Identification of a Vitamin-D Receptor Antagonist, MeTC7, which Inhibits the Growth of Xenograft and Transgenic Tumors In Vivo

Vitamin-D receptor (VDR) mRNA is overexpressed in neuroblastoma and carcinomas of lung, pancreas, and ovaries and predicts poor prognoses. VDR antagonists may be able to inhibit tumors that overexpress VDR. However, the current antagonists are arduous to synthesize and are only partial antagonists, limiting their use. Here, we show that the VDR antagonist MeTC7 (5), which can be synthesized from 7-dehydrocholesterol (6) in two steps, inhibits VDR selectively, suppresses the viability of cancer cell-lines, and reduces the growth of the spontaneous transgenic TH-MYCN neuroblastoma and xenografts in vivo. The VDR selectivity of 5 against RXRα and PPAR-γ was confirmed, and docking studies using VDR-LBD indicated that 5 induces major changes in the binding motifs, which potentially result in VDR antagonistic effects. These data highlight the therapeutic benefits of targeting VDR for the treatment of malignancies and demonstrate the creation of selective VDR antagonists that are easy to synthesize

Molecular Biomarker Analyses Using Circulating Tumor Cells

Evaluation of cancer biomarkers from blood could significantly enable biomarker assessment by providing a relatively non-invasive source of representative tumor material. Circulating Tumor Cells (CTCs) isolated from blood of metastatic cancer patients hold significant promise in this regard.Using spiked tumor-cells we evaluated CTC capture on different CTC technology platforms, including CellSearch and two biochip platforms, and used the isolated CTCs to develop and optimize assays for molecular characterization of CTCs. We report similar performance for the various platforms tested in capturing CTCs, and find that capture efficiency is dependent on the level of EpCAM expression. We demonstrate that captured CTCs are amenable to biomarker analyses such as HER2 status, qRT-PCR for breast cancer subtype markers, KRAS mutation detection, and EGFR staining by immunofluorescence (IF). We quantify cell surface expression of EGFR in metastatic lung cancer patient samples. In addition, we determined HER2 status by IF and FISH in CTCs from metastatic breast cancer patients. In the majority of patients (89%) we found concordance with HER2 status from patient tumor tissue, though in a subset of patients (11%), HER2 status in CTCs differed from that observed in the primary tumor. Surprisingly, we found CTC counts to be higher in ER+ patients in comparison to HER2+ and triple negative patients, which could be explained by low EpCAM expression and a more mesenchymal phenotype of tumors belonging to the basal-like molecular subtype of breast cancer.Our data suggests that molecular characterization from captured CTCs is possible and can potentially provide real-time information on biomarker status. In this regard, CTCs hold significant promise as a source of tumor material to facilitate clinical biomarker evaluation. However, limitations exist from a purely EpCAM based capture system and addition of antibodies to mesenchymal markers could further improve CTC capture efficiency to enable routine biomarker analysis from CTCs

Molecular cytogenetic analysis and gene amplification in rhabdomyosarcoma and neuroblastoma

grantor: University of TorontoAn understanding of the genetic changes that lead to malignancy is of fundamental importance in the study of carcinogenesis. As tumors progress, multiple genetic aberrations are accumulated which often correlate with biological and clinical behavior. Recent advances in cancer cytogenetics can provide a greater depth of information regarding chromosomal changes in tumors. This thesis addresses the molecular genetic consequences of chromosomal and DNA copy number alterations in two pediatric neoplasms: rhabdomyosarcoma and neuroblastoma. Rhabdomyosarcoma predominantly presents as two major subtypes: embryonal RMS (RMS-E) and alveolar RMS (RMS-A). Comparative genomic hybridization performed on 21 RMSs identified distinct differences between the two subtypes. Whole chromosomal gains of 2, 5, 7, 8, 11 and 12 were observed in RMS-E, and high copy number amplification, involving the 2p24 region (6/12 tumors) and less frequently the 12q13-21, 9p22 and 17q22-25 regions was detected in RMS-A. Patients with amplification had a shorter disease free interval and a higher mortality. Spectral karyotype analysis of a RMS cell line of each subtype identified a surprisingly high level of structural change. Gene expression studies using the Atlas Human Cancer Array (588 genes) showed that 45 genes showed differential expression. Currently, very little information is available on the evolution of ' MYCN' amplicon as either double minute chromosomes (dmins) or homogeneously staining regions (hsrs). Free chromatin fluorescence in situ hybridization was therefore performed to provide relational mapping of 'MYCN' and 'DDX1' within band 2p24 and to analyze the configuration of the two genes in dmin and hsr. 'DDX1' was found to be telomeric to 'MYCN' and no regular reiterated amplicon repeat unit was present in hsrs. A more detailed analysis of the configurations of ' DDX1' and 'MYCN' within each array detected a complex pattern of amplicons within hsrs suggesting that multiple rearrangements must have taken place. Analysis of dmins showed a simpler pattern of amplicon structure suggesting that deletions and/or duplications of 'MYCN' and ' DDX1' may have occurred. These data are consistent with a molecular mechanism for amplification involving many rearrangements as it develops resulting in complex amplicon structures leading to changes in both relative gene copy number and in the intragenic distances between co-amplified genes.Ph.D

A study on morbidity pattern among geriatric population of an urban slum, Dehradun, India

Background: Ageing is a process of deterioration in the individual’s functional capacity that results from structural changes as age advances. India has acquired the label of “an ageing nation” with 7.7% of its population being more than 60 years old. Research on geriatric morbidity and related risk factors are required to improve the delivery of health care to the elderly. This study was an attempt to study the morbidity status of geriatric people that may serve as a baseline data and also help in planning the health services. Aims and Objectives: To ascertain determinants and predictors as potential socio-epidemiological correlates of the prevalent morbidity pattern and recommend appropriate measures / interventions to address felt and unmet needs in geriatric health. Material and Methods: An observational study conducted in field practice area of Department of Community Medicine, Himalayan Institute of Medical Sciences, HIHT, Dehradun. Study participants aged above 60 years were administered by pre tested, semi structured questionnaire after obtained informed consent. Descriptive statistics were calculated by using SPSS version 17. Results: Among 520 subjects most common morbidity in study subjects was arthritis (49.61%) followed by cataract (46.34%), chronic gastritis (44.23%), COPD (25%), diabetes (21.15%), skin lesions (19.03%), hypertension (11.39%). Conclusion: There is high morbidity rate identified in this present study so there is an urgent need to develop geriatric health care services at primary health centre leve

Malignant and benign ganglioglioma: A pathological and molecular study1

Gangliogliomas are generally benign tumors, composed of transformed neuronal and glial elements, with rare malignant progression of the glial component. The current study of a rare case of a woman harboring a ganglioglioma with areas of malignant transformation addresses two fundamental questions: (1) Are the ganglioglioma and its malignant component clonal in origin? (2) What are the genetic alterations associated with the initiation and subsequent malignant progression of ganglioglioma? By using the human androgen receptor gene (HUMARA) assay, we found the ganglioglioma and the malignant component to be clonal in origin, suggestive of initial transformation of a single neuroglial precursor cell with subsequent malignant progression. Conventional and array comparative genomic hybridization (approximately 2.5-Mb resolution) analyses found chromosomal losses to be predominant in the benign areas of the ganglioglioma, with gains more prevalent in the malignant component. Regions of chromosomal loss, postulated to harbor genes involved in the initiation of ganglioglioma, included 1p35-36, 2p16-15, 3q13.1-13.3, 3q24-25.3, 6p21.3-21.2, 6q24-25.2, 9p12, Xp11.3-11.22, and Xq22.1-22.3. Direct analysis demonstrated loss of p19 expression and p53 mutation in the malignant areas, highly suggestive of these alterations being involved in the malignant progression of the ganglioglioma. Additional chromosomal alterations specific to the malignancy involved gains on 1p35-34.2, 2q24.1-32.3, 3q13.1-13.3, 6q13-16.2, 7q11.2-31.3, 8q21.1-23, 11q12-31, and 12q13.2-21.3. This molecular-pathological study has provided insight into the pathogenesis of gangliogliomas and associated rare malignant progression. Deciphering the specific genes residing in these chromosomal regions may further our understanding of not only these rare tumors but also the more common gliomas

Application of Comparative Genomic Hybridization, Spectral Karyotyping, and Microarray Analysis in the Identification of Subtype-Specific Patterns of Genomic Changes in Rhabdomyosarcoma

Rhabdomyosarcoma (RMS) in children occurs predominantly as two major histologically defined subtypes called embryonal RMS (RMS-E) and the prognostically less favorable alveolar RMS (RMS-A). Comparative genomic hybridization (CGH) was performed on 21 RMS and identified consistent gains affecting chromosomes 2 (8/10), 5 (5/10), 6 (3/10), 7 (7/10), 8 (9/10), 11 (6/10), and 12 (5/10) in RMS-E. Losses/deletions involved chromosomes 19 (2/10) and chromosomes 4, 9, 10, 17, 21 (1/10 each). High copy number amplification, involving the 2p24 region (5/11) and less frequently, the 12813-21 (2/11), 9p22 (1/11), and 17822-25 (1/11) regions, was detected in RMS-A. Gene amplification at band 2p24 was present in 6/12 alveolar tumors, and in each case, MYCN was amplified, together with the distally placed DDX1 gene. For these patients there was a shorter disease free interval and a higher mortality than patients with tumors without amplification. Detailed spectral karyotype analysis (SKY) was performed on two RMS cell lines (one of each subtype) and identified a surprisingly high level of structural change. Gene expression studies with the Atlas Human Cancer Array (588 genes) showed that 153 genes generated a signal of similar intensity in both cell lines, and 45 genes appeared to have subtype-specific expression. The chromosomal location of differentially expressed genes was compared to the pattern of genomic alteration in RMS as determined by CGH in this study and the literature