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Osteoarthritis (OA) is a chronic degenerative joint disease. The pathogenesis is poorly understood. What is known is that OA is characterized by imbalance in anabolic and catabolic gene expression in articular chondrocytes. This results in bone on bone articulations resulting in impaired mobility and joint pain. Although the cause of OA is unknown, comorbidities include: aging, obesity, and mechanical stress. Currently the only diagnostic modalities are radiology and physical examination, and early detection is rare. Biomarkers are quantifiable substances, and their presence can be suggestive of a certain phenomenon or disease. Biomarkers are popular for early diagnosis for pathological conditions in the fields of oncology, cardiology, and endocrinology. This review has systematically reviewed the literature about biomarkers in the field of OA, specifically protein, miRNA, and metabolic biomarkers found in the blood, urine, and synovial fluid.

In Europid populations, low birth weight of offspring predicts insulin resistance in the mother and cardiovascular disease in both parents. We investigated the association between birth weight of offspring and obesity and cardiovascular risk in the parents of 477 8-year-old children born at the King Edward Memorial Hospital, Pune, India. Eight years after the birth of the child, mothers (33 years of age, n = 459) of heavier babies were taller and more obese (BMI, fat mass, and waist circumference, all P < 0.001) than mothers of lighter babies. Increasing offspring birth weight predicted higher homeostasis model assessment for insulin resistance (P < 0.01) and metabolic syndrome in mothers (P < 0.001) (adjusted for offspring sex and birth order, maternal age, and socioeconomic status) but not hyperglycemia. Fathers (39 years of age, n = 398) of heavier babies were taller and heavier, independent of maternal size (P < 0.01, both), but were not more insulin resistant. Unlike other reports, lower offspring birth weight did not predict insulin resistance in fathers. Thus, urban Indian parents have a higher risk of being obese 8 years after delivery of a heavier child. Mothers but not fathers of heavier babies also have a higher risk of being insulin resistant and developing the metabolic syndrome. Our findings highlight the need for a better understanding of the relation between fetal growth and future health before contemplating public health interventions to improve fetal growth. [ABSTRACT FROM AUTHOR]

Background: Transcript variants and protein isoforms are central to unique tissue functions and maintenance of homeostasis, in addition to being associated with aberrant states such as cancer, where their crosstalk with the mutated tumor suppressor p53 may contribute to genomic instability and chromosomal rearrangements. We previously identified several novel splice variants in ovarian cancer RNA-sequencing datasets; herein, we aimed to elucidate the biological effects of the Integrin Subunit Beta 8 variant (termed pITGB8-205)., Methods: Resolution of the full-length sequence of pITGB8-205 through rapid amplification of cDNA ends (RACE-PCR). Cell cycle analysis and karyotype studies were performed to further explore genomic instability. RNA-seq and proteomics analyses were used to identify the differential expression of the genes., Results: This full-length study revealed a unique 5' sequence in pITGB8-205 that differed from the reported ITGB8-205 sequence, suggesting differential regulation of this novel transcript. Under a p53 mutant background, overexpression of pITGB8-205 triggered genetic instability reminiscent of oncogene-induced replicative stress with extensive abnormal mitoses and chromosomal and nuclear aberrations indicative of chromosomal instability, leading to near whole-genome duplication that imposes energy stress on cellular resources. Micronuclei and aneuploidy are striking features of pITGB8-205-overexpressing p53-mutant cells but are not enhanced in p53 wild-type (WT) cells. RNA-seq and proteomics analyses further suggested that p53 inactivation in ovarian cancer provides a permissive intracellular molecular niche for pITGB8-205 to mediate its effects on genomic instability. This observation is pivotal considering that most high-grade serous ovarian carcinoma (HGSC) tumors express mutant p53. The resulting aneuploid clones with enhanced self-renewal and survival capabilities disrupt clonal dominance under stress yet maintain a balance between replicative stress and prosurvival advantages., Conclusion: pITGB8-205-overexpressing clones sustain ovarian tumor cell survival, achieve homeostasis and are formidable opponents of therapy., (© 2024. The Author(s).)

High-grade serous ovarian carcinoma (HGSC) is associated with late-stage disease presentation and poor prognosis, with a limited understanding of early transformation events. Our study analyzes HGSC tumor progression and organ-specific metastatic dissemination to identify hypoxia-associated molecular, cellular, and histological alterations. Clinical characteristics of the HGSC were replicated in orthotopic xenografts, which involve metastatic dissemination and the prevalence of group B tumors (volume: >0.0625 ≤ 0.5 cm3). Enhanced hyaluronic acid (HA) deposition, expanded tumor vasculature, and increased necrosis contributed to the remodeling of tumor tissue architecture. The proliferative potential of tumor cells and the ability to form glands were also altered during tumor growth. Flow cytometry and label chase-based molecular profiling across the tumor regenerative hierarchy identified the hypoxia-vasculogenic niche and the hybrid epithelial-mesenchymal tumor-cell state as determinants of self-renewal capabilities of progenitors and cancer stem cells. A regulatory network and mathematical model based on tumor histology and molecular signatures predicted hypoxia-inducible factor 1-alpha (HIF1A) as a central node connecting HA synthesis, epithelial-mesenchymal transition, metabolic, vasculogenic, inflammatory, and necrotic pathways in HGSC tumors. Thus, our findings provide a temporal resolution of hypoxia-associated events that sculpt HGSC tumor growth; an in-depth understanding of it may aid in the early detection and treatment of HGSC., (© 2022 S. Karger AG, Basel.)

The molecular and functional diversity generated by chimeric transcripts (CTs) that are derived from two genes is indicated to contribute to tumor cell survival. Several gaps yet exist. The present research is a systematic study of the spectrum of CTs identified in RNA sequencing datasets of 160 ovarian cancer samples in the The Cancer Genome Atlas (TCGA) (https://portal.gdc.cancer.gov). Structural annotation revealed complexities emerging from chromosomal localization of partner genes, differential splicing and inclusion of regulatory, untranslated regions. Identification of phenotype-specific associations further resolved a dynamically modulated mesenchymal signature during transformation. On an evolutionary background, protein-coding CTs were indicated to be highly conserved, while non-coding CTs may have evolved more recently. We also realized that the current premise postulating structural alterations or neighbouring gene readthrough generating CTs is not valid in instances wherein the parental genes are genomically distanced. In addressing this lacuna, we identified the essentiality of specific spatiotemporal arrangements mediated gene proximities in 3D space for the generation of CTs. All these features together suggest non-random mechanisms towards increasing the molecular diversity in a cell through chimera formation either in parallel or with cross-talks with the indigenous regulatory network., Competing Interests: None of the authors have any competing financial interests to declare in this work., (© 2023 The Authors.)

RNA-binding proteins (RBPs) are pivotal for regulating gene expression as they are involved in each step of RNA metabolism. Several RBPs are essential for viable growth and development in mammals. RNA-binding motif 47 (RBM47) is an RRM-containing RBP whose role in mammalian embryonic development is poorly understood yet deemed to be essential since its loss in mouse embryos leads to perinatal lethality. In this study, we attempted to elucidate the significance of RBM47 in cell-fate decisions of mouse embryonic stem cells (mESCs). Downregulation of Rbm47 did not affect mESC maintenance and the cell cycle but perturbed the expression of primitive endoderm (PrE) markers and increased GATA4 + PrE-like cells. However, the PrE misregulation could be reversed by either overexpressing Rbm47 or treating the knockdown mESCs with the inhibitors of FGFR or MEK, suggesting an implication of RBM47 in regulating FGF-ERK signaling. Rbm47 knockdown affected the multi-lineage differentiation potential of mESCs as it regressed teratoma in NSG mice and led to a skewed expression of differentiation markers in serum-induced monolayer differentiation. Further, lineage-specific differentiation revealed that Rbm47 is essential for proper differentiation of mESCs towards neuroectodermal and endodermal fate. Taken together, we assign a hitherto unknown role(s) to RBM47 in a subtle regulation of mESC differentiation., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The rapid development of advanced high throughput technologies and introduction of high resolution "omics" data through analysis of biological molecules has revamped medical research. Single-cell sequencing in recent years, is in fact revolutionising the field by providing a deeper, spatio-temporal analyses of individual cells within tissues and their relevance to disease. Like conventional sequencing, the single-cell approach deciphers the sequence of nucleotides in a given Deoxyribose Nucleic Acid (DNA), Ribose Nucleic Acid (RNA), Micro Ribose Nucleic Acid (miRNA), epigenetically modified DNA or chromatin DNA; however, the unit of analyses is changed to single cells rather than the entire tissue. Further, a large number of single cells analysed from a single tissue generate a unique holistic perception capturing all kinds of perturbations across different cells in the tissue that increases the precision of data. Inherently, execution of the technique generates a large amount of data, which is required to be processed in a specific manner followed by customised bioinformatic analysis to produce meaningful results. The most crucial role of single-cell sequencing technique is in elucidating the inter-cell genetic, epigenetic, transcriptomic and proteomic heterogeneity in health and disease. The current review presents a brief overview of this cutting-edge technology and its applications in medical research., Competing Interests: The authors have none to declare., (© 2022 Director General, Armed Forces Medical Services. Published by Elsevier, a division of RELX India Pvt. Ltd.)

Background: Dental remineralization is the process of transporting minerals from the surrounding environment (i.e., saliva and biofilm) into partially demineralized tooth structures. Remineralization can be induced by professional therapies such as fluoride-based treatments that have the highest level of supporting evidence. High-intensity LASER and nonthermal atmospheric pressure plasma therapy have been known to increase the resistance of enamel to demineralization by surface modification., Aim: The aim of the study was to evaluate and compare the surface remineralization of enamel using ErCr:YSGG LASER and nonthermal atmospheric pressure plasma (NTP) activation with three different fluoride varnishes., Methodology: Sixty-eight extracted premolars were used which were sectioned mesiodistally to obtain 135 specimens and artificial caries were induced on the experimental surface. They were then randomly divided into three groups (n = 45): MI Varnish (GC Japan), Vanish Varnish (3M ESPE), and Embrace Varnish (Pulpdent). After varnish application, these groups were further divided into three subgroups based on the activation therapy used. Fifteen samples from each group were treated with ErCr:YSGG LASER, 15 samples with NTP, and 15 samples were the control that did not undergo activation. After 9 days of pH cycling, the mean ion concentration of the surface calcium and phosphate ions was recorded using FEG-SEM and EDX analysis. The data were statistically analyzed., Results: One-way ANOVA and post hoc Tukey test accepting P < 0.05 were performed for comparisons between all analyses groups. Vanish Varnish showed a higher Ca/P ratio in LASER, NTP, and control subgroups, followed by MI Varnish and Embrace Varnish. ErCr:YSGG LASER therapy showed an improved Ca/P ratio in all varnishes than NTP therapy and control., Conclusion: ErCr:YSGG LASER therapy showed positive effects toward improving the Ca/P, followed by NTP therapy as compared to control in all three varnishes indicating their role in enhancing the effects of remineralization. Vanish Varnish showed a higher Ca/P ratio indicating better remineralization post activation., Competing Interests: None

Background: Glioblastoma initiation and progression is believed to be driven by Glioma stem cells (GSCs). Activation of NOTCH1 and WNT, and more recently, non-canonical WNT5A signaling, has been demonstrated to regulate self-renewal and differentiation of the GSCs crucially. High expression levels of NOTCH1 and WNT in GBM tumors contribute to the sustenance of GSCs and mediate characteristic phenotypic plasticity, which is reflected by the different subtypes and tremendous intra-tumor heterogeneity. However, the coregulation of NOTCH1 and WNT5A is not well understood. Here, we studied the role of these molecules in regulating the characteristics of different GSC subtypes., Methods: We established a novel GSC-enriched cell model, referred to as NSG-70, from a patient with recurrent GBM. NSG-70 cells harbor a unique cytogenetic feature, viz. isochromosome 9q. At the same time, its expression profiles indicate that it is a mixed lineage comprising proneural and mesenchymal subtypes. We examined the relevance of NOTCH1 and WNT5A signaling and their coordinated action in GBM using these cells and other patient-derived models representing different GSC subtypes., Results: Our data revealed that the downregulation of NOTCH1 resulted in the suppression of stem cell and mesenchymal markers and significantly reduced the levels of WNT5A. NOTCH1 knockdown also led to a notable reduction in the vasculogenic mimicry of GSCs. Interestingly, knockdown of WNT5A exhibited similar effects and drove quiescent GSC towards proliferation. In a complementary manner, ectopic expression of WNT5A or rhWNT5A treatment rescued the effects of NOTCH1 knockdown., Conclusion: The resistance of GSCs towards conventional therapies in part due to subtype interconversion demands therapies targeting specific GSC subtype. Our study suggests the need for a combinatorial approach that could effectively target the NOTCH1-WNT5A signaling axis toward eliminating GSCs., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The involvement of cancer stem cells (CSCs) in driving tumor dormancy and drug resistance is well established. Most therapeutic regimens however are ineffective in targeting these regenerative populations. We report the development and evaluation of a monoclonal antibody, mAb150, which targets the metastasis associated antigen, Annexin A2 (AnxA2) through recognition of a N-terminal epitope. Treatment with mAb150 potentiated re-entry of CSCs into the cell cycle that perturbed tumor dormancy and facilitated targeting of CSCs as was validated by in vitro and in vivo assays. Epigenetic potentiation further improved mAb150 efficacy in achieving total tumor regression by targeting regenerative populations to achieve tumor regression, specifically in high-grade serous ovarian adenocarcinoma., (Copyright © 2021. Published by Elsevier Inc.)

RNA-binding proteins (RBPs) are critical players in the post-transcriptional regulation of gene expression and are associated with each event in RNA metabolism. The term 'RNA-binding motif' (RBM) is assigned to novel RBPs with one or more RNA recognition motif (RRM) domains that are mainly involved in the nuclear processing of RNAs. RBM47 is a novel RBP conserved in vertebrates with three RRM domains whose contributions to various aspects of cellular functions are as yet emerging. Loss of RBM47 function affects head morphogenesis in zebrafish embryos and leads to perinatal lethality in mouse embryos, thereby assigning it to be an essential gene in early development of vertebrates. Its function as an essential cofactor for APOBEC1 in C to U RNA editing of several targets through substitution for A1CF in the A1CF-APOBEC1 editosome, established a new paradigm in the field. Recent advances in the understanding of its involvement in cancer progression assigned RBM47 to be a tumor suppressor that acts by inhibiting EMT and Wnt/[Formula: see text]-catenin signaling through post-transcriptional regulation. RBM47 is also required to maintain immune homeostasis, which adds another facet to its regulatory role in cellular functions. Here, we review the emerging roles of RBM47 in various biological contexts and discuss the current gaps in our knowledge alongside future perspectives for the field., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Cellular plasticity and transitional phenotypes add to complexities of cancer metastasis that can be initiated by single cell epithelial to mesenchymal transition (EMT) or cooperative cell migration (CCM). Our study identifies novel regulatory cross-talks between Tcf21 and Slug in mediating phenotypic and migration plasticity in high-grade serous ovarian adenocarcinoma (HGSC). Differential expression and subcellular localization associate Tcf21, Slug with epithelial, mesenchymal phenotypes, respectively; however, gene manipulation approaches identify their association with additional intermediate phenotypic states, implying the existence of a multistep epithelial-mesenchymal transition program. Live imaging further associated distinct migratory modalities with the Tcf21/Slug status of cell systems and discerned proliferative/passive CCM, active CCM and EMT modes of migration. Tcf21-Slug balance identified across a phenotypic spectrum in HGSC cell lines, associated with microenvironment-induced transitions and the emergence of an epithelial phenotype following drug exposure. Phenotypic transitions and associated functionalities following drug exposure were affirmed to ensue from occupancy of Slug promoter E-box sequences by Tcf21. Our study effectively provides a framework for understanding the relevance of ovarian cancer plasticity as a function of two transcription factors., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The experimental evaluation of metastasis overly focuses on the gain of migratory and invasive properties, while disregarding the contributions of cellular plasticity, extra-cellular matrix heterogeneity, niche interactions, and tissue architecture. Traditional cell-based assays often restrict the inclusion of these processes and warrant the implementation of approaches that provide an enhanced spatiotemporal resolution of the metastatic cascade. Time lapse imaging represents such an underutilized approach in cancer biology, especially in the context of disease progression. The inclusion of time lapse microscopy and microfluidic devices in routine assays has recently discerned several nuances of the metastatic cascade. Our review emphasizes that a complete comprehension of metastasis in view of evolving ideologies necessitates (i) the use of appropriate, context-specific assays and understanding their inherent limitations; (ii) cautious derivation of inferences to avoid erroneous/overestimated clinical extrapolations; (iii) corroboration between multiple assay outputs to gauge metastatic potential; and (iv) the development of protocols with improved in situ implications. We further believe that the adoption of improved quantitative approaches in these assays can generate predictive algorithms that may expedite therapeutic strategies targeting metastasis via the development of disease relevant model systems. Such approaches could potentiate the restructuring of the cancer metastasis paradigm through an emphasis on the development of next-generation real-time assays.

Molecular stratification of high-grade serous ovarian carcinoma (HGSC) for targeted therapy is a pertinent approach in improving prognosis of this highly heterogeneous disease. Enabling the same necessitates identification of class-specific biomarkers and their robust detection in the clinic. We have earlier resolved three discrete molecular HGSC classes associated with distinct functional behavior based on their gene expression patterns, biological networks, and pathways. An important difference revealed was that Class 1 is likely to exhibit cooperative cell migration (CCM), Class 2 undergoes epithelial to mesenchymal transition (EMT), while Class 3 is possibly capable of both modes of migration. In the present study, we define clinical stratification of HGSC tumors through the establishment of standard operating procedures for immunohistochemistry and histochemistry based detection of a panel of biomarkers including TCF21, E-cadherin, PARP1, Slug, AnnexinA2, and hyaluronan. Further development and application of scoring guidelines based on expression of this panel in cell line-derived xenografts, commercial tissue microarrays, and patient tumors led to definitive stratification of samples. Biomarker expression was observed to vary significantly between primary and metastatic tumors suggesting class switching during disease progression. Another interesting feature in the study was of enhanced CCM-marker expression in tumors following disease progression and chemotherapy. These stratification principles and the new information thus generated is the first step towards class-specific personalized therapies in the disease.

Cancer metastasis and therapy resistance are the major unsolved clinical challenges, and account for nearly all cancer-related deaths. Both metastasis and therapy resistance are fueled by epithelial plasticity, the reversible phenotypic transitions between epithelial and mesenchymal phenotypes, including epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET). EMT and MET have been largely considered as binary processes, where cells detach from the primary tumor as individual units with many, if not all, traits of a mesenchymal cell (EMT) and then convert back to being epithelial (MET). However, recent studies have demonstrated that cells can metastasize in ways alternative to traditional EMT paradigm; for example, they can detach as clusters, and/or occupy one or more stable hybrid epithelial/mesenchymal (E/M) phenotypes that can be the end point of a transition. Such hybrid E/M cells can integrate various epithelial and mesenchymal traits and markers, facilitating collective cell migration. Furthermore, these hybrid E/M cells may possess higher tumor-initiation and metastatic potential as compared to cells on either end of the EMT spectrum. Here, we review in silico, in vitro, in vivo and clinical evidence for the existence of one or more hybrid E/M phenotype(s) in multiple carcinomas, and discuss their implications in tumor-initiation, tumor relapse, therapy resistance, and metastasis. Together, these studies drive the emerging notion that cells in a hybrid E/M phenotype may occupy 'metastatic sweet spot' in multiple subtypes of carcinomas, and pathways linked to this (these) hybrid E/M state(s) may be relevant as prognostic biomarkers as well as a promising therapeutic targets., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Retinoid and rexinoid receptors are known to regulate key processes during development, differentiation, and cell death in vertebrates. However, their contributions to progression of malignant disease remain largely elusive although it is realized that transformed cancer cells, which essentially evade apoptosis, may display altered molecular expressions or functions associated with retinoid signaling. Here, using a progression model of ovarian cancer, we describe a proteomics-based approach including experimental procedures toward identification and validation of altered protein profiles during transformation. Effectively, this specifies loss of RXR-γ during progression of epithelial ovarian cancer.

Metastatic dissemination generates an aggressive disease facilitated by enhanced migratory and invasive properties. Experimental approaches employ several in vitro and in vivo assays toward quantification of these functionalities. In vitro assessments of cell motility often employ endpoint assays that rely on the global efficacy of wound closure and thwart quantification of migratory phenotypes observed during metastatic dissemination. Recent studies highlight the distinct signatures associated with individual vs. collective cell migration and necessitate the incorporation of these modalities into routine analyses. Advances in live cell imaging that permit real-time visualization of pathophysiological processes can be employed toward elucidating phenotypic plasticity associated with cell migration to overcome caveats inherent to end-point assays. Herein, we corroborate live cell imaging with the in vitro scratch assay toward quantification of migratory modalities in transformed cells. Our protocol describes a step-by-step approach for live cell setup of the scratch assay, and details analyses employed toward definition of three quantitative metrics viz., displacement, velocity and number of nearest neighbors. The current protocol (from scratch induction to data acquisition) is implemented for ~30 h and provides global/single-cell resolution of migratory phenotypes as opposed to the endpoint assays. Routine application of this protocol in cancer biology can aid the design of therapeutic regimes targeting specific migratory modalities and significantly contribute to the dissection of associated molecular networks.

A major focus of contemporary drug screening strategies is the identification of novel anticancer compounds, which often results in underutilization of resources. Current drug evaluation involves in vivo tumor (xenograft) regression as proof-of-principle for cytotoxicity (POC). However, this end-point lacks any assessment of drug resistance of the residual tumor and its capability to establish refractory and/or recurrent disease, which would represent more appropriate indicators of therapeutic failure. We have recently developed a flow cytometry-based approach for the analyses of intra-tumor cellular heterogeneity across stem cell hierarchies, genetic instability and differential cell cycling fractions, which can potentially be predictive of refractory disease and tumor relapse. Iterating this approach after initial POC screening in the drug discovery pipeline would have a great impact in terms of precision of drug evaluation, design of optimal drug combinations and/or drug repositioning. In this perspective, we highlight how through embracing of a comprehensive, informative and analytical assessment of the cellular content of residual tumors, the fidelity and statistical robustness of preclinical drug discovery can be greatly improved., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Intratumor heterogeneity presents a major hurdle in cancer therapy. Most current research studies consider tumors as single entities and overlook molecular diversity between heterogeneous state(s) of different cells assumed to be homogenous. The present approach was designed for fluorescence-activated cell sorting-based resolution of heterogeneity arising from cancer stem cell (CSC) hierarchies and genetic instability in ovarian tumors, followed by microarray-based expression profiling of sorted fractions. Through weighted gene correlation network analyses, we could assign enriched modules of co-regulated genes to each fraction. Such gene modules often correlate with biological functions; one such specific association was the enrichment of CD53 expression in CSCs, functional validation indicated CD53 to be a tumor-initiating cell- rather than quiescent CSC-marker. Another association defined a state of poise for stress-induced metastases in aneuploid cells. Our results thus emphasize the need for studying cell-specific functionalities relevant to regeneration, drug resistance and disease progression in discrete tumor cell fractions.

Tumor heterogeneity and the presence of drug-sensitive and refractory populations within the same tumor are almost never assessed in the drug discovery pipeline. Such incomplete assessment of drugs arising from spatial and temporal tumor cell heterogeneity reflects on their failure in the clinic and considerable wasted costs in the drug discovery pipeline. Here we report the derivation of a flow cytometry-based tumor deconstruction platform for resolution of at least 18 discrete tumor cell fractions. This is achieved through concurrent identification, quantification and analysis of components of cancer stem cell hierarchies, genetically instable clones and differentially cycling populations within a tumor. We also demonstrate such resolution of the tumor cytotype to be a potential value addition in drug screening through definitive cell target identification. Additionally, this real-time definition of intra-tumor heterogeneity provides a convenient, incisive and analytical tool for predicting drug efficacies through profiling perturbations within discrete tumor cell subsets in response to different drugs and candidates. Consequently, possible applications in informed therapeutic monitoring and drug repositioning in personalized cancer therapy would complement rational design of new candidates besides achieving a re-evaluation of existing drugs to derive non-obvious combinations that hold better chances of achieving remission.

Purpose: Resolution of aberrant epigenetic changes leading to altered gene expression during transformation and tumor progression is pertinent for mechanistic understanding of disrupted pathways in cancer. Such changes provide for biomarkers that can be applied in drug screening and improved disease management., Experimental Design: Genome-wide profiling and analyses of promoter DNA methylation, histone modifications, and gene expression of an in vitro progression model of serous ovarian adenocarcinoma were carried out. Similar in silico analyses and comparison of methylation and gene expression of early- and late-grade ovarian cancer samples in The Cancer Genome Atlas assigned a clinical relevance to our study. Candidate biomarkers were evaluated for epigenetic drug treatments in experimental animal models on a background of differing tumor cell responses arising from intratumor heterogeneity., Results: Differentially regulated genes during tumor progression were identified through the previously mentioned analyses as candidate biomarkers. In examining the tumor suppressor PTGIS as a potential biomarker for treatment with either 5-Aza-dC or TSA, 5-Aza-dC effectively stabilized cell cycling, restricted genetic instability, and derepressed PTGIS expression, while TSA led to emergence of drug-resistant progenitors lacking PTGIS expression. Profiling MEST and RXRγ for curcumin and CBB1007, respectively, indicated an inability of curcumin and CBB1007 in restricting residual tumor regenerative capabilities., Conclusions: Our study provides novel insights into epigenetic regulation in ovarian cancer progression and potential biomarkers for evaluating efficacy of epigenetic drugs in restricting residual tumor regeneration. Such approaches may assign a new functional interpretation of drug efficacy and cell tumor responses in ovarian cancer., (©2015 American Association for Cancer Research.)

Slug, a five C2H2 zinc finger (ZF) motif transcription factor mediates cell migration in development, adult tissue repair and regeneration, as well as during tumor metastases through epithelial to mesenchymal transition. At the molecular level, this involves interactions with E-box (CACC/GGTG) consensus elements within target gene promoters to achieve transcriptional repression. However, precise elucidation of events involved in this DNA recognition and binding of specific promoters to regulate target genes have not been achieved. In the present study, we show that besides transcriptional repression, Slug can also directly activate its own expression by preferential binding to specific E-box elements in the distal binding region of its promoter. Our findings suggest that while the first ZF does not contribute to the transcription-associated functions of Slug, all the remaining four ZFs are involved in regulating the expression of target genes with ZF3 and ZF4 being more crucial than ZF2 or ZF5. We also report that recognition and binding preferences of ZFs are defined through intrinsic differences in the E-box core base pairs and/or flanking sequences, with the S2 E-box element being most critical during autoregulation. However, specific target E-box recognition and binding are also defined by the cellular context, which implies that in silico and/or biochemical DNA binding preferences may not necessarily be able to accurately predict in situ events. Our studies thus constitute a novel understanding of transcriptional regulation., (Copyright © 2015. Published by Elsevier B.V.)

The hallmarks of cancer deem biological pathways and molecules to be conserved. This approach may be useful for deriving a prognostic gene signature. Weighted Gene Co-expression Network Analysis of gene expression datasets in eleven cancer types identified modules of highly correlated genes and interactive networks conserved across glioblastoma, breast, ovary, colon, rectal and lung cancers, from which a universal classifier for tumor stratification was extracted. Specific conserved gene modules were validated across different microarray platforms and datasets. Strikingly, preserved genes within these modules defined regulatory networks associated with immune regulation, cell differentiation, metastases, cell migration, metastases, oncogenic transformation, and resistance to apoptosis and senescence, with AIF1 and PRRX1 being suggested to be master regulators governing these biological processes. A universal classifier from these conserved networks enabled execution of common set of principles across different cancers that revealed distinct, differential correlation of biological functions with patient survival in a cancer-specific manner. Correlation analysis further identified a panel of 15 risk genes with potential prognostic value, termed as the GBOCRL-IIPr panel [(GBM-Breast-Ovary-Colon-Rectal-Lung)-Immune-Invasion-Prognosis], that surprisingly, were not amongst the master regulators or important network hubs. This panel may now be integrated in predicting patient outcomes in the six cancers.

Purpose: Tumor heterogeneity and subsistence of high-grade serous ovarian adenocarcinoma (HGSC) classes can be speculated from clinical incidences suggesting passive tumor dissemination versus active invasion and metastases., Experimental Design: We explored this theme toward tumor classification through two approaches of gene expression pattern clustering: (i) derivation of a core set of metastases-associated genes and (ii) resolution of independent weighted correlation networks. Further identification of appropriate cell and xenograft models was carried out for resolution of class-specific biologic functions., Results: Both clustering approaches achieved resolution of three distinct tumor classes, two of which validated in other datasets. Networks of enriched gene modules defined biologic functions of quiescence, cell division-differentiation-lineage commitment, immune evasion, and cross-talk with niche factors. Although deviant from normal homeostatic mechanisms, these class-specific profiles are not totally random. Preliminary validation of these suggests that Class 1 tumors survive, metastasize in an epithelial-mesenchymal transition (EMT)-independent manner, and are associated with a p53 signature, aberrant differentiation, DNA damage, and genetic instability. These features supported by association of cell-specific markers, including PAX8, PEG3, and TCF21, led to the speculation of their origin being the fimbrial fallopian tube epithelium. On the other hand, Class 2 tumors activate extracellular matrix-EMT-driven invasion programs (Slug, SPARC, FN1, THBS2 expression), IFN signaling, and immune evasion, which are prospectively suggestive of ovarian surface epithelium associated wound healing mechanisms. Further validation of these etiologies could define a new therapeutic framework for disease management.

Background: Earlier, proteomic profiling of a Serous Ovarian Carcinoma (SeOvCa) progression model in our lab had identified significantly enriched expression of three double-strand break (DSB) -repair proteins viz. RAD50, NPM1, and XRCC5 in transformed cells over pre-transformed, non-tumorigenic cells. Analysis of the functional relevance of enhanced levels of these proteins was explored in transformed ovarian cancer cells., Methods: Expression profiling, validation and quantitation of the DSB-repair proteins at the transcriptional and protein levels were carried out. Further analyses included identification of their localization, distribution and modulation on exposure to Estradiol (E2) and cisplatin. Effects on silencing of each of these under conditions of genomic-stress were studied with respect to apoptosis, alterations in nuclear morphology and DNA fragmentation; besides profiling known mitotic and spindle check-point markers in DSB-repair., Results: We identified that levels of these DSB-repair proteins were elevated not only in our model, but generally in cancer and are specifically triggered in response to genotoxic stress. Silencing of their expression led to aberrant DSB repair and consequently, p53/p21 mediated apoptosis. Further compromised functionality generated genomic instability., Conclusions: Present study elucidates a functional relevance of NPM1, RAD50 and XRCC5 DSB-repair proteins towards ensuring survival and evasion of apoptosis during ovarian transformation, emphasizing their contribution and association with disease progression in high-grade SeOvCa.

The process of cellular transformation involves cascades of molecular changes that are modulated through altered epigenetic, transcription, post-translational and protein regulatory networks. Thus, identification of transformation-associated protein alterations can provide an insight into major regulatory pathways activated during disease progression. In the present protein expression profiling approach, we identified differential sets of proteins in a two-dimensional gel electrophoresis screen of a serous ovarian adenocarcinoma progression model. Function-based categorization of the proteins exclusively associated with pre-transformed cells identified four cellular processes of which RXR-γ is known to modulate cellular differentiation and apoptosis. We thus probed the functional relevance of RXR-γ expression and signaling in these two pathways during tumor progression. RXR-γ expression was observed to modulate cellular differentiation and apoptosis in steady-state pre-transformed cells. Interestingly, retinoid treatment was found to enhance RXR-γ expression in transformed cells and sensitize them towards apoptosis in vitro, and also reduce growth of xenografts derived from transformed cells. Our findings emphasize that loss of RXR-γ levels appears to provide mechanistic benefits to transformed cells towards the acquisition of resistance to apoptosis hallmark of cancer, while effective retinoid treatment may present a viable approach towards sensitization of tumor cells to apoptosis through induction of RXR-γ expression.

The concept of cancer as a stem cell disease has slowly gained ground over the last decade. A 'stem-like' state essentially necessitates that some cells in the developing tumor express the properties of remaining quiescent, self-renewing and regenerating tumors through establishment of aberrant cellular hierarchies. Alternatively, such capacities may also be reacquired through a de-differentiation process. The abnormal cellular differentiation patterns involved during either process during carcinogenesis are likely to be driven through a combination of genetic events and epigenetic regulation. The role(s) of the latter is increasingly being appreciated in acquiring the requisite genomic specificity and flexibility required for phenotypic plasticity, specifically in a context wherein genome sequences are not altered for differentiation to ensue. In this chapter, the recent advances in elucidating epigenetic mechanisms that govern the self-renewal, differentiation and regenerative potentials of cancer stem cells will be presented.

"Epigenetic plasticity" refers to the capability of mammalian cells to alter their differentiation status via chromatin remodeling-associated alterations in gene expression. While epigenetic plasticity has been best associated with lineage commitment of embryonic stem cells, recent studies have demonstrated chromatin remodeling even in terminally differentiated normal cells, and advanced-stage melanoma and breast cancer cells, in context-dependent responses to alterations in their microenvironment. In the current study, we extend this attribute of epigenetic plasticity to aggressive ovarian cancer cells, by using an integrative approach to associate cellular phenotypes with chromatin modifications ("ChIP-chip") and mRNA and microRNA expression. While we identified numerous gene promoters possessing the well-known "bivalent mark" of H3K27me3/H3K4me2, we also report 14 distinct, lesser-known bi-, tri-, and tetravalent combinations of activating and repressive chromatin modifications, in platinum-resistant CP70 ovarian cancer cells. The vast majority (>90%) of all the histone marks studied localized to regions within 2000 bp of transcription start sites, supporting a role in gene regulation. Upon a simple alteration in the microenvironment, transition from two- to three-dimensional culture, an increase (17% to 38%) in repressive-only marked promoters was observed, concomitant with a decrease (31% to 21%) in multivalent (i.e., juxtaposed permissive and repressive histone marked) promoters. Like embryonic/tissue stem and other (non-ovarian) carcinoma cells, ovarian cancer cell epigenetic plasticity reflects an inherent transcriptional flexibility for context-responsive alterations in phenotype. It is possible that this plasticity could be therapeutically exploited for the management of this lethal gynecologic malignancy.

The isolation and identification of stem-like cells in solid tumors or cancer stem cells (CSCs) have been exciting developments of the last decade, although these rare populations had been earlier identified in leukemia. CSC biology necessitates a detailed delineation of normal stem cell functioning and maintenance of homeostasis within the organ. Ovarian CSC biology has unfortunately not benefited from a pre-established knowledge of stem cell lineage demarcation and functioning in the normal organ. In the absence of such information, some of the classical parameters such as long-term culture-initiating assays to isolate stem cell clones from tumors, screening and evaluation of other epithelial stem cell surface markers, dye efflux, and label retention have been applied toward the putative isolation of CSCs from ovarian tumors. The present review presents an outline of the various approaches developed so far and the various perspectives revealed that are now required to be dealt with toward better disease management.

Multiple, dissimilar genetic defects in cancers of the same origin contribute to heterogeneity in tumor phenotypes and therapeutic responses of patients, yet the associated molecular mechanisms remain elusive. Here, we show at the systems level that serous ovarian carcinoma is marked by the activation of interconnected modules associated with a specific gene set that was derived from three independent tumor-specific gene expression data sets. Network prediction algorithms combined with preestablished protein interaction networks and known functionalities affirmed the importance of genes associated with ovarian cancer as predictive biomarkers, besides "discovering" novel ones purely on the basis of interconnectivity, whose precise involvement remains to be investigated. Copy number alterations and aberrant epigenetic regulation were identified and validated as significant influences on gene expression. More importantly, three functional modules centering on c-Myc activation, altered retinoblastoma signaling, and p53/cell cycle/DNA damage repair pathways have been identified for their involvement in transformation-associated events. Further studies will assign significance to and aid the design of a panel of specific markers predictive of individual- and tumor-specific pathways. In the parlance of this emerging field, such networks of gene-hub interactions may define personalized therapeutic decisions.

Rubinstein-Taybi syndrome (RSTS), a developmental disorder comprising abnormalities that include mental retardation, an unusual facial appearance, broad thumbs and big toes is frequently associated with molecular lesions in the CREB-binding protein gene, CREBBP. The objective of the present study was to identify and analyse CREBBP mutations in Indian RSTS patients on which there are no data. Direct sequencing of CREBBP performed in 13 RSTS patients identified the three zinc fingers (CH1, CH2, CH3) and HAT domain as mutational hotspots in which ten novel pathogenic mutations were localized. Functional analysis revealed that three of these mutations affecting amino acids Glu1459, Leu1668 and Glu1724 were critical for histone acetyltransferase activity. Twenty-eight novel CREBBP single-nucleotide polymorphisms (SNPs) were also identified in the Indian population. Linkage disequilibrium studies revealed associations between (i) SNP (rs129974/c.3836-206G greater than C) and mutation (p.Asp1340Ala); (ii) (rs130002) with mutation (p.Asn435Lys) and (iii) SNPs rs129974, rs130002 and SNP (c.3836-206G greater than C) signifying a disease affection status. In conclusion, the present study reports the highest detection rate of CREBBP mutations (76.9%) in RSTS patients to date, of which ten are predicted to be pathogenic and three critical for histone acetyltransferase activity. Moreover, identification of the association of CREBBP polymorphisms with disease susceptibility could be an important risk factor for the pathogenesis of RSTS.

Biallelic inactivation of the CREB-binding protein (CREBBP) a transcriptional co-activator produces an embryonic lethal phenotype in mice. In humans, re-arrangements in CREBBP are associated with the Rubinstein-Taybi Syndrome (RSTS) that is characterised by craniofacial, skeletal and neuronal symptoms. Neuronal defects in RSTS can be attributed to genetic re-arrangements in CREBBP, which has been implicated in synaptic plasticity and long-term memory. The present study was designed to investigate the role of CREBBP re-arrangements during neuronal differentiation. Towards this, deletion constructs of pCREBBP, viz. pDeltaCB-HAT and pDeltaHAT-CT were generated and transfected into NT2 cells. Expression profiling of the components of Notch, Wnt, SHH and Retinoid signaling along with screening of the neuronal markers was carried out in the NT2 cells and their mutant derivatives. ChIP-PCRs along with co-immunoprecipitations were also performed in these cells to investigate defects due to inappropriate interaction of mutated CREEBP with the corresponding transcription factor and other transcription regulatory proteins both at steady state as well as during differentiation. Mutant NT2 cells lacking the CREB, BROMO and HAT domains (CB-HAT) were highly proliferative and showed limited differentiation; while mutant NT2 cells expressing CREBBP lacking the HAT and CTAD domains (HAT-CT) are proliferation deficient and differentiate rapidly albeit generating an insufficient number of neurons. Altered CREBBP structure resulted in changes in HAT activity, cell cycle profiles and expression of basal levels of components of Notch, SHH, Wnt and retinoid pathways known to be critical in the proliferation and differentiation of neuronal progenitors. At the chromatin level, aberrant signaling correlated with altered binding affinities of the (CREBBP-transcription factor) complexes to promoter regions of components of these pathways. Thus, differentiation defects are manifested early at the genomic level leading to aberrant transcription of the genes involved in differentiation along the neuronal lineage.

DNA methylation and histone modifications often function concomitantly to drive an aberrant program of gene expression in most cancers. Consequently, they have also been identified as being associated with ovarian cancer. However, several basic issues remain unclear - are these marks established early during normal ovarian functioning, or at a preneoplastic stage, or through a gradual accumulation, or do they arise de novo during transformation? Such issues have been difficult to address in ovarian cancer wherein preneoplastic lesions and progression models have not yet been established and drug-refractive disease progression is rapid and aggressive. The review presents an overview of the known involvement of histone modifications in various cellular states that might contribute to our understanding of epithelial ovarian cancer.

Tumor formation involves substantial cell division and genetic instability, but the relationship between quiescent cancer stem cells (CSC) and dividing progenitors in these events is poorly understood. Likewise, the implication of aneuploid cells in solid tumors is uncertain. CSCs are postulated to contribute to tumor dormancy and present a formidable obstacle in limiting treatment outcomes for a majority of cancers, whereas the genetic heterogeneity conjured by aneuploid cells may influence tumor drug resistance. However, direct confirmation of these events remains forthcoming. In the present study, we addressed the identification of tumor dormancy in terms of isolation of therapy-refractory residual tumor cells from tumors that persist in a state of quiescence as label-retaining cells. The choices of label were PKH67/PKH26 dyes that irreversibly bind to the lipid bilayer on cell membranes and get equally partitioned among daughter cells subsequent to each cell division. Consequent characterization revealed that label-retaining cells encompass two different populations capable of remaining in a state of quiescence, i.e., stem-like cells and aneuploid cells. The former express a reversibility of quiescence through retention of functionality and also exhibit therapeutic refractoriness; the latter seem to be either quiescent or proliferation-arrested at steady-state. Subsequent to exposure to selective pressure of chemotherapy, a fraction of these cells may acquire the potential to proliferate in a drug-refractory manner and acquire stem-like characteristics. Collectively, the findings of the present study reveal that tumor-derived CSCs and aneuploid populations contribute to drug resistance and tumor dormancy in cancer progression.

The transcriptional repressors Snail and Slug contribute to cancer progression by mediating epithelial-mesenchymal transition (EMT), which results in tumor cell invasion and metastases. We extend this current understanding to demonstrate their involvement in the development of resistance to radiation and paclitaxel. The process is orchestrated through the acquisition of a novel subset of gene targets that is repressed under conditions of stress, effectively inactivating p53-mediated apoptosis, while another subset of targets continues to mediate EMT. Repressive activities are complemented by a concurrent derepression of specific genes resulting in the acquisition of stem cell-like characteristics. Such cells are bestowed with three critical capabilities, namely EMT, resistance to p53-mediated apoptosis, and a self-renewal program, that together define the functionality and survival of metastatic cancer stem cells. EMT provides a mechanism of escape to a new, less adverse niche; resistance to apoptosis ensures cell survival in conditions of stress in the primary tumor; whereas acquisition of "stemness" ensures generation of the critical tumor mass required for progression of micrometastases to macrometastases. Our findings, besides achieving considerable expansion of the inventory of direct genes targets, more importantly demonstrate that such elegant cooperative modulation of gene regulation mediated by Snail and Slug is critical for a cancer cell to acquire stem cell characteristics toward resisting radiotherapy- or chemotherapy-mediated cellular stress, and this may be a determinative aspect of aggressive cancer metastases.

Recruitment and localization of endothelial precursors within tumors is a potential area for the development of therapeutics, because their functional contribution to tumor vasculature is realized to be important for cancer cell survival. However, the exact nature of the recruited cell type and cellular events orchestrating the entire phenomenon remains obscure. We report that human ovarian cancer is frequently associated with cells expressing the stem cell surface marker CD133. We further show that these CD133-expressing cells are nontumorigenic in nature, and they augment tumor development through their vasculogenic potential. This cell population is attracted by cancer stem cells (CSCs) and retains a direct physical association within the CSC-derived spheroids. Our study further delineates the contribution of these vasculogenic CD133(+) stem cells, termed by us as endothelial stem cells (EnSCs) to the developing tumor vasculature during disease progression. In support of their being stem cells, the EnSCs have a capability of establishing an entire endothelial cell hierarchy. We conclude that such EnSCs play a crucial role in ensuring the development of long-term tumor vasculature to complement CSC-driven tumor development and disease progression.

Aggressive epithelial ovarian cancer (EOC) is genetically and epigenetically distinct from normal ovarian surface epithelial cells (OSE) and early neoplasia. Co-expression of epithelial and mesenchymal markers in EOC suggests an involvement of epithelial-mesenchymal transition (EMT) in cancer initiation and progression. This phenomenon is often associated with acquisition of a stem cell-like phenotype and chemoresistance that correlate with the specific gene expression patterns accompanying transformation, revealing a plasticity of the ovarian cancer cell genome during disease progression.Differential gene expressions between normal and transformed cells reflect the varying mechanisms of regulation including genetic changes like rearrangements within the genome, as well as epigenetic changes such as global genomic hypomethylation with localized promoter CpG island hypermethylation. The similarity of gene expression between ovarian cancer cells and the stem-like ovarian cancer initiating cells (OCIC) are surprisingly also correlated with epigenetic mechanisms of gene regulation in normal stem cells. Both normal and cancer stem cells maintain genetic flexibility by co-placement of activating and/or repressive epigenetic modifications on histone H3. The co-occupancy of such opposing histone marks is believed to maintain gene flexibility and such bivalent histones have been described as being poised for transcriptional activation or epigenetic silencing. The involvement of both-microRNA (miRNA) mediated epigenetic regulation, as well as epigenetic-induced changes in miRNA expression further highlight an additional complexity in cancer stem cell epigenomics.Recent advances in array-based whole-genome/epigenome analyses will continue to further unravel the genomes and epigenomes of cancer and cancer stem cells. In order to illuminate phenotypic signatures that delineate ovarian cancer from their associated cancer stem cells, a priority must lie in the expansion of current technologies and further implementation of bioinformatics to handle the complexity of the cancer epigenome and the various networks that coordinate disease initiation and progression. Great potential lies in the translation of these findings into epigenetic-based therapies. Additionally, targeting chemo-resistant cancer stem cells may provide a much needed breakthrough in treatment of advanced ovarian cancer and chemoresistant disease.

The mechanism of free radical production by complex I deficiency is ill-defined, although it is of significant contemporary interest. This study studied the ROS production and antioxidant defenses in children with mitochondrial NADH dehydrogenase deficiency. ROS production has remained significantly elevated in patients compared to controls. The expression of all antioxidant enzymes significantly increased at mRNA level. However, the enzyme activities did not correlate with high mRNA or protein expression. Only the activity of superoxide dismutase (SOD) was found to correlate with higher mRNA expression in patient derived cell lines. The activities of the enzymes such as glutathione peroxidase (GPx), Catalase (CAT) and glutathione-S-transferase (GST) were significantly reduced in patients (p<0.05 or p<0.01). Glutathione reductase (GR) activity and intracellular glutathione (GSH) levels were not changed. Decreased enzyme activities could be due to post-translational or oxidative modification of ROS scavenging enzymes. The information on the status of ROS and marking the alteration of ROS scavenging enzymes in peripheral lymphocytes or lymphoblast cell lines will provide a better way to design antioxidant therapies for such disorders.

Background: Mitochondrial encephalomyopathies are a heterogeneous group of clinical disorders generally caused due to mutations in either mitochondrial DNA (mtDNA) or nuclear genes encoding oxidative phosphorylation (OXPHOS). We analyzed the mtDNA sequences from a group of 23 pediatric patients with clinical and morphological features of mitochondrial encephalopathies and tried to establish a relationship of identified variants with the disease., Methodology/principle Findings: Complete mitochondrial genomes were amplified by PCR and sequenced by automated DNA sequencing. Sequencing data was analyzed by SeqScape software and also confirmed by BLASTn program. Nucleotide sequences were compared with the revised Cambridge reference sequence (CRS) and sequences present in mitochondrial databases. The data obtained shows that a number of known and novel mtDNA variants were associated with the disease. Most of the non-synonymous variants were heteroplasmic (A4136G, A9194G and T11916A) suggesting their possibility of being pathogenic in nature. Some of the missense variants although homoplasmic were showing changes in highly conserved amino acids (T3394C, T3866C, and G9804A) and were previously identified with diseased conditions. Similarly, two other variants found in tRNA genes (G5783A and C8309T) could alter the secondary structure of Cys-tRNA and Lys-tRNA. Most of the variants occurred in single cases; however, a few occurred in more than one case (e.g. G5783A and A10149T)., Conclusions and Significance: The mtDNA variants identified in this study could be the possible cause of mitochondrial encephalomyopathies with childhood onset in the patient group. Our study further strengthens the pathogenic score of known variants previously reported as provisionally pathogenic in mitochondrial diseases. The novel variants found in the present study can be potential candidates for further investigations to establish the relationship between their incidence and role in expressing the disease phenotype. This study will be useful in genetic diagnosis and counseling of mitochondrial diseases in India as well as worldwide.

Silencing of tumor suppressor genes (TSGs), by DNA methylation, is well known in adult cancers. However, based on the "stem cell" theory of tumorigenesis, the early epigenetic events arising in malignant precursors remain unknown. A recent report demonstrates that, while pluripotent embryonic stem cells lack DNA methylation and possess a "bivalent" pattern of activating and repressive histone marks in numerous TSGs, analogous multipotent malignant cells derived from germ cell tumors (embryonic carcinoma cells) gain additional silencing modifications to those same genes. These results suggest a possible mechanism by which aberrant differentiation, mediated by histone and DNA methylation, instigates tumor progression.

Cancer as a disease driven by cancer stem cells is a concept that has emerged over the last few years. However, several issues relating to this phenomenon as yet remain unaddressed. A fundamental question is one relating to the identification of events leading to transformation of a normal tissue stem cell to a cancer stem cell. Complete knowledge of this evolutionary process may be crucial for the development of novel effective therapies that influence patient prognosis. The scope of this review is to discuss reports that have begun to elucidate stem cell transformation either as an isolated event or as a progression as an attempt towards understanding some of the critical events involved in the process.

Analyses of genome orthologs in cancer on the background of tumor heterogeneity, coupled with the recent identification that the tumor propagating capacity resides within a very small fraction of cells (the tumor stem cells-TSCs), has not been achieved. Here, we describe a strategy to explore genetic drift in the mitochondrial genome accompanying varying stem cell dynamics in epithelial ovarian cancer. A major and novel outcome is the identification of a specific mutant mitochondrial DNA profile associated with the TSC lineage that is drastically different from the germ line profile. This profile, however, is often camouflaged in the primary tumor, and sometimes may not be detected even after metastases, questioning the validity of whole tumor profiling towards determining individual prognosis. Continuing mutagenesis in subsets with a mutant mitochondrial genome could result in transformation through a cooperative effect with nuclear genes - a representative example in our study is a tumor suppressor gene viz. cAMP responsive element binding binding protein. This specific profile could be a critical predisposing step undertaken by a normal stem cell to overcome a tightly regulated mutation rate and DNA repair in its evolution towards tumorigenesis. Our findings suggest that varying stem cell dynamics and mutagenesis define TSC progression that may clinically translate into increasing tumor aggression with serious implications for prognosis.

Widespread applications of stem cell-based therapies in the clinic are being reported, yet there is a surprising lack of resolution of the factors that lead to failure of such therapies on a long-term basis. While classical pharmacogenomics aids the prediction of drug responses in an individual based on genetic variation and pharmacological responses, stem cell therapy involves an additional dimension of host-donor cell interactions and adaptability. We propose the development of concise guidelines based on pharmacogenetic and donor/host-related cellular factors studied more extensively in larger data sets, and used in prospective studies to individualize stem cell therapy.

The cellular mechanisms underlying the increasing aggressiveness associated with ovarian cancer progression are poorly understood. Coupled with a lack of identification of specific markers that could aid early diagnoses, the disease becomes a major cause of cancer-related mortality in women. Here we present direct evidence that the aggressiveness of human ovarian cancer may be a result of transformation and dysfunction of stem cells in the ovary. A single tumorigenic clone was isolated among a mixed population of cells derived from the ascites of a patient with advanced ovarian cancer. During the course of the study, yet another clone underwent spontaneous transformation in culture, providing a model of disease progression. Both the transformed clones possess stem cell-like characteristics and differentiate to grow in an anchorage-independent manner in vitro as spheroids, although further maturation and tissue-specific differentiation was arrested. Significantly, tumors established from these clones in animal models are similar to those in the human disease in their histopathology and cell architecture. Furthermore, the tumorigenic clones, even on serial transplantation continue to establish tumors, thereby confirming their identity as tumor stem cells. These findings suggest that: (a) stem cell transformation can be the underlying cause of ovarian cancer and (b) continuing stochastic events of stem and progenitor cell transformation define the increasing aggression that is characteristically associated with the disease.

Objectives: The transcriptional factors Snail and Slug have been reported to be important in cell migration during development and also during tumor metastasis. Their expression and role in ovarian cancer, hitherto unexplored, was examined to understand the molecular events in ovarian cancer metastases since the latter is responsible for the high degree of mortality associated with the disease., Methods: Ectopic expression of mSnail and mSlug in the epithelial ovarian cancer cell line SKOV3 was carried out and stable clones were selected. These were used to examine specific repression of the adherens, tight and desmosomal junction components by the two transcription factors. Furthermore, functional implications with respect to enhanced migration of cells, tumorigenecity and metastasis were also studied., Results: The ectopic expression of Snail or Slug resulted in epithelial-mesenchymal transition (EMT), enhanced motility, invasiveness and tumorigenecity in the cell line SKOV3. In addressing the mechanism by which Snail and Slug lead to loss of intercellular adhesion, specific repression of adherens junction components (E-cadherin and betacatenin), tight junction components (Occludin and ZO-1) and desmosomal junction components (Dsg2) were observed. Snail suppresses expression of adherens and tight junction components, while Slug suppresses expression of all the three junction components; concertedly, bringing down the intercellular adhesion between cells. Further activation of these transcriptional factors in hypoxic conditions revealed a rapid upregulation of Slug expression as an immediate reaction that probably triggers off a signaling cascade leading to Snail expression., Conclusions: These results indicate distinct roles of the transcriptional factors Snail and Slug during ovarian cancer metastasis and cell survival through mediation of EMT.

Therapeutic stem cell applications represent a newly evolving approach for the treatment of several genetic and degenerative diseases. The advent of pharmacogenomics too, holds promise for an individualized, optimal treatment regime for a large variety of medical conditions. A combination of the benefits of these two technologies creates a new niche in therapeutic medicine research viz. that of stem cell pharmacogenomics (SCP). The development of this approach requires the application of existing technologies in genomics, proteomics and bioinformatics to resolve the various issues involved in advancing the therapeutic applications of stem cell medicine. In this brief overview of the subject, we attempt to provide fresh insights into the exclusive niche of stem cell pharmacogenomics and discuss some of the priority issues that need to be targeted, based on the existing principles of pharmacogenomics, stem cell characteristics and transplantation medicine. Advances in these areas are imperative in realizing the dream of stem cell therapies contributing towards the improvisation of the quality of human life.

Immature embryos of thirty-three genotypes of wheat were cultured on 2,4-D containing medium. Occurrence of precocious germination of the zygotic and somatic embryos simultaneously on the same medium was a striking feature observed during the course of work. The percentage of precocious germination was seen to vary extensively from 0-88% and 0-84% for zygotic and somatic embryos respectively. In the genotypes NI-5439 and NI-5643 which are characterized by a high tillering capacity, the phenomenon of precocious germination seems to take a different path from that observed in the other genotypes. This is evident since these two genotypes require total absence of hormone for shoot elongation although multiple shoot primordia are formed on auxin containing medium.Precocious germination also seems to be relevant to somatic embryogenesis and plantlet regeneration. This conclusion stems from the observation that a majority of the genotypes that show precocious germination of zygotic embryos have greater embryogenic potential. Consecutively, most of the genotypes that show precocious germination of somatic embryos exhibit a higher frequency and faster rate of plantlet regeneration.