In contrast to normal cells, cancer cells avidly take up glucose and metabolize it to lactate eve... more In contrast to normal cells, cancer cells avidly take up glucose and metabolize it to lactate even when oxygen is abundant, a phenomenon referred to as the Warburg effect. This fundamental alteration in glucose metabolism in cancer cells enables their specific detection by positron emission tomography (PET) following i.v. injection of the glucose analogue (18)F-fluorodeoxy-glucose ((18)FDG). However, this useful imaging technique is limited by the fact that not all cancers avidly take up FDG. To identify molecular determinants of (18)FDG retention, we interrogated the transcriptomes of human-cancer cell lines and primary tumors for metabolic pathways associated with (18)FDG radiotracer uptake. From ninety-five metabolic pathways that were interrogated, the glycolysis, and several glycolysis-related pathways (pentose phosphate, carbon fixation, aminoacyl-tRNA biosynthesis, one-carbon-pool by folate) showed the greatest transcriptional enrichment. This "FDG signature" predic...
The octamer sequence 5'-ATGCAAAT, in either orientation, serves as an upstream element in... more The octamer sequence 5'-ATGCAAAT, in either orientation, serves as an upstream element in a variety of promoters and also occurs as a modular enhancer element. It is of particular interest in immunoglobulin genes since it is found in the upstream regions of all heavy and light chain promoters and in the heavy chain enhancer, both of which are known to be necessary for cell-specific expression. We report here the chromatographic separation of ubiquitous and B cell-specific octamer-binding proteins. The B cell factor was purified to homogeneity using affinity chromatography and consists of three peptides of 62, 61, and 58.5 +/- 1.5 kd. Each of the polypeptides was renatured after SDS-PAGE and shown to bind to the octamer sequence. The specific DNA binding activity of the pure B cell-specific factor was indistinguishable from that of the affinity-purified ubiquitous factor. This B cell-specific octamer-binding factor, in pure form, activated transcription from a kappa light chain promoter in vitro, thus demonstrating that it is indeed a B cell-specific transcription factor for this gene. In addition to the ubiquitous and B cell-specific octamer-binding factors, we identified several additional proteins, one of which is B cell-specific, that interact with the kappa promoter.
We describe a region of human DNA containing four metallothionein (hMT) genes. One of these genes... more We describe a region of human DNA containing four metallothionein (hMT) genes. One of these genes, hMT-IA, was found to encode a functional protein that confers heavy metal resistance to NIH 3T3 cells after transfer on a bovine papilloma virus-derived vector. This gene is expressed in cultured human cell lines, but at a lower basal level than the hMT-IIA gene; it shows a different induction response to heavy metals and glucocorticoids than the hMT-IIA gene. Induction of the human MT family therefore does not represent an equivalent elevation in the level of expression of individual genes, but is the sum of the differential responses of active members. The differential response is due to functional differences of the respective promoter/regulatory regions of the genes as shown by gene-fusion experiments. While the hMT-IIA promoter is responsive to Cd++, Zn++, and glucocorticoids, the hMT-IA promoter mediates response only to Cd++.
H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine r... more H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine residue at position 27 (K27M) or glycine at position 34 (G34R/V). Sixteen genes encode histone H3, each variant differing in only a few amino acids. Therefore, how mutations in a single H3 gene contribute to carcinogenesis is unknown. H3F3A K27M mutations are predicted to alter methylation of H3K27. H3K27me3 is a repressive mark critical to stem cell maintenance and is mediated by EZH2, a member of the polycomb-group (PcG) family. We evaluated H3K27me3 and EZH2 expression using immunohistochemistry in 76 pediatric brain tumors. H3K27me3 was lowered/absent in tumor cells but preserved in endothelial cells and infiltrating lymphocytes in six out of 20 GBMs. H3K27me3 showed strong immunoreactivity in all other tumor subtypes. Sequencing of GBMs showed H3F3A K27M mutations in all six cases with lowered/absent H3K27me3. EZH2 expression was high in GBMs, but absent/focal in other tumors. However, no significant differences in EZH2 expression were observed between H3F3A K27M mutant and wild type GBMs, suggesting that EZH2 mediated trimethylation of H3K27 is inhibited in GBM harboring K27M mutations. Our results indicate that H3F3A K27M mutant GBMs show decreased H3K27me3 that may be of both diagnostic and biological relevance.
The pleural space is an attractive site for using viral vectors to deliver gene products to the l... more The pleural space is an attractive site for using viral vectors to deliver gene products to the lung parenchyma, other thoracic structures and the systemic circulation. The advantages of intrapleural gene transfer using viral vectors include: (i) easy accessibility; (ii) large surface area; (iii) ability to provide high concentrations of secreted gene products to chest structures; (iv) low risk of detrimental effects of possible vector-induced inflammation compared with intravascular delivery; and (v) because it is local, lower vector doses can be used to deliver therapeutic genes to thoracic structures than less efficient systemic routes. Examples of pleural gene transfer include the use of adenovirus vectors to treat mesothelioma by transiently expressing genes that encode toxic proteins, immunomodulatory molecules or anti-angiogenesis factors. Intrapleural delivery of adeno-associated viral vectors represents an efficient strategy to treat alpha1-antitrypsin (alpha1AT) deficiency, achieving high lung and systemic therapeutic levels of alpha1AT. Intrapleural delivery of gene transfer vectors holds promise for the treatment of diseases requiring transient, localized gene expression, as well as sustained expression of genes to correct hereditary disorders requiring localized or systemic expression of the therapeutic protein.
The cytoplasmic domain of the human T cell-type interleukin-1 receptor (hIL-1R) is not involved i... more The cytoplasmic domain of the human T cell-type interleukin-1 receptor (hIL-1R) is not involved in the binding, internalization, or nuclear localization of interleukin-1 (IL-1), but is essential for signal transduction. We have previously localized a 50-amino acid region (residues 477-527) critical for IL-1-mediated activation of the interleukin-2 promoter in T cells. This region displays a striking degree of amino acid conservation in human, murine, and chicken IL-1Rs. Here we report the results of a site-directed mutational analysis of the cytoplasmic domain of the hIL-1R. We have introduced single-amino acid substitutions at positions conserved in all three receptors and at nonconserved positions and identified key amino acids for IL-1R function in signal transduction. Three basic (Arg431, Lys515, and Arg518) and 3 aromatic (Phe513, Trp514, and Tyr519) amino acids that are conserved in human, murine, and chicken IL-1Rs could not be replaced without abolishing IL-1R-mediated signal transduction. A substitution at another conserved position (Pro521) reduces significantly the ability of the IL-1R to transmit the IL-1 signal. Nonconserved residues could be replaced without affecting signal transduction. The cytoplasmic domain of the IL-1R is related to that of the Drosophila Toll protein, with a 26% identity and a 43% similarity in amino acid sequence. The amino acids shown to be essential for IL-1R function are conserved in the Toll protein. Our experimental data indicate that the amino acid sequence similarity between the IL-1R and the Drosophila toll protein reflects a functional homology between the two proteins.
Malignant pleural mesothelioma (MPM) is an aggressive neoplasm associated with asbestos exposure.... more Malignant pleural mesothelioma (MPM) is an aggressive neoplasm associated with asbestos exposure. Although previous studies based on candidate gene approaches have identified important common somatic mutations in MPM, these studies have focused on small sets of genes and have provided a limited view of the genetic alterations underlying this disease. Here, we performed whole-exome sequencing on DNA from 22 MPMs and matched blood samples, and identified 517 somatic mutations across 490 mutated genes. Integrative analysis of mutations and somatic copy-number alterations revealed frequent genetic alterations in BAP1, NF2, CDKN2A, and CUL1. Our study presents the first unbiased view of the genomic basis of MPM.
In contrast to normal cells, cancer cells avidly take up glucose and metabolize it to lactate eve... more In contrast to normal cells, cancer cells avidly take up glucose and metabolize it to lactate even when oxygen is abundant, a phenomenon referred to as the Warburg effect. This fundamental alteration in glucose metabolism in cancer cells enables their specific detection by positron emission tomography (PET) following i.v. injection of the glucose analogue (18)F-fluorodeoxy-glucose ((18)FDG). However, this useful imaging technique is limited by the fact that not all cancers avidly take up FDG. To identify molecular determinants of (18)FDG retention, we interrogated the transcriptomes of human-cancer cell lines and primary tumors for metabolic pathways associated with (18)FDG radiotracer uptake. From ninety-five metabolic pathways that were interrogated, the glycolysis, and several glycolysis-related pathways (pentose phosphate, carbon fixation, aminoacyl-tRNA biosynthesis, one-carbon-pool by folate) showed the greatest transcriptional enrichment. This "FDG signature" predic...
The octamer sequence 5'-ATGCAAAT, in either orientation, serves as an upstream element in... more The octamer sequence 5'-ATGCAAAT, in either orientation, serves as an upstream element in a variety of promoters and also occurs as a modular enhancer element. It is of particular interest in immunoglobulin genes since it is found in the upstream regions of all heavy and light chain promoters and in the heavy chain enhancer, both of which are known to be necessary for cell-specific expression. We report here the chromatographic separation of ubiquitous and B cell-specific octamer-binding proteins. The B cell factor was purified to homogeneity using affinity chromatography and consists of three peptides of 62, 61, and 58.5 +/- 1.5 kd. Each of the polypeptides was renatured after SDS-PAGE and shown to bind to the octamer sequence. The specific DNA binding activity of the pure B cell-specific factor was indistinguishable from that of the affinity-purified ubiquitous factor. This B cell-specific octamer-binding factor, in pure form, activated transcription from a kappa light chain promoter in vitro, thus demonstrating that it is indeed a B cell-specific transcription factor for this gene. In addition to the ubiquitous and B cell-specific octamer-binding factors, we identified several additional proteins, one of which is B cell-specific, that interact with the kappa promoter.
We describe a region of human DNA containing four metallothionein (hMT) genes. One of these genes... more We describe a region of human DNA containing four metallothionein (hMT) genes. One of these genes, hMT-IA, was found to encode a functional protein that confers heavy metal resistance to NIH 3T3 cells after transfer on a bovine papilloma virus-derived vector. This gene is expressed in cultured human cell lines, but at a lower basal level than the hMT-IIA gene; it shows a different induction response to heavy metals and glucocorticoids than the hMT-IIA gene. Induction of the human MT family therefore does not represent an equivalent elevation in the level of expression of individual genes, but is the sum of the differential responses of active members. The differential response is due to functional differences of the respective promoter/regulatory regions of the genes as shown by gene-fusion experiments. While the hMT-IIA promoter is responsive to Cd++, Zn++, and glucocorticoids, the hMT-IA promoter mediates response only to Cd++.
H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine r... more H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine residue at position 27 (K27M) or glycine at position 34 (G34R/V). Sixteen genes encode histone H3, each variant differing in only a few amino acids. Therefore, how mutations in a single H3 gene contribute to carcinogenesis is unknown. H3F3A K27M mutations are predicted to alter methylation of H3K27. H3K27me3 is a repressive mark critical to stem cell maintenance and is mediated by EZH2, a member of the polycomb-group (PcG) family. We evaluated H3K27me3 and EZH2 expression using immunohistochemistry in 76 pediatric brain tumors. H3K27me3 was lowered/absent in tumor cells but preserved in endothelial cells and infiltrating lymphocytes in six out of 20 GBMs. H3K27me3 showed strong immunoreactivity in all other tumor subtypes. Sequencing of GBMs showed H3F3A K27M mutations in all six cases with lowered/absent H3K27me3. EZH2 expression was high in GBMs, but absent/focal in other tumors. However, no significant differences in EZH2 expression were observed between H3F3A K27M mutant and wild type GBMs, suggesting that EZH2 mediated trimethylation of H3K27 is inhibited in GBM harboring K27M mutations. Our results indicate that H3F3A K27M mutant GBMs show decreased H3K27me3 that may be of both diagnostic and biological relevance.
The pleural space is an attractive site for using viral vectors to deliver gene products to the l... more The pleural space is an attractive site for using viral vectors to deliver gene products to the lung parenchyma, other thoracic structures and the systemic circulation. The advantages of intrapleural gene transfer using viral vectors include: (i) easy accessibility; (ii) large surface area; (iii) ability to provide high concentrations of secreted gene products to chest structures; (iv) low risk of detrimental effects of possible vector-induced inflammation compared with intravascular delivery; and (v) because it is local, lower vector doses can be used to deliver therapeutic genes to thoracic structures than less efficient systemic routes. Examples of pleural gene transfer include the use of adenovirus vectors to treat mesothelioma by transiently expressing genes that encode toxic proteins, immunomodulatory molecules or anti-angiogenesis factors. Intrapleural delivery of adeno-associated viral vectors represents an efficient strategy to treat alpha1-antitrypsin (alpha1AT) deficiency, achieving high lung and systemic therapeutic levels of alpha1AT. Intrapleural delivery of gene transfer vectors holds promise for the treatment of diseases requiring transient, localized gene expression, as well as sustained expression of genes to correct hereditary disorders requiring localized or systemic expression of the therapeutic protein.
The cytoplasmic domain of the human T cell-type interleukin-1 receptor (hIL-1R) is not involved i... more The cytoplasmic domain of the human T cell-type interleukin-1 receptor (hIL-1R) is not involved in the binding, internalization, or nuclear localization of interleukin-1 (IL-1), but is essential for signal transduction. We have previously localized a 50-amino acid region (residues 477-527) critical for IL-1-mediated activation of the interleukin-2 promoter in T cells. This region displays a striking degree of amino acid conservation in human, murine, and chicken IL-1Rs. Here we report the results of a site-directed mutational analysis of the cytoplasmic domain of the hIL-1R. We have introduced single-amino acid substitutions at positions conserved in all three receptors and at nonconserved positions and identified key amino acids for IL-1R function in signal transduction. Three basic (Arg431, Lys515, and Arg518) and 3 aromatic (Phe513, Trp514, and Tyr519) amino acids that are conserved in human, murine, and chicken IL-1Rs could not be replaced without abolishing IL-1R-mediated signal transduction. A substitution at another conserved position (Pro521) reduces significantly the ability of the IL-1R to transmit the IL-1 signal. Nonconserved residues could be replaced without affecting signal transduction. The cytoplasmic domain of the IL-1R is related to that of the Drosophila Toll protein, with a 26% identity and a 43% similarity in amino acid sequence. The amino acids shown to be essential for IL-1R function are conserved in the Toll protein. Our experimental data indicate that the amino acid sequence similarity between the IL-1R and the Drosophila toll protein reflects a functional homology between the two proteins.
Malignant pleural mesothelioma (MPM) is an aggressive neoplasm associated with asbestos exposure.... more Malignant pleural mesothelioma (MPM) is an aggressive neoplasm associated with asbestos exposure. Although previous studies based on candidate gene approaches have identified important common somatic mutations in MPM, these studies have focused on small sets of genes and have provided a limited view of the genetic alterations underlying this disease. Here, we performed whole-exome sequencing on DNA from 22 MPMs and matched blood samples, and identified 517 somatic mutations across 490 mutated genes. Integrative analysis of mutations and somatic copy-number alterations revealed frequent genetic alterations in BAP1, NF2, CDKN2A, and CUL1. Our study presents the first unbiased view of the genomic basis of MPM.
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Papers by A. Heguy