Papers by Hosein Kouros-Mehr
Trends in Molecular Medicine, 2010
Several ongoing clinical studies are designed to test the efficacy of antiangiogenic therapies in... more Several ongoing clinical studies are designed to test the efficacy of antiangiogenic therapies in the adjuvant setting, where the goal is to increase the cure rate in patients who have just undergone surgical resection of all visible disease. Tumors depend on angiogenesis to support their growth and progression and blockade of this process has proven to be a valid strategy for treating multiple types of advanced metastatic cancer. However, results from the first of these clinical adjuvant studies were disappointing, stimulating extensive debate as to the potential of this approach. It will require additional clinical studies before we realize whether the effects of angiogenic blockade are durable, and if they are able to cure a subset of patients with early stage cancer.
Chemical Senses, 2001
In mammals, the vomeronasal organ (VNO) contains chemosensory receptor cells that bind to pheromo... more In mammals, the vomeronasal organ (VNO) contains chemosensory receptor cells that bind to pheromones and induce a variety of social and reproductive behaviors. It has been traditionally assumed that the human VNO (Jacobson's organ) is a vestigial structure, although recent studies have shown minor evidence for a structurally intact and possibly functional VNO. The presence and function of the human VNO remains controversial, however, as pheromones and VNO receptors have not been well characterized. In this study we screened a human Bacterial Artificial Chromosome (BAC) library with multiple primer sets designed from human cDNA sequences homologous to mouse VNO receptor genes. Utilizing these BAC sequences in addition to mouse VNO receptor sequences, we screened the High Throughput Genome Sequence (HTGS) database to find additional human putative VNO receptor genes. We report the identification of 56 BACs carrying 34 distinct putative VNO receptor gene sequences, all of which appear to be pseudogenes. Sequence analysis indicates substantial homology to mouse V1R and V2R VNO receptor families. Furthermore, chromosomal localization via FISH analysis and RH mapping reveal that the majority of the BACs are localized to telomeric and centromeric chromosomal localizations and may have arisen through duplication events. These data yield insight into the present state of pheromonal olfaction in humans and into the evolutionary history of human VNO receptors. (Sequence data from this article have been deposited with the DDBJ/EMBL/Genbank Data Libraries under accession nos.
Dna Research, 2000
Heterotrimeric guanine nucleotide binding proteins (G proteins) transduce extracellular signals r... more Heterotrimeric guanine nucleotide binding proteins (G proteins) transduce extracellular signals received by transmembrane receptors to effector proteins. Each subunit of the G protein complex is encoded by a member of one of three corresponding gene families. Currently, 16 different members of the a subunit family, 5 different members of the (3 subunit family, and 11 different members of the 7 subunit family have been described in mammals. Here we have identified and characterized Bacterial Artificial Chromosomes (BACs) containing the human homologs of each of the a, (3, and 7 subunit genes as well as a Gall pseudogene and a previously undiscovered G75-like gene. The gene structure and chromosome location of each gene was determined, as were the orientations of paired genes. These results provide greater insight into the evolution and functional diversity of the mammalian G protein subunit genes.
Current Opinion in Cell Biology, 2008
The GATA family of transcription factors plays essential roles in the specification and maintenan... more The GATA family of transcription factors plays essential roles in the specification and maintenance of differentiated cell types. GATA-3 was identified in a microarray screen of the mouse mammary gland as the most highly expressed transcription factor in the mammary epithelium and is expressed exclusively in the luminal epithelial cell population. Targeted deletion of GATA-3 in mammary glands leads to profound defects in mammary development and inability to specify and maintain the luminal cell fate in the adult mouse. In breast cancer, GATA-3 has emerged as a strong and independent predictor of tumor differentiation, estrogen receptor status and clinical outcome. GATA-3 maintains tumor differentiation and suppresses tumor dissemination in a mouse model of breast cancer. This review explores our current understanding of GATA-3 signaling in luminal cell differentiation, both in mammary development and breast cancer.
Cell, 2006
The GATA family of transcription factors plays fundamental roles in cell-fate specification. Howe... more The GATA family of transcription factors plays fundamental roles in cell-fate specification. However, it is unclear if these genes are necessary for the maintenance of cellular differentiation after development. We identified GATA-3 as the most highly enriched transcription factor in the mammary epithelium of pubertal mice. GATA-3 was found in the luminal cells of mammary ducts and the body cells of terminal end buds (TEBs). Upon conditional deletion of GATA-3, mice exhibited severe defects in mammary development due to failure in TEB formation during puberty. After acute GATA-3 loss, adult mice exhibited undifferentiated luminal cell expansion with basement-membrane detachment, which led to caspase-mediated cell death in the long term. Further, FOXA1 was identified as a downstream target of GATA-3 in the mammary gland. This suggests that GATA-3 actively maintains luminal epithelial differentiation in the adult mammary gland, which raises important implications for the pathogenesis of breast cancer.
Development, 2005
Epithelial-mesenchymal crosstalk is essential for tissue morphogenesis, but incompletely understo... more Epithelial-mesenchymal crosstalk is essential for tissue morphogenesis, but incompletely understood. Postnatal mammary gland development requires epidermal growth factor receptor (EGFR) and its ligand amphiregulin (AREG), which generally must be cleaved from its transmembrane form in order to function. As the transmembrane metalloproteinase ADAM17 can process AREG in culture and Adam17 −/− mice tend to phenocopy Egfr −/− mice, we examined the role of each of these molecules in mammary development. Tissue recombination and transplantation studies revealed that EGFR phosphorylation and ductal development occur only when ADAM17 and AREG are expressed on mammary epithelial cells, whereas EGFR is required stromally, and that local AREG administration can rescue Adam17 −/− transplants. Several EGFR agonists also stimulated Adam17 −/− mammary organoid growth in culture, but only AREG was expressed abundantly in the developing ductal system in vivo. Thus, ADAM17 plays a crucial role in mammary morphogenesis by releasing AREG from mammary epithelial cells, thereby eliciting paracrine activation of stromal EGFR and reciprocal responses that regulate mammary epithelial development.
Differentiation, 2006
Unlike other branched organs, the mammary gland undergoes most of its branching during adolescent... more Unlike other branched organs, the mammary gland undergoes most of its branching during adolescent rather than embryonic development. Its morphogenesis begins in utero, pauses between birth and puberty, and resumes in response to ovarian estrogens to form an open ductal tree that eventually fills the entire mammary fat pad of the young female adult. Importantly, this ''open'' architecture leaves room during pregnancy for the organ to develop milk-producing alveoli like leaves on otherwise bare branches. Thereafter, the ducts serve to deliver the milk that is produced throughout lactation. The hormonal cues that elicit these various phases of mammary development utilize local signaling cascades and reciprocal stromal-epithelial interactions to orchestrate the tissue reorganization, differentiation and specific activities that define each phase. Fortunately, the mammary gland is rather amenable to experimental inquiry and, as a result, we have a fair, although incomplete, understanding of the mechanisms that control its development. This review discusses our current sense and understanding of those mechanisms as they pertain to mammary branching, with the caveat that many more aspects are still waiting to be solved.
Developmental Dynamics, 2006
The mammary gland develops in a process known as branching morphogenesis, whereby a distal epithe... more The mammary gland develops in a process known as branching morphogenesis, whereby a distal epithelial bud extends and bifurcates to form an extensive ductal network. Compared with other branched organs, such as the lung and kidney, little is known about the molecular basis of branching in the mammary gland. Here we report a microarray profiling strategy to identify novel genes that may regulate mammary branching. We microdissected terminal end bud (TEB) and mature duct microenvironments from β-actin-green fluorescent protein reporter mice and compared their RNA expression profiles with epithelium-free mammary stroma by means of microarray. We identified 1,074 genes enriched in the TEB microenvironment, 222 genes enriched in the mature duct microenvironment, and 385 genes enriched in both TEB and mature duct microenvironments. The microarray correctly predicted the expression of genes known to be enriched in the epithelium (Ets-5) and stroma (MMP-14) of TEBs and in the mature duct microenvironment (MMP-3). The microarray also correctly predicted the localization of previously uncharacterized genes, such as the TEBenriched SPRR-1a, the duct-enriched casein-γ, and the general epithelial marker pleiotrophin. Analysis of genes enriched in TEBs revealed several genes in the Wnt (Wnt-2, Wnt-5a, Wnt-7b, Dsh-3, Frizzled-1, Frizzled-2), hedgehog (Dhh), ephrin (Ephrin-B1, Eph-A2), and transcription factor (Twist-1, Twist-2, Snail) families. In situ hybridization verified that these genes were enriched in the TEB epithelium (Wnt-5a, Wnt-7b, Dhh, Eph-A2) or TEB stroma (Wnt-2, Frizzled-1, Ephrin-B1). We discuss the potential roles of these genes in mammary branching morphogenesis.
Cancer Cell, 2008
How breast cancers are able to disseminate and metastasize is poorly understood. Using hyperplasi... more How breast cancers are able to disseminate and metastasize is poorly understood. Using hyperplasia transplant system, we show that tumor dissemination and metastasis occur in discrete steps during tumor progression. Bioinformatic analysis revealed that loss of the transcription factor GATA-3 marked progression from adenoma to early carcinoma and onset of tumor dissemination. Restoration of GATA-3 in late carcinomas induced tumor differentiation suppressed tumor dissemination. Targeted deletion of GATA-3 in early tumors led to apoptosis of differentiated cells, indicating that its loss is not sufficient for malignant conversion. Rather, malignant progression occurred with an expanding GATA-3 negative tumor cell population. These data indicate that GATA-3 regulates tumor differentiation and suppresses tumor dissemination in breast cancer.
BMC Bioinformatics, 2006
Background: Monoclonal antibodies are used extensively throughout the biomedical sciences for det... more Background: Monoclonal antibodies are used extensively throughout the biomedical sciences for detection of antigens, either in vitro or in vivo. We, for example, have used them for quantitation of proteins on "reverse-phase" protein lysate arrays. For those studies, we quality-controlled > 600 available monoclonal antibodies and also needed to develop precise information on the genes that encode their antigens. Translation among the various protein and gene identifier types proved nontrivial because of one-to-many and many-to-one relationships. To organize the antibody, protein, and gene information, we initially developed a relational database in Filemaker for our own use. When it became apparent that the information would be useful to many other researchers faced with the need to choose or characterize antibodies, we developed it further as AbMiner, a fully relational web-based database under MySQL, programmed in Java.
Because most potential molecular markers and targets are proteins, proteomic profiling is expecte... more Because most potential molecular markers and targets are proteins, proteomic profiling is expected to yield more direct answers to functional and pharmacological questions than does transcriptional profiling. To aid in such studies, we have developed a protocol for making reverse-phase protein lysate microarrays with larger numbers of spots than previously feasible. Our first application of these arrays was to profiling of the 60 human cancer cell lines (NCI-60) used by the National Cancer Institute to screen compounds for anticancer activity. Each glass slide microarray included 648 lysate spots representing the NCI-60 cell lines plus controls, each at 10 two-fold serial dilutions to provide a wide dynamic range. Mouse monoclonal antibodies and the catalyzed signal amplification system were used for immunoquantitation. The signal levels from the >30,000 data points for our first 52 antibodies were analyzed by using P-SCAN and a quantitative dose interpolation method. Clustered image maps revealed biologically interpretable patterns of protein expression. Among the principal early findings from these arrays were two promising pathological markers for distinguishing colon from ovarian adenocarcinomas. When we compared the patterns of protein expression with those we had obtained for the same genes at the mRNA level by using both cDNA and oligonucleotide arrays, a striking regularity appeared: cell-structure-related proteins almost invariably showed a high correlation between mRNA and protein levels across the NCI-60 cell lines, whereas non-cell-structure-related proteins showed poor correlation.
To study the molecular mechanisms by which drug resistance develops, we compared DU145 human pros... more To study the molecular mechanisms by which drug resistance develops, we compared DU145 human prostate cancer cells with a subline selected for resistance to camptothecin. Differences in gene expression level were assessed by hybridizing the two cell types against each other using quadruplicate "Oncochip" cDNA microarrays that included 1648 cancerrelated genes. Expression levels differing by a factor of >1.5 were detected for 181 of the genes. These differences were judged statistically reliable on the basis of a stratum-adjusted Kruskal-Wallis test, after taking into account a dye-dependent variable. The 181 expression-altered genes included a larger than expected number of the "apoptosis-related" genes (P ؍ 0.04). To assess whether this observation reflected a generalized resistance of RCO.1 to apoptosis, we exposed the cells to a range of stresses (cisplatin, staurosporine, UV, ionizing radiation, and serum starvation) and found greatly reduced apoptotic responses for RC0.1 (relative to DU145) using flow cytometric Annexin V and terminal deoxynucleotidyl transferase-mediated nick end labeling assays. We next examined the apoptosis-related genes in the context of a molecular interaction map and found expression differences in the direction "expected" on the basis of the apoptosis-resistance of RC0.1 for BAD, caspase-6, and genes that signal via the Akt pathway. Exposure of the cells to wortmannin, an inhibitor of the Akt effector phosphatidylinositol 3-kinase, provided functional support for involvement of the Akt pathway. However, closer examination of the molecular interaction map revealed a paradox: many of the expression differences observed for apoptosis-related genes were in the direction "contrary" to that expected given the resistance of RC0.1. The map indicated that most of these unexpected expression differences were associated with genes involved in the nuclear factor B and transforming growth factor  pathways. Overall, the patterns that emerged suggested a two-step model for the selection process that led to resistance in RC0.1 cells. The first hypothesized step would involve a decrease in apoptotic susceptibility through changes in the apoptosis-control machinery associated with the Bcl-2 and caspase gene families, and also in antiapoptotic pathways operating through Akt/PKB. The second step would involve changes in multifunctional upstream genes (including some genes in the nuclear factor B and transforming growth factor  pathways) that can facilitate apoptosis but that would also tend to contribute to cell proliferation in the presence of drug. Thus, we propose that a downstream blockade of apoptosis was "permissive" for the selection of upstream pathway changes that would otherwise have induced apoptosis. This model is analogous to one suggested previously for the relationship between oncogene function and apoptosis in carcinogenesis.
Molecular Cancer Therapeutics, 2007
E-cadherin (E-cad) is a transmembrane adhesion glycoprotein, the expression of which is often red... more E-cadherin (E-cad) is a transmembrane adhesion glycoprotein, the expression of which is often reduced in invasive or metastatic tumors. To assess E-cad's distribution among different types of cancer cells, we used bisulfite-sequencing for detailed, base-by-base measurement of CpG methylation in E-cad's promoter region in the NCI-60 cell lines. The mean methylation levels of the cell lines were distributed bimodally, with values pushed toward either the high or low end of the methylation scale. The 38 epithelial cell lines showed substantially lower (28%) mean methylation levels compared with the nonepithelial cell lines (58%). The CpG site at -143 with respect to the transcriptional start was commonly methylated at intermediate levels, even in cell lines with low overall DNA methylation. We also profiled the NCI-60 cell lines using Affymetrix U133 microarrays and found E-cad expression to be correlated with E-cad methylation at highly statistically significant levels. Above a threshold of approximately 20% to 30% mean methylation, the expression of E-cad was effectively silenced. Overall, this study provides a type of detailed analysis of methylation that can also be applied to other cancer-related genes. As has been shown in recent years, DNA methylation status can serve as a biomarker for use in choosing therapy.
Proceedings of The National Academy of Sciences, 2003
Because most potential molecular markers and targets are proteins, proteomic profiling is expecte... more Because most potential molecular markers and targets are proteins, proteomic profiling is expected to yield more direct answers to functional and pharmacological questions than does transcriptional profiling. To aid in such studies, we have developed a protocol for making reverse-phase protein lysate microarrays with larger numbers of spots than previously feasible. Our first application of these arrays was to profiling of the 60 human cancer cell lines (NCI-60) used by the National Cancer Institute to screen compounds for anticancer activity. Each glass slide microarray included 648 lysate spots representing the NCI-60 cell lines plus controls, each at 10 two-fold serial dilutions to provide a wide dynamic range. Mouse monoclonal antibodies and the catalyzed signal amplification system were used for immunoquantitation. The signal levels from the >30,000 data points for our first 52 antibodies were analyzed by using P-SCAN and a quantitative dose interpolation method. Clustered image maps revealed biologically interpretable patterns of protein expression. Among the principal early findings from these arrays were two promising pathological markers for distinguishing colon from ovarian adenocarcinomas. When we compared the patterns of protein expression with those we had obtained for the same genes at the mRNA level by using both cDNA and oligonucleotide arrays, a striking regularity appeared: cell-structure-related proteins almost invariably showed a high correlation between mRNA and protein levels across the NCI-60 cell lines, whereas non-cell-structure-related proteins showed poor correlation.
Chromosome rearrangement, a hallmark of cancer, has profound effects on carcinogenesis and tumor ... more Chromosome rearrangement, a hallmark of cancer, has profound effects on carcinogenesis and tumor phenotype. We used a panel of 60 human cancer cell lines (the NCI-60) as a model system to identify relationships among DNA copy number, mRNA expression level, and drug sensitivity. For each of 64 cancer-relevant genes, we calculated all 4,096 possible Pearson's correlation coefficients relating DNA copy number (assessed by comparative genomic hybridization using bacterial artificial chromosome microarrays) and mRNA expression level (determined using both cDNA and Affymetrix oligonucleotide microarrays). The analysis identified an association of ERBB2 overexpression with 3p copy number, a finding supported by data from human tumors and a mouse model of ERBB2-induced carcinogenesis. When we examined the correlation between DNA copy number for all 353 unique loci on the bacterial artificial chromosome microarray and drug sensitivity for 118 drugs with putatively known mechanisms of action, we found a striking negative correlation ( −0.983; 95% bootstrap confidence interval, −0.999 to −0.899) between activity of the enzyme drug L-asparaginase and DNA copy number of genes near asparagine synthetase in the ovarian cancer cells. Previous analysis of drug sensitivity and mRNA expression had suggested an inverse relationship between mRNA levels of asparagine synthetase and L-asparaginase sensitivity in the NCI-60. The concordance of pharmacogenomic findings at the DNA and mRNA levels strongly suggests further study of L-asparaginase for possible treatment of a low-synthetase subset of clinical
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Papers by Hosein Kouros-Mehr