in patients with primary amenorrhea in an Indian population. METHODS: Cytogenetic analysis was carried out among women referred from all parts of India for primary amenorrhea between 2002 and 2010. Clinical history and laboratory findings were taken into consideration to determine the diagnosis. G-banding with trypsin-Giemsa was performed to detect chromosome abnormalities. RESULTS: There were 15 balanced autosomal translocations in 1100 patients. Two novel translocations were identified: 1 with mosaic pattern of X chromosome monosomy and male karyotype, together with balanced autosomal translocation of chromosomes 11 and 20 in both cell lines; and 1 with double Robertsonian translocation of chromosomes 14 and 21. CONCLUSION: Autosomal genes have a crucial role in reproductive development. More candidate genes need to be recognized for appropriate genetic counseling and clinical management.

Variant on 9p21 is strongly associated with coronary artery disease but lacks association with myocardial infarction and disease severity in a population in Western India.

BACKGROUND AND AIMS: Coronary artery disease (CAD) is the leading cause of death worldwide, especially so in Indians. Recently, genome-wide studies have implicated SNPs in the 58 kb region of chromosome 9p21 to be associated with CAD. In the current study we evaluated the association of single nucleotide polymorphism (SNP) rs10757278 at the 9p21 locus with CAD in a population from Western India. METHODS: Genotyping for rs10757278 A/G was done by direct sequencing in 215 cases with confirmed CAD and 150 controls. RESULTS: A significantly higher frequency of the G allele was seen in cases as compared to controls (0.64 vs. 0.53). In the current study the G allele showed association with risk of CAD (OR 1.832 per G allele 95% 1.035-3.242, P 0.042; OR 2.452 GG vs. AA 95% 1.358-4.4431, P 0.004). Addition of the 9p21 allele to Framingham risk score (FRS) resulted in a shift of 17% of individuals from the low-risk category to the intermediate-low (>5-<10% 10-year risk) and 7% from intermediate-low to intermediate-high (>10-<20% 10-year risk) categories. CONCLUSIONS: The rs10757278 G variant at the 9p21 locus is significantly associated with the risk of CAD in our population of Western India, similar to the observed trend in other populations; however, the association is much stronger in the present cohort and, considering the high propensity of Indians to develop CAD, it is an important marker even in terms of risk classification.