A novel algorithm to differentiate between primary lung tumors and distant liver metastasis in lung cancers using an exosome based multi gene biomarker panel.

The lack of non-invasive methods for detection of early metastasis is a crucial reason for the poor prognosis of lung cancer (LC) liver metastasis (LM) patients. In this study, the goal was to identify circulating biomarkers based on a biomarker model for the early diagnosis and monitoring of patients with LCLM. An 8-gene panel identified in our previous study was validated in CTC, cfRNA and exosomes isolated from primary lung cancer with & without metastasis. Further multivariate analysis including PCA & ROC was performed to determine the sensitivity and specificity of the biomarker panel. Model validation cohort (n = 79) was used to verify the stability of the constructed predictive model. Further, clinic-pathological factors, survival analysis and immune infiltration correlations were also performed. In comparison to our previous tissue data, exosomes demonstrated a good discriminative value with an AUC of 0.7247, specificity (72.48%) and sensitivity (96.87%) for the 8-gene panel. Further individual gene patterns led us to a 5- gene panel that showed an AUC of 0.9488 (p = < 0.001) and 0.9924 (p = < 0.001) respectively for tissue and exosomes. Additionally, on validating the model in a larger cohort a risk score was obtained (RS > 0.2) for prediction of liver metastasis with an accuracy of 95%. Survival analysis and immune filtration markers suggested that four exosomal markers were independently associated with poor overall survival. We report a novel blood-based exosomal biomarker panel for early diagnosis, monitoring of therapeutic response, and prognostic evaluation of patients with LCLM.

Androgen receptor: Structure, signaling, function and potential drug discovery biomarker in different breast cancer subtypes.

The Androgen Receptor (AR) is emerging as an important factor in the pathogenesis of breast cancer (BC), which is the most common malignancy worldwide. >70 % of AR expression in primary and metastatic breast tumors has been observed which suggests that AR may be a new marker and a potential therapeutic target among AR-positive BC patients. Biological insight into AR-positive breast cancer reveals that AR may cross-talk with several vital signaling pathways, including key molecules and receptors. Downstream signaling of AR might also affect many clinically important pathways that are emerging as clinical targets in BC. AR exhibits different behaviors depending on the breast cancer molecular subtype. Preliminary clinical research using AR-targeted drugs, which have already been FDA-approved for prostate cancer (PC), has given promising results for AR-positive breast cancer patients. However, since AR positivity's prognostic and predictive value remains uncertain, it is difficult to identify and stratify patients who would benefit from AR-targeted therapies alone. Thus, the need of the hour is to target the androgen receptor as a monotherapy or in combination with other conventional therapies which has proven to be an effective clinical strategy for the treatment of prostate cancer patients, and these therapeutic strategies are increasingly being investigated in breast cancer. Therefore, in this manuscript, we review the role of AR in various cellular processes that promote tumorigenesis and aggressiveness, in different subtypes of breast cancer, as well as discuss ongoing efforts to target AR for the more effective treatment and prevention of breast cancer.

Lipoprotein glomerulopathy: a rare cause of steroid-resistant nephrotic syndrome in a child.

Lipoprotein glomerulopathy (LPG) is a rare condition of renal lipidosis characterized by lipoprotein thrombi in glomeruli, an abnormal plasma lipoprotein profile, and a marked increase in serum apolipoprotein E (apo E) levels. It is a monogenic disorder with autosomal dominant inheritance and the average age of presentation is 32 years (4-69 years). It is rare in children. The presentation can be nephrotic syndrome, hematuria, or progressive renal failure. Here we report the first described case of LPG in an Indian 7.5-year-old boy who presented with steroid-resistant nephrotic syndrome with normal renal function. A renal biopsy was suggestive of lipoprotein glomerulopathy. The detection of a pathogenic variant in apo E, Kyoto type, by exome sequencing, confirmed the diagnosis of lipoprotein glomerulopathy. Complete response was achieved with Angiotensin-converting Enzyme inhibitor and fenofibrates.

Unravelling the role of Silibinin in targeting CD44+ cancer stem cells: Therapeutic implications, effective strategies and approaches.

CD44+ cancer stem cells (CSCs) are believed to account for drug resistance and tumour recurrence due to their potential to self-renew and differentiate into heterogeneous lineages. Therefore, efficient treatment strategies targeting and eliminating these CSCs are required. The flavonolignan, Silibinin, has gained immense attention in targeting CD44+ CSCs as it alters functional properties like cell cycle arrest, apoptosis, inhibition of invasion and metastasis and also inhibits a range of molecular pathways. However, its limited bioavailability is a major hurdle in asserting Silibinin as a translational therapeutic agent. Combinatorial therapy of Silibinin with conventional chemotherapeutic drugs is an alternative approach in targeting CD44+ CSCs as it increases the efficacy and reduces the cytotoxicity of chemotherapeutic drugs, thus preventing drug resistance. Certain Silibinin-conjugated nano-formulations have also been successfully developed, through which there is improved absorptivity/bioavailability of Silibinin and a decrease in the concentration of therapeutic drugs leading to reduced cytotoxicity. In this review, we summarise the effectiveness of the synergistic therapeutic approach for Silibinin in targeting the molecular mechanisms of CD44+ CSCs and emphasise the potential role of Silibinin as a novel therapeutic agent.

A case series on tofacitinib-induced weight gain.

Janus kinase inhibitor tofacitinib belongs to a group of targeted synthetic disease-modifying anti-rheumatic drugs, also known as small molecule inhibitors. They are oral drugs with a novel strategy to treat inflammatory diseases. The major concern with the use of these drugs is a high risk for infections and other potential side effects. Here, we have focused on reporting one of the rare side effects of tofacitinib, weight gain. We have reported six cases of tofacitinib-induced weight gain in patients of ankylosing spondylitis, rheumatoid arthritis, and vasculitis.

MX2 Viral Substrate Breadth and Inhibitory Activity Are Regulated by Protein Phosphorylation.

Human immunodeficiency virus type-1 (HIV-1) infection is potently inhibited by human myxovirus resistance 2 (MX2/MxB), which binds to the viral capsid and blocks the nuclear import of viral DNA. We have recently shown that phosphorylation is a key regulator of MX2 antiviral activity, with phosphorylation of serine residues at positions 14, 17, and 18 repressing MX2 function. Here, we extend the study of MX2 posttranslational modifications and identify serine and threonine phosphorylation in all domains of MX2. By substituting these residues with aspartic acid or alanine, hence mimicking the presence or absence of a phosphate group, respectively, we identified key positions that control MX2 antiviral activity. Aspartic acid substitutions of residues Ser306 or Thr334 and alanine substitutions of Thr343 yielded proteins with substantially reduced antiviral activity, whereas the presence of aspartic acid at positions Ser28, Thr151, or Thr343 resulted in enhanced activity: referred to as hypermorphic mutants. In some cases, these hypermorphic mutations, particularly when paired with other MX2 mutations (e.g., S28D/T151D or T151D/T343A) acquired the capacity to inhibit HIV-1 capsid mutants known to be insensitive to wild-type MX2, such as P90A or T210K, as well as MX2-resistant retroviruses such as equine infectious anemia virus (EIAV) and murine leukemia virus (MLV). This work highlights the complexity and importance of MX2 phosphorylation in the regulation of antiviral activity and in the selection of susceptible viral substrates. IMPORTANCE Productive infection by human immunodeficiency virus type-1 (HIV-1) requires the import of viral replication complexes into the nuclei of infected cells. Myxovirus resistance 2 (MX2/MxB) blocks this step, halting nuclear accumulation of viral DNA and virus replication. We recently demonstrated how phosphorylation of a stretch of three serines in the amino-terminal domain of MX2 inhibits the antiviral activity. Here, we identify additional positions in MX2 whose phosphorylation status reduces or enhances antiviral function (hypomorphic and hypermorphic variants, respectively). Importantly, hypermorphic mutant proteins not only increased inhibitory activity against wild-type HIV-1 but can also exhibit antiviral capabilities against HIV-1 capsid mutant viruses that are resistant to wild-type MX2. Furthermore, some of these proteins were also able to inhibit retroviruses that are insensitive to MX2. Therefore, we propose that phosphorylation comprises a major element of MX2 regulation and substrate determination.

Salinomycin mediated therapeutic targeting of circulating stem like cell population in oral cancer.

CD44+ circulating tumor stem cells (CTSCs) have been significantly associated with aggressiveness, resistance and poor prognosis of oral cancer patients. Thus, targeted elimination of these CTSCs could be a new conceptual framework for enhancing the therapeutic outcome of patients. Docking of potential investigational molecules and simulation results identified Salinomycin as a potential lead compound that could effectively inhibit CD44 receptor. To assess the cytotoxic effect, immuno-magnetically sorted circulatory CD44+ cells were subjected to increasing concentrations of 5FU, Cisplatin and Salinomycin. Salinomycin demonstrated significant cytotoxic effect towards the CD44+ subpopulation in a dose and time dependent manner. Further the effect of these compounds was investigated on apoptosis, cell cycle, signaling pathways and gene expression profiles using MuseTM flow cytometer and Real-Time PCR. It was observed that mRNA expression patterns of CD44v6, Nanog, AKT1, CDKN2A and ß-catenin of Salinomycin treated CD44+ cells. Moreover, Salinomycin significantly induced programmed cell death by inducing G2/M cell cycle arrest and inhibiting MAPK/PI3K pathways in this chemo-resistant population. Thus, this study demonstrated the potential of Salinomycin to target the chemo-resistant circulating CD44 population by attenuating its proliferation and survival.Communicated by Ramaswamy H. Sarma.

EGFR Mutation Analysis in Non-small Cell Lung Carcinoma Patients: A Liquid Biopsy Approach.

In the era of the targeted therapy identification of EGFR mutation detection in lung cancer is extremely helpful to predict the treatment efficacy of EGFR tyrosine kinase inhibitors (TKIs). Unfortunately, the inadequacy and quality of the biopsy samples are the major obstacles in molecular testing of EGFR mutation in lung cancer. To address this issue, the present study intended to use liquid biopsy as the non-invasive method for EGFR mutation detection. A total of 31 patients with an advanced stage of lung cancer were enrolled in the study from which cell-free DNA (cfDNA) and FFPE tissue DNA was extracted. Extracted DNA samples were analyzed for further EGFR exon specific mutation analysis by ARMS-PCR. Data were analyzed statistically using SPSS software. In cfDNA samples, the prevalence of wild type EGFR was 48% while the prevalence of TKI resistant and TKI sensitive mutations were 3%. Conversely, in tissue DNA samples, the prevalence of wild type, TKI sensitive and TKI resistant mutations were 48%, 19%, and 3%, respectively. The overall concordance of EGFR mutation between cfDNA and tissue DNA was 83%. McNemar's test revealed that there was no significant difference between EGFR expression of cfDNA and tissue DNA samples. Additionally, the significant-high incidence of TKI resistant mutations was observed in tobacco habituates, indicating the role of carcinogens present in the tobacco in developing resistant mutations. In conclusion, our data suggest that evaluation of EGFR mutation from cfDNA samples is practicable as a non-invasive tool in patients with advanced-stage of lung cancer.

Putative stemness markers octamer-binding transcription factor 4, sex-determining region Y-box 2, and NANOG in non-small cell lung carcinoma: A clinicopathological association.

BACKGROUND: The promising improvement in the clinical outcome of lung cancer can be possibly achieved by identification of the molecular events that underlie its pathogenesis. Cancer stem cell (CSC) being one of the subsets of tumor majorly participates in drug resistance and treatment failure because of the moderate cell cycle, lower proliferation, and increased expression of DNA repair and anti-apoptosis genes. Although many putative CSC markers exist, a precise characterization for non-small cell lung cancer is of utmost importance due to increased mortality rate and lack of targeted therapies. Hence, the article focuses on the expression of stemness-associated markers, namely octamer-binding transcription factor 4 (OCT4), NANOG, and sex-determining region Y-box 2 (SOX2) in non-small cell lung cancer (NSCLC) patients. METHODS: The expression of OCT4, NANOG, and SOX2 were evaluated in 32 histopathologically confirmed NSCLC tissues using real-time polymerase chain reaction. The obtained expression was correlated with clinical and pathological manifestations using the statistical test such as Student's t-test and Pearson correlation in varied statistical software. RESULTS: Results showed a significantly higher expression of OCT4 and NANOG compared to SOX2 in the tumor tissues. When the expression of these markers was correlated with the clinical parameters, higher expression was seen in males, patients with age above 60 years, and in adenocarcinoma subtype. In correlation with the habit, higher expression of OCT4 and SOX2 was observed in habituated patients. Expression of NANOG and OCT4 was higher even in patients with poor differentiation. CONCLUSION: The expression and prognostic significance of CSC markers obviously vary depending on histological NSCLC subtype. Importantly, our findings suggest that OCT4, SOX2, and NANOG network together may be promising for ongoing targeted therapies in specific NSCLC subgroups.

A Prognostic and Predictive Study of BCR-ABL Expression Based on Characterization of Fusion Transcripts.

BCR-ABL translocation is a key hallmark of chronic myeloid leukemia (CML). The chemistry involves transcription of a novel 8.5 kb mRNA with a b3a2 and/ or b2a2 junction. Though there is an improvement in survival using the TKIs, there are causes for the treatment failures. Thus, the study focuses on the causes underlying the failed response. This study comprises BCR-ABL expression in correlation with age, gender and transcript type and disease monitoring in follow-up samples. Eighty-seven chronic phase CML patients were enrolled in the study. Out of these 24 patients were followed further. Quantitative Real time PCR was performed to assess the BCR-ABL expression followed by gel electrophoresis of PCR products. The results were expressed as CN/µg RNA. The results obtained were correlated with SPSS 16.0 software. We found three different types of the expression that includes b2a2, b3a2 and co-expression (b2a2 + b3a2). Higher incidence of b2a2 with a relatively equal incidence of co-expression was observed. The BCR-ABL expression was higher in males, in young patients and in those exhibiting co-expression of the transcripts. Greater relapse was seen in females and in older patients. The patients with co-expression of transcripts exhibited the highest expression; however these patients showed the best treatment response suggest the co-expression is the favourable parameter for CML patients. The transcript type and BCR-ABL expression are well correlated and hence can be considered as a prognostic as well as the predictive indicator considering the BCR-ABL expression for CML patients.

Transcriptome-wide miRNA identification of Bacopa monnieri: a cross-kingdom approach.

Bacopa monnieri known as 'Brahmi' is a well-known medicinal plant belonging to Scrophulariaceae family for its nootropic properties. To the best of our knowledge, no characterization data is available on the potential role of micro RNAs (miRNAs) from this plant till date. We present here the first report of computational characterizations of miRNAs from B. monnieri. Owing to the high conservation of miRNAs in nature, new and potential miRNAs can be identified in plants using in silico techniques. Using the plant miRNA sequences present in the miRBase repository, a total of 12 miRNAs were identified from B. monnieri which pertained to 11 miRNA families from the shoot and root transcriptome data. Furthermore, gene ontology analysis of the identified 68 human target genes exhibited significance in various biological processes. These human target genes were associated with signaling pathways like NF-kB and MAPK with TRAF2, CBX1, IL1B, ITGA4 and ITGB1BP1 as the top five hub nodes. This cross-kingdom study provides initial insights about the potential of miRNA-mediated cross-kingdom regulation and unravels the essential target genes of human with implications in numerous human diseases including cancer.

Computational identification of miRNA and their cross kingdom targets from expressed sequence tags of Ocimum basilicum.

MicroRNAs (miRNAs) are conserved small non coding RNAs, which are typically 22-24 nucleotides long and play an important role in post transcription regulation andin various biological processes in both animals and plants. Ocimum basilicum is an important medicinal plant having different bioactive compounds eugenol and essential oils that possess numerous therapeutic properties. However, only a few miRNAs of Ocimum basilicum and its function have been studied till date. The present study focusses on the identification of miRNA from expressed sequenced tags by carrying out computational approaches based on the homology search method. A total of 10 potential miRNAs with 8 different families were predicted in O.basilicum. Furthermore, the psRNA target server was used to predict cross kingdom target genes on human transcriptome for identification ofpotential miRNAs. Eight miRNA families were found to modulate the 87 human target genes which were associated with RAS/MAPK signalling cascade, cardiomyopathy, HIV, breast cancer, lung cancer, Alzheimer's diseases and several neurological disorders. Moreover, O.basilicum miRNAs regulate the key human target genes having significance in various diseases and important biological networks with 10 hub nodes interactions. Thus this study gives the pave for further studies to explore the potential of miRNA mediated cross kingdom regulation and treatment of various diseases including cancer.

Genetic and Epigenetic Modulation of Drug Resistance in Cancer: Challenges and Opportunities.

Cancer is a complex disease that has the ability to develop resistance to traditional therapies. The current chemotherapeutic treatment has become increasingly sophisticated, yet it is not 100% effective against disseminated tumours. Anticancer drugs resistance is an intricate process that ascends from modifications in the drug targets suggesting the need for better targeted therapies in the therapeutic arsenal. Advances in the modern techniques such as DNA microarray, proteomics along with the development of newer targeted drug therapies might provide better strategies to overcome drug resistance. This drug resistance in tumours can be attributed to an individual's genetic differences, especially in tumoral somatic cells but acquired drug resistance is due to different mechanisms, such as cell death inhibition (apoptosis suppression) altered expression of drug transporters, alteration in drug metabolism epigenetic and drug targets, enhancing DNA repair and gene amplification. This review also focusses on the epigenetic modifications and microRNAs, which induce drug resistance and contributes to the formation of tumour progenitor cells that are not destroyed by conventional cancer therapies. Lastly, this review highlights different means to prevent the formation of drug resistant tumours and provides future directions for better treatment of these resistant tumours.

Publisher Correction: Silibinin, A Natural Blend In Polytherapy Formulation For Targeting Cd44v6 Expressing Colon Cancer Stem Cells.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Silibinin, A Natural Blend In Polytherapy Formulation For Targeting Cd44v6 Expressing Colon Cancer Stem Cells.

Colon cancer stem cells have been attributed to poor prognosis, therapeutic resistance and aggressive nature of the malignancy. Recent reports associated CD44v6 expression with relapse, metastasis and reduced 5-year survival of colon cancer patients, thereby making it a potential therapeutic target. Thus, in this study, comprehensive prediction and screening of CD44v6 against 1674 lead compounds was conducted. Silibinin was identified as a potential compound targeting CD44v6. Inorder to substantiate these findings, the cytotoxic effect of 5FU, Silibinin and 5FU+ Silibinin was assessed on human colon carcinoma cell line HCT116 derived CD44+ subpopulation. 5FU+ Silibinin inhibited cell proliferation of CD44+ subpopulation at lower concentration than Silibinin standalone. Further, corresponding to CD44v6 knockdown cells, 5FU+ Silibinin treatment significantly decreased CD44v6, Nanog, CTNNB1 and CDKN2A expression whereas increased E-cadherin expression in HCT116 derived CD44+ cells. Moreover, synergistic effect of these drugs suppressed sphere formation, inhibited cell migration, triggered PARP cleavage and perturbation in mitochondrial membrane potential, thereby activating intrinsic apoptotic pathways and induced autophagic cell death. Importantly, 5FU+ Silibinin could inhibit PI3K/MAPK dual activation and arrest the cell cycle at G0/G1 phase. Thus, our study suggests that inhibition of CD44v6 attenuates stemness of colon cancer stem cells and holds a prospect of potent therapeutic target.

A multi-gene expression profile panel for predicting liver metastasis: An algorithmic approach.

BACKGROUND & AIM: Liver metastasis has been found to affect outcome in prostate, pancreatic and colorectal cancers, but its role in lung cancer is unclear. The 5 year survival rate remains extensively low owing to intrinsic resistance to conventional therapy which can be attributed to the genetic modulators involved in the pathogenesis of the disease. Thus, this study aims to generate a model for early diagnosis and timely treatment of liver metastasis in lung cancer patients. METHODS: mRNA expression of 15 genes was quantified by real time PCR on lung cancer specimens with (n = 32) and without (n = 30) liver metastasis and their normal counterparts. Principal Component analysis, linear discriminant analysis and hierarchical clustering were conducted to obtain a predictive model. The accuracy of the models was tested by performing Receiver Operating Curve analysis. RESULTS: The expression profile of all the 15 genes were subjected to PCA and LDA analysis and 5 models were generated. ROC curve analysis was performed for all the models and the individual genes. It was observed that out of the 15 genes only 8 genes showed significant sensitivity and specificity. Another model consisting of the selected eight genes was generated showing a specificity and sensitivity of 90.0 and 96.87 respectively (p <0.0001). Moreover, hierarchical clustering showed that tumors with a greater fold change lead to poor prognosis. CONCLUSION: Our study led to the generation of a concise, biologically relevant multi-gene panel that significantly and non-invasively predicts liver metastasis in lung cancer patients.

Unravelling the link between embryogenesis and cancer metastasis.

PURPOSE: Cancer as opposed to embryonic development is characterized by dysregulated, uncontrolled and clonal growth of cells. Inspite of that they share certain commonality in gene expression patterns and a number of cellular & molecular features. Consequently, in the present study we aimed to evaluate the role of a definite set of genes in fetal liver, primary liver cancers and metastatic liver tissue. METHODS: The relative expression of fourteen candidate genes obtained by data mining and manual curation of published data (CXCL12, CXCR4, CK7, CDH1, CTNNB1, CLDN4, VEGFA, HIF1A, MMP9, p53, OPN, CDKN2A, TGFBR2, MUC16, ß-actin) were performed on 62 tissues (32 liver metastasis tissues and 30 primary Liver cancer tissues), Fetal liver tissues (below and above 20weeks of gestation) and 2 sets of control samples by real-time quantitative reverse transcription PCR (qRT-PCR). RESULTS: Results showed significant down-regulation of MMP9 and TP53 in Fetal liver above 20weeks of gestation whereas it was up-regulated in fetal liver below 20weeks of gestation, primary liver cancers and liver metastasis. Contradictory to that OPN and CDKN2A were significantly up-regulated in primary liver cancer, liver metastasis; down-regulated in fetal liver above 20weeks of gestation but were not expressed during early embryo development (below 20weeks of gestation). Moreover, MMP9 and TP53 demonstrated a strong correlation with MUC16 whereas CDKN2A and OPN showed correlation with CXCL12/CXCR4 signifying that MUC16, CXCL12/CXCR4 might be involved in the complex process of cancer metastasis. CONCLUSION: MMP9, OPN, TP53 and CDKN2A were the identified markers that were expressed in a similar pattern in early embryonic development and cancer development & invasion suggesting that these genes are activated during embryogenesis and might be re-expressed in cancer metastasis. Moreover, these genes govern a pathway that might be activated during cancer metastasis. Thus, targeting these molecules may provide better treatment for metastatic liver cancers.

Natural Compounds as Epigenetic Regulators of Human Dendritic Cell-mediated Immune Function.

Dendritic cells (DCs) are the most potent professional antigen-presenting cells (APCs) and are poised to capture antigen, migrate to draining lymphoid organs, and postmaturation process. Recent evidences have suggested that tumor microenvironment has an effect on DCs by inactivating various components of the immune system responsible for tumor clearance, eventually leading to tumorigenesis. This inactivation is owed to the epigenetic modifications [ie, microRNA (miRNA)] at the posttranscriptional level, thus regulating the differentiation patterns and functional behavior of DCs. Thus, need of the hour is to develop protocols for ex vivo generation of DCs which may provide a foundation for designing and developing DC-based vaccination for treatment of solid tumors. To achieve this, it is crucial to modulate DCs by identifying miRNAs which may increase the efficacy of DC-based vaccines by reprogramming the immunosuppressive nature of tumor microenvironment. Furthermore, it would be an interesting aspect to check the immunomodulatory potential of natural compounds in reprogramming the immune responses through DCs. Thus, this review aims to improvise the understanding of DC immune biology and miRNAs at genetic level in cancer which can be pivotal for designing novel or improved therapeutic approaches that will allow proper functioning of DCs in patient care. Furthermore, we have highlighted the candidate target molecules and signaling mechanisms having a vital role in the immune-modulatory activities of natural compounds and its derived phytocompounds. This review also establishes a link between miRNA expressions and biological roles of natural compounds modulating the activity of DCs.

Insights into the structural perturbations of spliced variants of CD44: a modeling and simulation approach.

Transient interactions between cancer stem cells and components of the tumor microenvironment initiate various signaling pathways crucial for carcinogenesis. Predominant hyaluronan (HA) receptor, CD44 is structurally and functionally one of the most variable cell surface receptors having the potential to generate a diverse repertory of CD44 isoforms by alternative splicing of variant exons and post-translational modifications. A structurally distinctive variant of CD44, CD44v10, has an inevitable role in malignant progression, invasion, and metastasis. This can be attributed to the binding of HA with CD44v10, which demonstrates a completely different behavioral pattern as compared to the other spliced variants of CD44 molecule. Absence of a comprehensively predicted crystal structure of human CD44s and CD44v10 is an impediment in understanding the resultant structural alterations caused by the binding of HA. Thus, in this study, we aim to predict the CD44s and CD44v10 structures to their closest native confirmation and study the HA binding-induced structural perturbations using homology modeling, molecular docking, and MD simulation approach. The results depicted that modeled 3D structures of CD44s and CD44v10 isoforms were found to be stable throughout MD simulations; however, a substantial decrease was observed in the binding affinity of HA with CD44v10 (-5.355 kcal/mol) as compared to CD44s. Furthermore, loss and gain of several H-bonds and hydrophobic interactions in CD44v10-HA complex during the simulation process not only elucidated the reason for decreased binding affinity for HA but also prompted toward the plausible role of HA-induced structural perturbations in occurrence and progression of carcinogenesis.

Circulating tumor stem like cells in oral squamous cell carcinoma: An unresolved paradox.

OBJECTIVE: Circulating tumor cells (CTCs) are increasingly gaining importance due to their immense potential in enhancing diagnosis, prognosis and response to therapy in solid malignancies. Therefore, we aimed to comprehend the molecular diversity and critical role of this disseminated tumor population in OSCC. METHODOLOGY: CD44+ subpopulation was isolated using immuno-magnetic cell separation and their purity was validated using flow cytometry. Characterisation of self renewal potential and resistance to chemotherapy was assessed using tumor sphere forming and cytotoxicity assay. Gene expression profile of pertinent CSC (CD44s, CD44v3, CD44v6) and stemness markers (Bmi1 and Nanog) was carried out in CD44+ cells using Real Time PCR. Predominantly expressed markers and their association with clinico-pathological conditions were substantiated in 30 OSCC patients. RESULT: Flow cytometry analysis depicted a predominant population of CD44+CD24-CD45- cells suggesting that circulating tumor cells had a subpopulation of CSC like cells in the circulation. These cells demonstrated increased sphere forming capability and intrinsic chemo-resistance compared to non-CSC, thus indicating the CSC features of self-renewal and chemo-resistance. Additionally, CD44+ cells showed significantly increased expression levels of CD44v6 and Nanog compared to CD44- cells. Clinically, expression pattern of CD44v6 and Nanog correlated with different anatomical subsites, loco-regional aggressiveness of the disease and recurrence, thus opening newer avenues that can be explored for better prognostic and therapeutic implications. CONCLUSION: This study explored the inevitable role of CD44v6 and Nanog as circulating stem like cell markers in assessment of loco-regional aggressiveness, detection of relapse and therapeutic response and resistance.