Association analysis of serotonin transporter gene polymorphism among the South-Indian migraineurs.

Background: Migraine is a multifactorial neurological disorder characterized by frequent moderate to severe intensity headaches. The genetic variations in synaptic and post-receptor signalling proteins have direct effect on the process of serotonergic neurotransmission. Methods: We aimed to investigate the genetic association of serotonin transporter (SERT) 5-hydroxytryptamine transporter-linked promoter region (5-HTTLPR) polymorphism and migraine risk in South-Indian population. A total of 304 subjects with migraine including with aura (MA) and without aura (MO) and 308 controls were included in the present study. The single nucleotide polymorphism (SNP) was detected using polymerase chain reaction (PCR) and confirmed by deoxyribonucleic acid (DNA) sequencing. Results: The genotyping analysis revealed insignificant relationship with migraine subjects when compared with controls (P > 0.05). The minor 'S' allele showed no association with odds ratio (OR) = 1.23 [95% confidence interval (CI): 0.90-1.66], heterozygote with OR = 1.18 (95% CI: 0.82-1.69), and homozygote with OR = 1.51 (95% CI: 0.52-4.35). Conclusion: Further clinical studies are required to validate the results of SERT 5-HTTLPR promoter polymorphism in diverse ethnic descents especially in Asian populations.

2D-DIGE-Based Proteomic Profiling with Validations Identifies Vimentin as a Secretory Biomarker Useful for Early Detection and Poor Prognosis in Oral Cancers.

Oral tongue squamous cell carcinoma (OTSCC) is an aggressive cancer with high morbidity and mortality rates, despite multimodality management. There are currently no clinically relevant molecular markers that identify patients at higher risk of recurrence and failure. We undertook 2D-DIGE proteomic profiling to study the differentially expressed proteins in OTSCC evaluating their role in prognosis. 2D-DIGE coupled with tandem mass spectrometry was performed on tissues obtained from early staged OTSCC along with its paired apparently adjacent normal tissue samples (n = 10). Top upregulated protein was validated using immunohistochemistry (n = 345), comprising of retrospective early stage OTSCC (n = 150) and prospective series of oral precancers, normal, and oral cancers (n = 195). Saliva samples collected from oral cancer and precancer samples were analyzed by ELISA (n = 146). We found statistically significant differential expression in 151 proteins out of 700 proteins quantified. Top ten differentially regulated proteins were identified using mass spectrometry analysis. We found vimentin, the mesenchymal protein, to be the most upregulated protein in tongue tumor tissues compared to adjacent apparent normal tissues. Vimentin was found to be significantly overexpressed in oral precancers along with cancers compared to normal tissues. The vimentin expression correlated significantly with differentiated states of oral precancers and cancers. Vimentin was also detected at significantly higher levels in saliva collected from oral precancer and cancer patients compared to normal healthy volunteers. Validation of vimentin in an independent series of retrospective early staged OTSCC showed that the vimentin expression is significantly associated with treatment failures and poorer DFS. The vimentin expression is useful as both poor prognostic and early detection marker in oral cancer. Vimentin detection in saliva can be a diagnostic test to detect oral precancers that may have malignant potential, needing closer follow-up, and disease monitoring.

Exome Sequencing with Validations and Expression of p16/CDKN2A Shows no Association with HPV in Oral Cancers.

PURPOSE: Despite many studies attributing HPV infection to oropharyngeal tumorigenesis, its involvement in non-oropharyngeal cancers is ambiguous. We have evaluated the mutation profile of p16 along with protein expression and correlated it with the HPV status in oral cancers. METHODS: Somatic mutations in p16 were studied by exome sequencing (n=25) and validated by Sequenom Mass spectrometry (n=50).  Expression of p16 was studied by immunohistochemistry (IHC) and correlated with HPV16/18 status evaluated by PCR, and IHC (n=221) in oral cancers. RESULTS: Out of 25 oral cancer patients' samples sequenced by Exome sequencing, p16 mutations were found in 4 samples (16%). All the p16 mutations were identified in patients with cancers in the site of gingivobuccal complex and not tongue subsite. All the 4 patients with p16 mutations had failed treatment, and showed a significantly poor disease-free survival. Insilico analysis of the types of p16 mutations showed mutated, truncated p16 protein having an increased intrinsic disorder, and all the mutations involved truncation post arginine. Validation of the p16 mutations by mass spectrometry showed 8/50 (16%) of patients harbouring pArg80Ter mutation, of which 7/8 (87.5%) had failed treatment.  Overexpression of p16 in >70% of the tumour cells was found in 21.4% (26/121) OSCC patients, 6.75% (5/74) OPML patients and p16 expression was significantly correlated (p=0.001; χ2 = 25.601) to the grade. All the samples were studied for HPV presence by PCR and IHC. We found that none of the p16 positive tumours showing expression in >70% of the tumour cells harbored HPV both by PCR as well as IHC. CONCLUSION: Our study emphasises the importance of p16 in oral cancers, and shows that oral cancer is not HPV associated and p16 expression is not a surrogate marker for HPV.

Genetic predisposition of TNFα gene polymorphism in South-Indian Migraineurs and meta-analysis.

Migraine (Mg) is a multifaceted neurovascular disorder caused by genetic and several environmental etiologies. We have implemented a case-control study of TNFα gene polymorphism in 212 Mg patients and 218 healthy controls utilizing the ARMS-PCR technique, followed by Sanger sequencing. Besides, we have conducted a meta-analysis of different genetic models (five genetic models) to combine and summarize the available data from 11 studies (including this present research). The strength of genetic associations in the meta-analysis used to assess by the pooled odds ratio (OR) and 95% confidence intervals (CI). The results of this case-control study discovered a significant relationship with Mg in recessive and homozygous genotype with OR = 2.35 (95% CI [0.96-5.74]), p-value = 0.045. Also, the outcomes of meta-analysis suggested an irrelevant relationship between TNFα gene (rs1800629) polymorphism and Mg susceptibility in the five genetic models. However, subgrouping based on ethnic background showed a significant association in the allelic genetic model with OR = 1.53 (95% CI [1.02-2.31]), p = 0.040 respectively. The meta-analysis results of TNFα gene polymorphism may represent a risk factor for Mg among Asians. In the future, large scale, multicentric case-control study by classification of patients with Mg with or without aura can be performed worldwide to identify the potential genetic risk factors leading to Mg pathogenesis.