Biomedical subjects

Nigel Clegg

Publications and source records attributed to Nigel Clegg.

8 recordsLinked to original sources

PubMed2006

Combining results of microarray experiments: a rank aggregation approach.

As technology for microarray analysis becomes widespread, it is becoming increasingly important to be able to compare and combine the results of experiments that explore the same scientific question. In this article, we present a rank-aggregation approach for combining results from several microarray studies. The motivation for this approach is twofold; first, the final results of microarray studies are typically expressed as lists of genes, rank-ordered by a measure of the strength of evidence that they are functionally involved in the disease process, and second, using the information on this rank-ordered metric means that we do not have to concern ourselves with data on the actual expression levels, which may not be comparable across experiments. Our approach draws on methods for combining top-k lists from the computer science literature on meta-search. The meta-search problem shares several important features with that of combining microarray experiments, including the fact that there are typically few lists with many elements and the elements may not be common to all lists. We implement two meta-search algorithms, which use a Markov chain framework to convert pairwise preferences between list elements into a stationary distribution that represents an aggregate ranking (Dwork et al, 2001). We explore the behavior of the algorithms in hypothetical examples and a simulated dataset and compare their performance with that of an algorithm based on the order-statistics model of Thurstone (Thurstone, 1927). We apply all three algorithms to aggregate the results of five microarray studies of prostate cancer.

Algorithms↗

PubMed2005

Substrates of the prostate-specific serine protease prostase/KLK4 defined by positional-scanning peptide libraries.

BACKGROUND: Prostase/KLK4 is a member of the human kallikrein (KLK) gene family that is expressed in prostate epithelial cells under the regulation of androgenic hormones. In this study, we sought to characterize the substrate specificity of KLK4 in order to gain insight into potential physiological roles of the enzyme. METHODS: A chimeric form of KLK4 was constructed in which the pro-region of KLK4 was replaced with the signal and propeptide sequence of trypsinogen (proT-KLK4) to create an activation site susceptible to enterokinase cleavage. proT-KLK4 was expressed in Drosophila S2 cells, purified, and activated with enterokinase to generate mature KLK4. The extended substrate specificity of KLK4 was defined by screening tetrapeptide positional scanning synthetic combinatorial libraries (PS-SCL). RESULTS: The preferred P1-P4 positions as determined by PS-SCL were: P1-Arg; P2-Gln/Leu/Val; P3-Gln/Ser/Val; P4-Ile/Val. The trypsin-like specificity of KLK4 was further confirmed using synthetic chromogenic peptides. Based upon the optimal cleavage site residues, a database search for potential KLK4 substrates identified several proteins with potential roles mediating normal prostate physiology or neoplastic growth including KLK3/PSA, parathyroid hormone-related peptide (PTHrP), and members of the bone morphogenetic protein (BMP) family. Recombinant KLK4 was able to activate pro-PSA/KLK3 and degrade members of the insulin-like growth factor (IGF) binding protein (IGFBP) family. CONCLUSIONS: These results identify potential KLK4 substrates that may serve to define the role of this protease in normal prostate physiology, and facilitate studies of the consequences of KLK4 expression in pathological conditions.

Animals↗

PubMed2004

Characterization and comparative analyses of transcriptomes from the normal and neoplastic human prostate.

BACKGROUND: The prostate gland is a highly specialized organ with functional attributes that serve to enhance the fertility of mammalian species. Pathological processes affecting the prostate include benign prostate hypertrophy and prostate carcinoma; diseases that account for major morbidity and mortality in middle-aged and elderly men. To facilitate studies of biological processes uniquely represented in the prostate and assess molecular alterations associated with prostate carcinoma, we sought to establish the diversity of gene expression in the normal and neoplastic prostate through the compilation and analysis of a prostate transcriptome. METHODS: We assembled and annotated ESTs derived from prostate cDNA libraries that were either produced in our laboratory or available from public sequence repositories such as CGAP, dbEST, and Unigene. Determinations of differential gene expression between the normal prostate, other normal tissues, and neoplastic prostate tissues was performed using statistical algorithms. Confirmation of differential expression was performed by quantitative PCR and Northern analysis. RESULTS: A total of 99,448 high-quality ESTs were assembled and annotated to produce a prostate transcriptome comprised of 24,580 distinct TUs. Comparative analyses of gene expression levels identified 61 TUs with exclusive expression in the prostate and 45 TUs with high levels of expression in the prostate relative to at least 25 other normal tissues (P > 0.99). Comparative analyses of ESTs derived from neoplastic prostate tissues identified 75 genes with dysregulated expression in cancer (P > 0.99). CONCLUSIONS: The human prostate expresses a diverse repertoire of genes that reflect a functionally complex organ. The identification of genes with prostate-restricted or enhanced expression may provide additional insights into the biochemical processes that interact to form the developmental, signaling, and metabolic pathways of the normal and neoplastic gland.

Algorithms↗

PubMed2003

Molecular characterization of prostatic small-cell neuroendocrine carcinoma.

OBJECTIVES: A subset of prostate carcinomas is composed predominantly, even exclusively, of neuroendocrine (NE) cells. In this report, we sought to characterize the gene expression profile of a prostate small cell NE carcinoma by assessing the diversity and abundance of transcripts in the LuCaP 49 prostate small cell carcinoma xenograft. METHODS: We constructed a cDNA library (PRCA3) from the LuCap 49 prostate small cell xenograft. Single pass DNA sequencing of randomly selected cDNA clones followed by sequence assembly and annotation produced a library of Expressed Sequence Tags (ESTs) representing the LuCaP 49 transcriptome. Comparative sequence analysis with ESTs derived from prostate adenocarcinoma libraries was performed using statistical algorithms designed to identify differentially expressed sequences. Putative NE cell-specific genes were further examined by Northern analysis. RESULTS: Sequence assembly and analysis identified 1,447 distinct genes expressed in the LuCaP 49 cDNA library. These include cDNAs encoding the NE markers secretogranin (SCG2), CD24, and ENO2. Northern analysis revealed that three additional genes, ASCL1, INA, and SV2B are expressed in LuCaP 49 but not in various prostate cancer cell lines or xenografts. Fifteen genes were identified with a statistical probability (P > 0.9) of being up-regulated in LuCaP 49 small cell carcinoma relative to prostate adenocarcinoma (two primary prostate adenocarcinomas and the LNCaP prostate adenocarcinoma cell line). CONCLUSIONS: Prostate small cell carcinoma expresses a diverse repertoire of genes that reflect characteristics of their NE cell of origin. ASCL1, INA, and SV2B are potential molecular markers for small cell NE tumors and NE cells of the prostate. This small cell NE carcinoma gene expression profile may yield insights into the development, progression, and treatment of subtypes of prostate cancer.

Aged↗

PubMed2002

The program of androgen-responsive genes in neoplastic prostate epithelium.

The human prostate gland is an important target organ of androgenic hormones. Testosterone and dihydrotestosterone interact with the androgen receptor to regulate vital aspects of prostate growth and function including cellular proliferation, differentiation, apoptosis, metabolism, and secretory activity. Our objective in this study was to characterize the temporal program of transcription that reflects the cellular response to androgens and to identify specific androgen-regulated genes (ARGs) or gene networks that participate in these responses. We used cDNA microarrays representing about 20,000 distinct human genes to profile androgen-responsive transcripts in the LNCaP adenocarcinoma cell line and identified 146 genes with transcript alterations more than 3-fold. Of these, 103 encode proteins with described functional roles, and 43 represent transcripts that have yet to be characterized. Temporal gene expression profiles grouped the ARGs into four distinct cohorts. Five uncharacterized ARGs demonstrated exclusive or high expression levels in the prostate relative to other tissues studied. A search of available DNA sequence upstream of 28 ARGs identified 25 with homology to the androgen response-element consensus-binding motif. These results identify previously uncharacterized and unsuspected genes whose expression levels are directly or indirectly regulated by androgens; further, they provide a comprehensive temporal view of the transcriptional program of human androgen-responsive cells.

Amino Acid Motifs↗

PubMed2002

The human (PEDB) and mouse (mPEDB) Prostate Expression Databases.

The Prostate Expression Databases (PEDB and mPEDB) are online resources designed to allow researchers to access and analyze gene expression information derived from the human and murine prostate, respectively. Human PEDB archives more than 84 000 Expressed Sequence Tags (ESTs) from 38 prostate cDNA libraries in a curated relational database that provides detailed library information including tissue source, library construction methods, sequence diversity and sequence abundance. The differential expression of each EST species can be viewed across all libraries using a Virtual Expression Analysis Tool (VEAT), a graphical user interface written in Java for intra- and inter-library sequence comparisons. Recent enhancements to PEDB include (i) the development of a murine prostate expression database, mPEDB, that complements the human gene expression information in PEDB, (ii) the assembly of a non-redundant sequence set or 'prostate unigene' that represents the diversity of gene expression in the prostate, and (iii) an expanded search tool that supports both text-based and BLAST queries. PEDB and mPEDB are accessible via the World Wide Web at http://www.pedb.org and http://www.mpedb.org.

Animals↗

PubMed2002

A neuroendocrine/small cell prostate carcinoma xenograft-LuCaP 49.

The late stages of progression of prostate carcinoma are typically characterized by an androgen-insensitive, rapidly proliferative state. Some late-stage tumors are composed predominantly of neuroendocrine cells. Virtually no animal models of a neuroendocrine/small cell variant of prostate carcinoma are available for experimental studies. We report a human neuroendocrine/small cell prostate carcinoma xenograft that was developed from a nodal metastasis of a human prostate carcinoma and that has been propagated as serial subcutaneous implants in severe combined immunodeficient mice for >4 years. Designated LuCaP 49, all tumor passages exhibit a neuroendocrine/small cell carcinoma phenotype-insensitivity to androgen deprivation, expression of neuroendocrine proteins, lack of expression of prostate-specific antigen or androgen receptor, and an unusually rapid growth (a doubling time of 6.5 days) for prostate cancer xenografts. Genetically this tumor exhibits loss of heterozygosity for the short arm of chromosome 8 and has a complex karyotype. This xenograft should prove to be useful in the investigation of mechanisms underlying the androgen-insensitive state of progressive prostate carcinoma.

Aged↗

PubMed2002

Digital expression profiles of the prostate androgen-response program.

The androgen receptor (AR) and cognate ligands regulate vital aspects of prostate cellular growth and function including proliferation, differentiation, apoptosis, lipid metabolism, and secretory action. In addition, the AR pathway also influences pathological processes of the prostate such as benign prostatic hypertrophy and prostate carcinogenesis. The pivotal role of androgens and the AR in prostate biology prompted this study with the objective of identifying molecular mediators of androgen action. Our approach was designed to compare transcriptomes of the LNCaP prostate cancer cell line under conditions of androgen depletion and androgen stimulation by generating and comparing collections of expressed sequence tags (ESTs). A total of 4400 ESTs were produced from LNCaP cDNA libraries and these ESTs assembled into 2486 distinct transcripts. Rigorous statistical analysis of the expression profiles indicated that 17 genes exhibited a high probability (P>0.9) of androgen-regulated expression. Northern analysis confirmed that the expression of KLK3/PSA, FKBP5, KRT18, DKFZP564K247, DDX15, and HSP90 is regulated by androgen exposure. Of these, only KLK3/PSA is known to be androgen-regulated while the other genes represent new members of the androgen-response program in prostate epithelium. LNCaP gene expression profiles defined by two independent experiments using the serial analysis of gene expression (SAGE) method were compared with the EST profiles. Distinctly different expression patterns were produced from each dataset. These results are indicative of the sensitivity of the methods to experimental conditions and demonstrate the power and the statistical limitations of digital expression analyses.

Adenocarcinoma↗