Cancer cell plasticity: from cellular, molecular, and genetic mechanisms to tumor heterogeneity and drug resistance.

Cancer is a complex disease displaying a variety of cell states and phenotypes. This diversity, known as cancer cell plasticity, confers cancer cells the ability to change in response to their environment, leading to increased tumor diversity and drug resistance. This review explores the intricate landscape of cancer cell plasticity, offering a deep dive into the cellular, molecular, and genetic mechanisms that underlie this phenomenon. Cancer cell plasticity is intertwined with processes such as epithelial-mesenchymal transition and the acquisition of stem cell-like features. These processes are pivotal in the development and progression of tumors, contributing to the multifaceted nature of cancer and the challenges associated with its treatment. Despite significant advancements in targeted therapies, cancer cell adaptability and subsequent therapy-induced resistance remain persistent obstacles in achieving consistent, successful cancer treatment outcomes. Our review delves into the array of mechanisms cancer cells exploit to maintain plasticity, including epigenetic modifications, alterations in signaling pathways, and environmental interactions. We discuss strategies to counteract cancer cell plasticity, such as targeting specific cellular pathways and employing combination therapies. These strategies promise to enhance the efficacy of cancer treatments and mitigate therapy resistance. In conclusion, this review offers a holistic, detailed exploration of cancer cell plasticity, aiming to bolster the understanding and approach toward tackling the challenges posed by tumor heterogeneity and drug resistance. As articulated in this review, the delineation of cellular, molecular, and genetic mechanisms underlying tumor heterogeneity and drug resistance seeks to contribute substantially to the progress in cancer therapeutics and the advancement of precision medicine, ultimately enhancing the prospects for effective cancer treatment and patient outcomes.

Associations between telomere attrition, genetic variants in telomere maintenance genes, and non-small cell lung cancer risk in the Jammu and Kashmir population of North India.

BACKGROUND: Telomeres are repetitive DNA sequences located at the ends of chromosomes, playing a vital role in maintaining chromosomal integrity and stability. Dysregulation of telomeres has been implicated in the development of various cancers, including non-small cell lung cancer (NSCLC), which is the most common type of lung cancer. Genetic variations within telomere maintenance genes may influence the risk of developing NSCLC. The present study aimed to evaluate the genetic associations of select variants within telomere maintenance genes in a population from Jammu and Kashmir, North India, and to investigate the relationship between telomere length and NSCLC risk. METHODS: We employed the cost-effective and high-throughput MassARRAY MALDI-TOF platform to assess the genetic associations of select variants within telomere maintenance genes in a population from Jammu and Kashmir, North India. Additionally, we used TaqMan genotyping to validate our results. Furthermore, we investigated telomere length variation and its relation to NSCLC risk in the same population using dual-labeled fluorescence-based qPCR. RESULTS: Our findings revealed significant associations of TERT rs10069690 and POT1 rs10228682 with NSCLC risk (adjusted p-values = 0.019 and 0.002, respectively), while TERF2 rs251796 and rs2975843 showed no significant associations. The TaqMan genotyping validation further substantiated the associations of TERT rs10069690 and rs2242652 with NSCLC risk (adjusted p-values = 0.02 and 0.003, respectively). Our results also demonstrated significantly shorter telomere lengths in NSCLC patients compared to controls (p = 0.0004). CONCLUSION: This study highlights the crucial interplay between genetic variation in telomere maintenance genes, telomere attrition, and NSCLC risk in the Jammu and Kashmir population of North India. Our findings suggest that TERT and POT1 gene variants, along with telomere length, may serve as potential biomarkers and therapeutic targets for NSCLC in this population. Further research is warranted to elucidate the underlying mechanisms and to explore the potential clinical applications of these findings.

Evaluation of 17 genetic variants in association with leukemia in the north Indian population using MassARRAY Sequenom.

Leukemia is a heterogeneous disorder, characterized by elevated proliferation of white blood cells. In this study, we explored the association of 17 genetic variants with leukemia patients in the Jammu and Kashmir region of north India. The variants were genotyped by using a high-throughput Agena MassARRAY platform in 758 individuals (166 cases and 592 controls). Of the 17 single-nucleotide polymorphisms (SNPs) studied, five SNPs were showing significant association with the high risk of leukemia in the north Indian population, which includes rs10069690 of telomere reverse transcriptase (TERT) with OR = 0.34 (95% CI, 0.20-0.58; p = .0008), rs2972392 (​​​PSCA) with OR 1.86 (95% CI, 1.04-3.81; p = .035), rs4986764 (BRIP1) with OR 1.34 (95% CI, 1.00-1.80; p = .04), rs6990097 (TNKS) with OR 1.81 (95% CI, 1.2-2.6; p = .001) and rs12190287 (TCF21) with OR 2.87 (95% CI, 1.72-4.7; p = .0001) by allelic association using Plink and analyzed by SPSS. This is the first study to explore these variants with leukemia in the studied population.

MassARRAY-based single nucleotide polymorphism analysis in breast cancer of north Indian population.

BACKGROUND: Breast Cancer (BC) is associated with inherited gene mutations. High throughput genotyping of BC samples has led to the identification and characterization of biomarkers for the diagnosis of BC. The most common genetic variants studied are SNPs (Single Nucleotide Polymorphisms) that determine susceptibility to an array of diseases thus serving as a potential tool for identifying the underlying causes of breast carcinogenesis. METHODS: SNP genotyping employing the Agena MassARRAY offers a robust, sensitive, cost-effective method to assess multiple SNPs and samples simultaneously. In this present study, we analyzed 15 SNPs of 14 genes in 550 samples (150 cases and 400 controls). We identified four SNPs of genes TCF21, SLC19A1, DCC, and ERCC1 showing significant association with BC in the population under study. RESULTS: The SNPs were rs12190287 (TCF21) having OR 1.713 (1.08-2.716 at 95% CI) p-value 0.022 (dominant), rs1051266 (SLC19A1) having OR 3.461 (2.136-5.609 at 95% CI) p-value 0.000000466 (dominant), rs2229080 (DCC) having OR 0.6867 (0.5123-0.9205 at 95% CI) p-value 0.0116 (allelic) and rs2298881 (ERCC1) having OR 0.669 (0.46-0.973 at 95% CI), p-value 0.035 (additive) respectively. The in-silico analysis was further used to fortify the above findings. CONCLUSION: It is further anticipated that the variants should be evaluated in other population groups that may aid in understanding the genetic complexity and bridge the missing heritability.

MassARRAY analysis of twelve cancer related SNPs in esophageal squamous cell carcinoma in J&K, India.

BACKGROUND: MassARRAY (Agena Bioscience™) combines competitive PCR with MALDI-TOF mass spectrometry (MS) analysis that gives highly accurate, sensitive, and high-throughput methods for the quantitative analysis of variation of gene expression in multiple samples. SNPs (Single Nucleotide Polymorphisms) have a very high potential of discovering disease-gene relationships. SNP-genotyping through MassARRAY is not only a cost-effective genotyping method but also provides a platform to validate variants observed through a high-throughput Next-generation sequencing (NGS). METHODS: In the present study, we have incorporated the use of matrix-assisted laser desorption/ionization-time of flight, mass spectrometry (MALDI-TOF) as a tool for differentiating genotypes based on the mass of variant. We have performed multiplex PCR and genotyped 12 SNPs in 758 samples (166 cases and 592 controls). The 12 studied SNPs were chosen with a rationale for their association with multiple cancers in literature. RESULTS: This is the first study to explore these SNPs with esophageal cancer within the J&K population. Out of 12 SNPs, two SNPs rs12190287 of TCF21 and rs10046 of CYP19A1 were significantly associated with esophageal cancer with Odds Ratio (OR) 1.412 (1.09-1.8 at 95% CI, p = 0.008) and 1.54 (1.21-2.072 at 95% CI, p = 0.0007) within the population of Jammu and Kashmir. CONCLUSION: We explored 12 SNPs that were found to be associated with multiple cancers in literature with esophageal cancer within the population of J&K. This is the first study to find the relation of these SNPs with ESCC within the studied population. This study explores the relation of genetic and environmental factors with the ESCC susceptibility.

Association of newly identified genetic variant rs2853677 of TERT with non-small cell lung cancer and leukemia in population of Jammu and Kashmir, India.

BACKGROUND: Telomere genetics has recently been emerged as an important field in molecular oncology. Various genome-wide association studies in different population groups have revealed that polymorphisms in Telomere maintenance gene (TERT) gene located on 5p15.33 is associated with susceptibility to leukemia and lung cancer risk. However, association of TERT with leukemia and lung cancer risk in north Indian population groups is still unknown. This study observed the association between genetic variant rs2853677 of TERT and leukemia and lung cancer in the state of Jammu and Kashmir, India. METHODS: A total of 781 subjects, out of which 381 cases (203 leukemic patients and 178 non-small cell lung cancer patients NSCLC) and 400 healthy controls were recruited for the study. Genetic variant rs2853677of TERT was detected using the real-time and Taqman Chemistry. Hardy-Weinberg Equilibrium was assessed using the chi square test. The allele and genotype- specific risks were estimated as odds ratio with 95% confidence interval. RESULTS: We observed that variant rs2853677 was strongly associated with lung cancer and leukemia risk with an odds ratio (OR) =1.8 (1.03-3.2 at 95% CI); p value (adjusted) = 0.03; odds ratio (OR) =2.9 (1.4-5.5.at 95% CI); p value (adjusted) = 0.002, respectively. CONCLUSION: The results of this study suggested that rs2853677 of TERT signifies association in multiple cancers and suggests that it can become potential marker for diagnosis of non-small cell lung cancer and leukemia. The study will provide an insight in understanding the genetic etiology and highlights the role of telomere-associated pathways in non-small cell lung cancer and leukemia. However, it would be quite interesting to explore the contribution of this variant in other cancers as well.

Genetic analysis of polymorphism rs10937405 of TP63 gene in breast and ovarian cancer patients of North Indian Cohort.

Introduction: Ovarian and breast cancers are highly prevalent in the population of Jammu and Kashmir (J&K). However, case-control association studies on breast and ovarian cancers are lacking in this population. Moreover, no case-control study is available on variant rs10937405 of TP63 in breast and ovarian cancers. Thus, we designed to replicate the cancer susceptible variant rs10937405 of TP63 in ovarian and breast cancers in the population of J&K because the TP63 gene act as a tumor suppressor gene and was previously associated with various cancers. Materials and Methods: This case-control association study conducted at the Shri Mata Vaishno Devi University, includes 150 breast, 150 ovarian cancer cases, and 210 healthy controls (age and sex-matched). Variant rs10937405 of the TP63 gene was determined by the TaqMan assay. Hardy-Weinberg equilibrium for the variant was assessed using the Chi-square test. The allele and genotype-specific risks were estimated by odds ratios (ORs) with 95% confidence intervals (CI). Results: In this study, variant rs10937405 of TP63 gene did not show any risk with ovarian and breast cancer with (P-value = 0.70) having OR 0.94, (0.69-1.28 at 95% CI) and (P-value = 0.16) having OR 0.80, (0.59-1.10). Discussion: Our results indicate that the variant rs10937405 of the TP63 gene did not impart any risk of breast and ovarian cancer in the population of J&K. Our results indicate that a larger sample size is needed for further statistical validation. As the study was for a particular variant, it warrants the analysis of other variants of this gene.

Innovative in Silico Approaches for Characterization of Genes and Proteins.

Bioinformatics is an amalgamation of biology, mathematics and computer science. It is a science which gathers the information from biology in terms of molecules and applies the informatic techniques to the gathered information for understanding and organizing the data in a useful manner. With the help of bioinformatics, the experimental data generated is stored in several databases available online like nucleotide database, protein databases, GENBANK and others. The data stored in these databases is used as reference for experimental evaluation and validation. Till now several online tools have been developed to analyze the genomic, transcriptomic, proteomics, epigenomics and metabolomics data. Some of them include Human Splicing Finder (HSF), Exonic Splicing Enhancer Mutation taster, and others. A number of SNPs are observed in the non-coding, intronic regions and play a role in the regulation of genes, which may or may not directly impose an effect on the protein expression. Many mutations are thought to influence the splicing mechanism by affecting the existing splice sites or creating a new sites. To predict the effect of mutation (SNP) on splicing mechanism/signal, HSF was developed. Thus, the tool is helpful in predicting the effect of mutations on splicing signals and can provide data even for better understanding of the intronic mutations that can be further validated experimentally. Additionally, rapid advancement in proteomics have steered researchers to organize the study of protein structure, function, relationships, and dynamics in space and time. Thus the effective integration of all of these technological interventions will eventually lead to steering up of next-generation systems biology, which will provide valuable biological insights in the field of research, diagnostic, therapeutic and development of personalized medicine.

Mutational Assessment in NKX2-5 and ACTC1 Genes in Patients with Congenital Cardiac Septal Defect (CCSD) from Ethnic Kashmiri Population.

(1) Background globe. The etiology of CHDs is complex and involves both genetic and non-genetic factors. Although, significant progress has been made in deciphering the genetic components involved in CHDs, recent reports have revealed that mutations in Nk2 homeobox5 (NKX2-5) and actin alpha cardiac muscle1 (ACTC1) genes play a key role in CHDs such as atrial and ventricular septum defects. Therefore, the present study evaluates the role of key hotspot mutations in NKX2-5 and ACTC1 genes of congenital cardiac septal defect (CCSD) in ethnic Kashmiri population. (2) Methods: A total of 112 confirmed CHD patients were included in the current study, of which 30 patients were evaluated for mutational analysis for hotspot mutations of NKX2-5 and ACTC1 genes. The total genomic DNA was extracted from the samples (cardiac tissue/blood) and were subjected to amplification for NKX2-5 (exon 1 and 2), and ACTC1 (exon 2) genes by using PCR specific primers to analyze the hotspot mutations in respective exons. The amplified products obtained were sent to Macrogen Korea for sequencing by Sanger's method. (3) Results: Our results confirmed that not a single mutation was found in either hotspot exon 1 and 2 of NKX2-5 and exon 2 of ACTC1 in the patients included in the current study. Interestingly, a novel synonymous nucleotide variation leading to G > C transversion (GCG > GCC) was found in exon 2 of NKX2-5 gene of CCSD patient. (4) Conclusions: The current findings demonstrated the role of NKX2-5 and ACTC1 in cardiac development. The study will provide an insight in understanding the genetic etiology and highlights the role of newly identified mutations in patients with CDS's in ethnic Kashmiri population. In silico findings revealed amino acid changes, splice site variation and the creation of new site. Furthermore, the study warrants complete screening of genes involved in CCSDs.

Transforming Growth Factor-Beta (TGF-ß) Signaling in Cancer-A Betrayal Within.

A ubiquitously expressed cytokine, transforming growth factor-beta (TGF-ß) plays a significant role in various ongoing cellular mechanisms. The gain or loss-of-function of TGF-ß and its downstream mediators could lead to a plethora of diseases includes tumorigenesis. Specifically, at the early onset of malignancy TGF-ß act as tumour suppressor and plays a key role in clearing malignant cells by reducing the cellular proliferation and differentiation thus triggers the process of apoptosis. Subsequently, TGF-ß at an advanced stage of malignancy promotes tumorigenesis by augmenting cellular transformation, epithelial-mesenchymal-transition invasion, and metastasis. Besides playing the dual roles, depending upon the stage of malignancy, TGF-ß also regulates cell fate through immune and stroma components. This oscillatory role of TGF-ß to fight against cancer or act as a traitor to collaborate and crosstalk with other tumorigenic signaling pathways and its betrayal within the cell depends upon the cellular context. Therefore, the current review highlights and understands the dual role of TGF-ß under different cellular conditions and its crosstalk with other signaling pathways in modulating cell fate.

Telomere Attrition With Concomitant hTERT Overexpression Involved in the Progression of Gastric Cancer May Have Prognostic and Clinical Implications in High-Risk Population Group From North India.

Genetic instabilities exacerbated by the dysfunction of telomeres can lead to the development of cancer. Nearly 90% of all human malignancies are linked with telomere dysregulation and overexpression of telomerase, an enzyme that catalyzes the synthesis of telomeric DNA repeats at the ends of chromosomes. The burden of gastric cancer continues to inflict a deterring impact on the global health scenario, accounting for over one million new cases in 2020. The disease is asymptomatic in its early stages of progression, which is attributed to the poor prognosis and overall surge in mortality rate worldwide. Exploiting telomere physiology can provide extensive mechanistic insight into telomere-associated gastric cancer progression and its use as a target in a variety of therapeutic interventions. In this study, we aimed to evaluate the clinical implications of c-Myc, human telomerase reverse transcriptase (hTERT) expression, and telomere length in patients with gastric cancer. A total of 57 gastric cancer cases and adjacent controls were included in the study. RT-PCR and immunohistochemistry were used to assess the expression levels of c-Myc and hTERT. The relative telomere length was measured by MMQPCR using the Cawthon method. Our results indicated that the shorter telomere and increased hTERT expression were associated with gastric cancer progression. The study also highlighted the role of short telomeres and increased expression of hTERT in gastric cancer progression and its association with various etiological risk factors, transcriptional activators, and overall survival among the ethnic Kashmiri population of North India.

Diagnostic implication of a circulating serum-based three-microRNA signature in hepatocellular carcinoma.

Owing to the diagnostic dilemma, the prognosis of hepatocellular carcinoma (HCC) remains impoverished, contributing to the globally high mortality rate. Currently, HCC diagnosis depends on the combination of imaging modalities and the measurement of serum alpha-fetoprotein (AFP) levels. Nevertheless, these conventional modalities exhibit poor performance in detecting HCC at early stages. Thus, there is a pressing need to identify novel circulating biomarkers to promote diagnostic accuracy and surveillance. Circulating miRNAs are emerging as promising diagnostic tools in screening various cancers, including HCC. However, because of heterogenous and, at times, contradictory reports, the universality of miRNAs in clinical settings remains elusive. Consequently, we proposed to explore the diagnostic potential of ten miRNAs selected on a candidate-based approach in HCC diagnosis. The expression of ten candidate miRNAs (Let-7a, miR-15a, miR-26a, miR-124, miR-126, miR-155, miR-219, miR-221, miR-222, and miR-340) was investigated in serum and tissue of 66 subjects, including 33 HCC patients and 33 healthy controls (HC), by rt-PCR. Receiver operating characteristic curve (ROC) analysis was used to determine the diagnostic accuracy of the prospective serum miRNA panel. To anticipate the potential biological roles of a three-miRNA signature, the target genes were evaluated using the Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathway. The serum and tissue expression of miRNAs (Let-7a, miR-26a, miR-124, miR-155, miR-221, miR-222, and miR-340) were differentially expressed in HCC patients (p < 0.05). The ROC analysis revealed promising diagnostic performance of Let-7a (AUC = 0.801), miR-221 (AUC = 0.786), and miR-2 (AUC = 0.758) in discriminating HCC from HC. Furthermore, in a logistic regression equation, we identified a three-miRNA panel (Let-7a, miR-221, and miR-222; AUC = 0.932) with improved diagnostic efficiency in differentiating HCC from HC. Remarkably, the combination of AFP and a three-miRNA panel offered a higher accuracy of HCC diagnosis (AUC = 0.961) than AFP alone. The functional enrichment analysis demonstrated that target genes may contribute to pathways associated with HCC and cell-cycle regulation, indicating possible crosstalk of miRNAs with HCC development. To conclude, the combined classifier of a three-miRNA panel and AFP could be indispensable circulating biomarkers for HCC diagnosis. Furthermore, targeting predicted genes may provide new therapeutic clues for the treatment of aggressive HCC.

Evaluation of the Cytotoxic, Anti-Inflammatory, and Immunomodulatory Effects of Withaferin A (WA) against Lipopolysaccharide (LPS)-Induced Inflammation in Immune Cells Derived from BALB/c Mice.

(1) Background: Inflammation is one of the primary responses of the immune system and plays a key role in the pathophysiology of various diseases. Recent reports suggest that various phytochemicals exhibit promising anti-inflammatory and immunomodulation activities with relatively few undesirable effects, thus offering a viable option to deal with inflammation and associated diseases. The current study evaluates the anti-inflammatory and immunomodulatory effects of withaferin A (WA) in immune cells extracted from BALB/c mice. (2) Methods: MTT assays were performed to assess the cell viability of splenocytes and anti-inflammatory doses of WA. Under aseptic conditions, the isolation of macrophages and splenocytes from BALB/c mice was performed to investigate the anti-inflammatory effects of WA. Analysis of the expression of proinflammatory cytokines and associated signaling mediators was performed using proinflammatory assay kits, real-time polymerase chain reaction (RT-PCR), and immunoblotting, while the quantification of B and T cells was performed by flow cytometry. (3) Results: Our results demonstrated that WA exhibits anti-inflammatory and immunomodulatory effects in LPS-stimulated macrophages and splenocytes derived from BALB/c mice, respectively. Mechanistically, we found that WA promotes an anti-inflammatory effect on LPS-stimulated macrophages by attenuating the secretion and expression of proinflammatory cytokines TNF-α, IL-1ß, IL-6, and the inflammation modulator NO, both at the transcriptional and translational level, respectively. Further, WA inhibits LPS-stimulated inflammatory signaling by dephosphorylation of p-Akt-Ser473 and p-ERK1/2. This dephosphorylation does not allow IĸB-kinase activation to disrupt IĸB-NF-ĸB interaction. The consistent interaction of IĸB with NF-ĸB in WA-treated cells attenuates the activation of downstream inflammatory signaling mediators Cox-2 and iNOS expression, which play crucial roles in inflammatory signaling. Additionally, we observed significant immunomodulation of LPS-stimulated spleen-derived lymphocytes by suppression of B (CD19) and T (CD4+/CD8+) cell populations after treatment with WA. (4) Conclusion: WA exhibits anti-inflammatory and immunomodulatory activity by modulating Akt/ERK/NF-kB-mediated inflammatory signaling in macrophages and immunosuppression of B (CD19) and T cell (CD4+/CD8+) populations in splenocytes after LPS stimulation. These results suggest that WA could act as a potential anti-inflammatory/immunomodulatory molecule and support its use in the field of immunopharmacology to modulate immune system cells.

JAK/STAT Signaling: Molecular Targets, Therapeutic Opportunities, and Limitations of Targeted Inhibitions in Solid Malignancies.

JAK/STAT signaling pathway is one of the important regulatory signaling cascades for the myriad of cellular processes initiated by various types of ligands such as growth factors, hormones, and cytokines. The physiological processes regulated by JAK/STAT signaling are immune regulation, cell proliferation, cell survival, apoptosis and hematopoiesis of myeloid and non-myeloid cells. Dysregulation of JAK/STAT signaling is reported in various immunological disorders, hematological and other solid malignancies through various oncogenic activation mutations in receptors, downstream mediators, and associated transcriptional factors such as STATs. STATs typically have a dual role when explored in the context of cancer. While several members of the STAT family are involved in malignancies, however, a few members which include STAT3 and STAT5 are linked to tumor initiation and progression. Other STAT members such as STAT1 and STAT2 are pivotal for antitumor defense and maintenance of an effective and long-term immune response through evolutionarily conserved programs. The effects of JAK/STAT signaling and the persistent activation of STATs in tumor cell survival; proliferation and invasion have made the JAK/STAT pathway an ideal target for drug development and cancer therapy. Therefore, understanding the intricate JAK/STAT signaling in the pathogenesis of solid malignancies needs extensive research. A better understanding of the functionally redundant roles of JAKs and STATs may provide a rationale for improving existing cancer therapies which have deleterious effects on normal cells and to identifying novel targets for therapeutic intervention in solid malignancies.

Head and neck cancer: Current challenges and future perspectives.

Head and neck cancers are a heterogeneous, aggressive and genetically complex collection of malignancies of the oral cavity, nasopharynx, oropharynx, hypopharynx, larynx, paranasal sinuses and salivary glands, which are difficult to treat. About 90% of all head and neck cancers are squamous cell carcinomas (HNSCC). Larynx and Oral cavity carcinomas are generally related with tobacco consumption, alcohol abuse (or both), but pharynx carcinomas are generally associated with infection of human papillomavirus (HPV), especially HPV-16 subtype. Thus, usually HNSCC can be separated into HPV-negative and HPV-positive categories. Despite substantial efforts invested into therapeutic development of HNSCC, the 5-year survival rate of patients with HNSCC still remains dismal. The primary reason being late diagnosis, recurrent metastasis, relapse and resistance to therapies. Currently surgery and radiotherapy represent the baseline treatment options for most initial stage HNSCC patients, but these treatments are associated with significant morbidity and poor prognosis. Moreover, the issue of resistance to both radiotherapy/chemotherapy and recurrent relapse are common in HNSCC. Elucidation of the genetic landscape, tumor microenvironment and aberrant signaling pathways have generated new insights into the molecular pathogenesis of this disease. Thus, the scientific research has therefore been focused on the understanding of HNSCC biology and immunobiology to identification of predictive/prognostic biomarkers, which will be key to develop more effective targeted therapies with less toxicity and high specificity.

Dual labeled fluorescence probe based qPCR assay to measure the telomere length.

Telomeres are highly repetitive regions capping the chromosomes and composed of multiple units of hexa-nucleotides, TTAGGG, making their quantification difficult. Most of the methods developed to estimate telomeres are extensively cumbersome or expensive. The quantitative polymerase chain reaction (qPCR) based assay is relatively easy and cheaper method that applies SyBr Green dye chemistry to measure telomere length. SyBr Green dye fluoresces after intercalation into the double stranded DNA (dsDNA), thus detection of unspecific products has been a limitation as it may affect quantitation of telomeres. To overcome this limitation of SyBr Green dye, we developed a dual labeled fluorescence probe based quantitative polymerase chain reaction (qPCR) to measure the telomere length. This highly efficient, yet cost effective and easy method, utilizes a probe that targets primarily the telomeric DNA and this increases accuracy of an existing qPCR method.

Evaluation of biomarkers, genetic mutations, and epigenetic modifications in early diagnosis of pancreatic cancer.

BACKGROUND: Pancreatic cancer (PC) is one of the deadliest malignancies with an alarming mortality rate. Despite significant advancement in diagnostics and therapeutics, early diagnosis remains elusive causing poor prognosis, marred by mutations and epigenetic modifications in key genes which contribute to disease progression. AIM: To evaluate the various biological tumor markers collectively for early diagnosis which could act as prognostic biomarkers and helps in future therapeutics of PC in Kashmir valley. METHODS: A total of 50 confirmed PC cases were included in the study to evaluate the levels of carbohydrate antigen 19-9 (CA 19-9), tissue polypeptide specific antigen (TPS), carcinoembryonic antigen (CEA), vascular endothelial growth factor-A (VEGF-A), and epidermal growth factor receptor (EGFR). Mutational analysis was performed to evaluate the mutations in Kirsten rat sarcoma (KRAS), Breast cancer type 2 (BRCA-2), and deleted in pancreatic cancer-4 (DPC-4) genes. However, epigenetic modifications (methylation of CpG islands) were performed in the promoter regions of cyclin-dependent kinase inhibitor 2A (p16; CDKN2A), MutL homolog 1 (hMLH1), and Ras association domain-containing protein 1(RASSF1A) genes. RESULTS: We found significantly elevated levels of biological markers CA 19-9 (P ≤ 0.05), TPS (P ≤ 0.05), CEA (P ≤ 0.001), and VEGF (P ≤ 0.001). Molecular genetic analysis revealed that KRAS gene mutation is predominant in codon 12 (16 subjects, P ≤ 0.05), and 13 (12 subjects, P ≤ 0.05). However, we did not find a mutation in DPC-4 (1203G > T) and BRCA-2 (617delT) genes. Furthermore, epigenetic modification revealed that CpG methylation in 21 (P ≤ 0.05) and 4 subjects in the promoter regions of the p16 and hMLH1 gene, respectively. CONCLUSION: In conclusion, CA 19-9, TPS, CEA, and VEGF levels were significantly elevated and collectively have potential as diagnostic and prognostic markers in PC. Global data of mutation in the KRAS gene commonly in codon 12 and rare in codon 13 could augment the predisposition towards PC. Additionally, methylation of the p16 gene could also modulate transcription of genes thereby increasing the predisposition and susceptibility towards PC.

Genetic evaluation of the variants using MassARRAY in non-small cell lung cancer among North Indians.

Lung cancer is genetically diverse and a major health burden. Non-small cell lung cancer (NSCLC) accounts for 80% of total lung cancer cases and 20% cases are Small cell lung cancer (SCLC). The present case-control association study focused on the cost effective high throughput genotyping using Agena MassARRAY matrix-assisted laser desorption/ionization-time of flight, mass spectrometry (MALDI-TOF) platform to analyze the genetic association of candidate genetic variants. We performed multiplex PCR and genotyped twelve single nucleotide polymorphisms (SNPs) in 723 samples (162 NSCLC cases and 592 healthy controls). These genetic variants were selected from literature for their association with various cancers worldwide and this is the first study from the region to examine these critically important genetic variants. With prospective case-control association study design, twelve variants from ten genes were evaluated. Amongst these six variants, TCF21 (rs12190287), ERCC1 (rs2298881, 11615), ERCC5 (rs751402), ARNTL (rs4757151), BRIP1 (rs4986764) showed significant association with NSCLC risk (p ≤ 0.003) in Jammu and Kashmir population. In-silico findings of these genetic variants showed remarkable functional roles that needs in-vitro validations. It is further anticipated that such case control studies will help us in understanding the missing heritability of non-small cell lung cancer.

Polymorphism in the TP63 gene imparts a potential risk for leukemia in the North Indian population.

BACKGROUND: The role of single nucleotide polymorphism rs10937405 (C>T) of the TP63 gene in cancer including leukemia has previously been studied in different world populations; however, the role of this variant in leukemia in the North Indian population of Jammu and Kashmir is still unknown. OBJECTIVES: In the present study, we investigated the association of genetic variant rs10937405 with leukemic in the Jammu and Kashmir population. METHODS: A total of 588 subjects, (188 cases and 400 controls) were recruited for the study. The rs10937405 variant was genotyped by using the real-time based TaqMan assay. RESULTS: A statistically significant association was observed between the rs10937405 and leukemia [OR of 1.94 (95% CI 1.51-2.48), p=1.2x10-6]. CONCLUSION: The current study concludes that the rs10937405 variant is a risk factor for the development of leukemia in the population of Jammu and Kashmir, North India. However, it would be interesting to explore the contribution of this variant in other cancers as well. Our findings will help in the development of diagnostic markers for leukemia in the studied population and potentially for other North Indian populations.

REV3L single nucleotide variants lead to increased susceptibility towards non-small cell lung cancer in the population of Jammu and Kashmir.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the most common lung cancer, accounting for 80-85% of all lung cancer cases. Various genetic studies have associated REV3L (Protein reversion less 3-like) gene mutations, which encodes the catalytic subunit of error prone translesion synthesis polymerase zeta with cancer, including lung cancer; however, no such data is available from any North Indian population. In this study we attempted to screen the North Indian population of Jammu and Kashmir (J&K) for the potential role of REV3L gene polymorphisms in NSCLC. METHODS: A total of four REV3L single nucleotide variants were selected for genotyping based on the available literature. The genotyping was carried out by using the TaqMan allele discrimination assay in 500 subjects (200 NSCLC patients and 300 age and sex matched healthy controls). The association of variants with NSCLC was evaluated by logistic regression. RESULTS: Out of the four REV3L variants genotyped; rs1002481, rs462779, and rs465646 were found significantly associated with NSCLC risk under the recessive model, with an Odds Ratio (OR) of 3.52(2.14-5.8 at 95% CI, p-value = 0.00000062), 3.7 (1.8-7.6 at 95% CI, p-value = 0.00031), and 2.2 (1.47-3.37 at 95% CI, p-value = 0.0003), respectively. DISCUSSION: Our data supports a strong association between variants rs1002481, rs462779, rs465646 and NSCLC, indicating a potential role of these REV3L variants in increasing the risk for the development of NSCLC in the studied population. Although a first report from any Indian population, these variants have been previously reported to be associated with lung and colorectal cancers in different world populations. Our data along with the existing data supports the notation that these variants can be used as potential genetic predisposition markers. AVAILABILITY OF DATA AND MATERIALS: Data generated and analysed during study is not available publicly but can be made available from the corresponding author upon reasonable request.