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Catechol-O-methyltransferase (COMT)-mediated metabolism of catechol estrogens : Comparison of wild-type and variant COMT isoforms
The oxidative metabolism of 17beta-estradiol (E2) and estrone (E1) to catechol estrogens (2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1) and estrogen quinones has been postulated to be a factor in mammary carcinogenesis. Catechol-O-methyltransferase (COMT) catalyzes the methylation of catechol estrogens to met...
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Published in: | Cancer research (Chicago, Ill.) Ill.), 2001-09, Vol.61 (18), p.6716-6722 |
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description | The oxidative metabolism of 17beta-estradiol (E2) and estrone (E1) to catechol estrogens (2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1) and estrogen quinones has been postulated to be a factor in mammary carcinogenesis. Catechol-O-methyltransferase (COMT) catalyzes the methylation of catechol estrogens to methoxy estrogens, which simultaneously lowers the potential for DNA damage and increases the concentration of 2-methoxyestradiol (2-MeOE2), an antiproliferative metabolite. We expressed two recombinant forms of COMT, the wild-type (108Val) and a common variant (108Met), to determine whether their catalytic efficiencies differ with respect to catechol estrogen inactivation. The His-tagged proteins were purified by nickel-nitrilo-triacetic acid chromatography and analyzed by electrophoresis and Western immunoblot. COMT activity was assessed by determining the methylation of 2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1, using gas chromatography/mass spectrometry for quantitation of the respective methoxy products. In the case of 2-OHE2 and 2-OHE1, methylation occurred at 2-OH and 3-OH groups, resulting in the formation of 2-MeOE2 and 2-OH-3-MeOE2, and 2-MeOE1 and 2-OH-3-MeOE1, respectively. In contrast, in the case of 4-OHE2 and 4-OHE1, methylation occurred only at the 4-OH group, yielding 4-MeOE2 and 4-MeOE1, respectively. Individual and competition experiments revealed the following order of product formation: 4-MeOE2 > 4-MeOE1 >> 2-MeOE2 > 2-MeOE1 > 2-OH-3-MeOE1 > 2-OH-3-MeOE2. The variant isoform differed from wild-type COMT by being thermolabile, leading to 2-3-fold lower levels of product formation. MCF-7 breast cancer cells with the variant COMT 108Met/Met genotype also displayed 2-3-fold lower catalytic activity than ZR-75 breast cancer cells with the wild-type COMT 108Val/Val genotype. Thus, inherited alterations in COMT catalytic activity are associated with significant differences in catechol estrogen and methoxy estrogen levels and, thereby, may contribute to interindividual differences in breast cancer risk associated with estrogen-mediated carcinogenicity. |
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Catechol-O-methyltransferase (COMT) catalyzes the methylation of catechol estrogens to methoxy estrogens, which simultaneously lowers the potential for DNA damage and increases the concentration of 2-methoxyestradiol (2-MeOE2), an antiproliferative metabolite. We expressed two recombinant forms of COMT, the wild-type (108Val) and a common variant (108Met), to determine whether their catalytic efficiencies differ with respect to catechol estrogen inactivation. The His-tagged proteins were purified by nickel-nitrilo-triacetic acid chromatography and analyzed by electrophoresis and Western immunoblot. COMT activity was assessed by determining the methylation of 2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1, using gas chromatography/mass spectrometry for quantitation of the respective methoxy products. In the case of 2-OHE2 and 2-OHE1, methylation occurred at 2-OH and 3-OH groups, resulting in the formation of 2-MeOE2 and 2-OH-3-MeOE2, and 2-MeOE1 and 2-OH-3-MeOE1, respectively. In contrast, in the case of 4-OHE2 and 4-OHE1, methylation occurred only at the 4-OH group, yielding 4-MeOE2 and 4-MeOE1, respectively. Individual and competition experiments revealed the following order of product formation: 4-MeOE2 > 4-MeOE1 >> 2-MeOE2 > 2-MeOE1 > 2-OH-3-MeOE1 > 2-OH-3-MeOE2. The variant isoform differed from wild-type COMT by being thermolabile, leading to 2-3-fold lower levels of product formation. MCF-7 breast cancer cells with the variant COMT 108Met/Met genotype also displayed 2-3-fold lower catalytic activity than ZR-75 breast cancer cells with the wild-type COMT 108Val/Val genotype. Thus, inherited alterations in COMT catalytic activity are associated with significant differences in catechol estrogen and methoxy estrogen levels and, thereby, may contribute to interindividual differences in breast cancer risk associated with estrogen-mediated carcinogenicity.</description><identifier>ISSN: 0008-5472</identifier><identifier>EISSN: 1538-7445</identifier><identifier>PMID: 11559542</identifier><identifier>CODEN: CNREA8</identifier><language>eng</language><publisher>Philadelphia, PA: American Association for Cancer Research</publisher><subject>Alleles ; Analytical, structural and metabolic biochemistry ; Biological and medical sciences ; Breast Neoplasms - enzymology ; Breast Neoplasms - genetics ; Catechol O-Methyltransferase - genetics ; Catechol O-Methyltransferase - metabolism ; Cloning, Molecular ; Enzyme Activation ; Enzyme Stability ; Enzyme-Linked Immunosorbent Assay ; Estrogens, Catechol - metabolism ; Fundamental and applied biological sciences. Psychology ; Gas Chromatography-Mass Spectrometry ; Gynecology. Andrology. Obstetrics ; Humans ; Isoenzymes - genetics ; Isoenzymes - metabolism ; Mammary gland diseases ; Medical sciences ; Methylation ; Other biological molecules ; Polymerase Chain Reaction ; Recombinant Proteins - genetics ; Recombinant Proteins - metabolism ; Terpenes, steroids. Hormones ; Tumor Cells, Cultured ; Tumors</subject><ispartof>Cancer research (Chicago, Ill.), 2001-09, Vol.61 (18), p.6716-6722</ispartof><rights>2001 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=1139369$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/11559542$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>DAWLING, Sheila</creatorcontrib><creatorcontrib>ROODI, Nady</creatorcontrib><creatorcontrib>MERNAUGH, Raymond L</creatorcontrib><creatorcontrib>XIAOHONG WANG</creatorcontrib><creatorcontrib>PARL, Fritz F</creatorcontrib><title>Catechol-O-methyltransferase (COMT)-mediated metabolism of catechol estrogens : Comparison of wild-type and variant COMT isoforms</title><title>Cancer research (Chicago, Ill.)</title><addtitle>Cancer Res</addtitle><description>The oxidative metabolism of 17beta-estradiol (E2) and estrone (E1) to catechol estrogens (2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1) and estrogen quinones has been postulated to be a factor in mammary carcinogenesis. Catechol-O-methyltransferase (COMT) catalyzes the methylation of catechol estrogens to methoxy estrogens, which simultaneously lowers the potential for DNA damage and increases the concentration of 2-methoxyestradiol (2-MeOE2), an antiproliferative metabolite. We expressed two recombinant forms of COMT, the wild-type (108Val) and a common variant (108Met), to determine whether their catalytic efficiencies differ with respect to catechol estrogen inactivation. The His-tagged proteins were purified by nickel-nitrilo-triacetic acid chromatography and analyzed by electrophoresis and Western immunoblot. COMT activity was assessed by determining the methylation of 2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1, using gas chromatography/mass spectrometry for quantitation of the respective methoxy products. In the case of 2-OHE2 and 2-OHE1, methylation occurred at 2-OH and 3-OH groups, resulting in the formation of 2-MeOE2 and 2-OH-3-MeOE2, and 2-MeOE1 and 2-OH-3-MeOE1, respectively. In contrast, in the case of 4-OHE2 and 4-OHE1, methylation occurred only at the 4-OH group, yielding 4-MeOE2 and 4-MeOE1, respectively. Individual and competition experiments revealed the following order of product formation: 4-MeOE2 > 4-MeOE1 >> 2-MeOE2 > 2-MeOE1 > 2-OH-3-MeOE1 > 2-OH-3-MeOE2. The variant isoform differed from wild-type COMT by being thermolabile, leading to 2-3-fold lower levels of product formation. MCF-7 breast cancer cells with the variant COMT 108Met/Met genotype also displayed 2-3-fold lower catalytic activity than ZR-75 breast cancer cells with the wild-type COMT 108Val/Val genotype. Thus, inherited alterations in COMT catalytic activity are associated with significant differences in catechol estrogen and methoxy estrogen levels and, thereby, may contribute to interindividual differences in breast cancer risk associated with estrogen-mediated carcinogenicity.</description><subject>Alleles</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Biological and medical sciences</subject><subject>Breast Neoplasms - enzymology</subject><subject>Breast Neoplasms - genetics</subject><subject>Catechol O-Methyltransferase - genetics</subject><subject>Catechol O-Methyltransferase - metabolism</subject><subject>Cloning, Molecular</subject><subject>Enzyme Activation</subject><subject>Enzyme Stability</subject><subject>Enzyme-Linked Immunosorbent Assay</subject><subject>Estrogens, Catechol - metabolism</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gas Chromatography-Mass Spectrometry</subject><subject>Gynecology. Andrology. Obstetrics</subject><subject>Humans</subject><subject>Isoenzymes - genetics</subject><subject>Isoenzymes - metabolism</subject><subject>Mammary gland diseases</subject><subject>Medical sciences</subject><subject>Methylation</subject><subject>Other biological molecules</subject><subject>Polymerase Chain Reaction</subject><subject>Recombinant Proteins - genetics</subject><subject>Recombinant Proteins - metabolism</subject><subject>Terpenes, steroids. Hormones</subject><subject>Tumor Cells, Cultured</subject><subject>Tumors</subject><issn>0008-5472</issn><issn>1538-7445</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2001</creationdate><recordtype>article</recordtype><recordid>eNpN0E1LxDAQBuAgiruu_gXJwYMeAknTNKk3KX7Byl7W8zLNh1tpm5JEZY_-c7O4gqdhmGcG5j1Ccya4IrIsxTGaU0oVEaUsZugsxvfcCkbFKZoxJkQtymKOvhtIVm99T1ZksGm761OAMTobIFp83axe1jd5YLrMDM4CWt93ccDeYX1YxTam4N_sGPEtbvwwQeiiH_fkq-sNSbvJYhgN_swDGBPeX8WZOB-GeI5OHPTRXhzqAr0-3K-bJ7JcPT43d0uyLSRNpJJS1lozWhpbWJBO01ZoKgUIRZVsOTOyVUYqbQpeiVo7ypRkonBGVa2xfIEuf-9OH21-aDOFboCw2_xlkcHVAUDU0Lucg-7iP8drXtX8Bx3wagE</recordid><startdate>20010915</startdate><enddate>20010915</enddate><creator>DAWLING, Sheila</creator><creator>ROODI, Nady</creator><creator>MERNAUGH, Raymond L</creator><creator>XIAOHONG WANG</creator><creator>PARL, Fritz F</creator><general>American Association for Cancer Research</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope></search><sort><creationdate>20010915</creationdate><title>Catechol-O-methyltransferase (COMT)-mediated metabolism of catechol estrogens : Comparison of wild-type and variant COMT isoforms</title><author>DAWLING, Sheila ; ROODI, Nady ; MERNAUGH, Raymond L ; XIAOHONG WANG ; PARL, Fritz F</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-h270t-67779cc104de2ea7fc0b5c075a58087b31d7b8d78cd23659cf0187152fd86bde3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2001</creationdate><topic>Alleles</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Biological and medical sciences</topic><topic>Breast Neoplasms - enzymology</topic><topic>Breast Neoplasms - genetics</topic><topic>Catechol O-Methyltransferase - genetics</topic><topic>Catechol O-Methyltransferase - metabolism</topic><topic>Cloning, Molecular</topic><topic>Enzyme Activation</topic><topic>Enzyme Stability</topic><topic>Enzyme-Linked Immunosorbent Assay</topic><topic>Estrogens, Catechol - metabolism</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Gas Chromatography-Mass Spectrometry</topic><topic>Gynecology. Andrology. Obstetrics</topic><topic>Humans</topic><topic>Isoenzymes - genetics</topic><topic>Isoenzymes - metabolism</topic><topic>Mammary gland diseases</topic><topic>Medical sciences</topic><topic>Methylation</topic><topic>Other biological molecules</topic><topic>Polymerase Chain Reaction</topic><topic>Recombinant Proteins - genetics</topic><topic>Recombinant Proteins - metabolism</topic><topic>Terpenes, steroids. Hormones</topic><topic>Tumor Cells, Cultured</topic><topic>Tumors</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>DAWLING, Sheila</creatorcontrib><creatorcontrib>ROODI, Nady</creatorcontrib><creatorcontrib>MERNAUGH, Raymond L</creatorcontrib><creatorcontrib>XIAOHONG WANG</creatorcontrib><creatorcontrib>PARL, Fritz F</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><jtitle>Cancer research (Chicago, Ill.)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>DAWLING, Sheila</au><au>ROODI, Nady</au><au>MERNAUGH, Raymond L</au><au>XIAOHONG WANG</au><au>PARL, Fritz F</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Catechol-O-methyltransferase (COMT)-mediated metabolism of catechol estrogens : Comparison of wild-type and variant COMT isoforms</atitle><jtitle>Cancer research (Chicago, Ill.)</jtitle><addtitle>Cancer Res</addtitle><date>2001-09-15</date><risdate>2001</risdate><volume>61</volume><issue>18</issue><spage>6716</spage><epage>6722</epage><pages>6716-6722</pages><issn>0008-5472</issn><eissn>1538-7445</eissn><coden>CNREA8</coden><abstract>The oxidative metabolism of 17beta-estradiol (E2) and estrone (E1) to catechol estrogens (2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1) and estrogen quinones has been postulated to be a factor in mammary carcinogenesis. Catechol-O-methyltransferase (COMT) catalyzes the methylation of catechol estrogens to methoxy estrogens, which simultaneously lowers the potential for DNA damage and increases the concentration of 2-methoxyestradiol (2-MeOE2), an antiproliferative metabolite. We expressed two recombinant forms of COMT, the wild-type (108Val) and a common variant (108Met), to determine whether their catalytic efficiencies differ with respect to catechol estrogen inactivation. The His-tagged proteins were purified by nickel-nitrilo-triacetic acid chromatography and analyzed by electrophoresis and Western immunoblot. COMT activity was assessed by determining the methylation of 2-OHE2, 4-OHE2, 2-OHE1, and 4-OHE1, using gas chromatography/mass spectrometry for quantitation of the respective methoxy products. In the case of 2-OHE2 and 2-OHE1, methylation occurred at 2-OH and 3-OH groups, resulting in the formation of 2-MeOE2 and 2-OH-3-MeOE2, and 2-MeOE1 and 2-OH-3-MeOE1, respectively. In contrast, in the case of 4-OHE2 and 4-OHE1, methylation occurred only at the 4-OH group, yielding 4-MeOE2 and 4-MeOE1, respectively. Individual and competition experiments revealed the following order of product formation: 4-MeOE2 > 4-MeOE1 >> 2-MeOE2 > 2-MeOE1 > 2-OH-3-MeOE1 > 2-OH-3-MeOE2. The variant isoform differed from wild-type COMT by being thermolabile, leading to 2-3-fold lower levels of product formation. MCF-7 breast cancer cells with the variant COMT 108Met/Met genotype also displayed 2-3-fold lower catalytic activity than ZR-75 breast cancer cells with the wild-type COMT 108Val/Val genotype. Thus, inherited alterations in COMT catalytic activity are associated with significant differences in catechol estrogen and methoxy estrogen levels and, thereby, may contribute to interindividual differences in breast cancer risk associated with estrogen-mediated carcinogenicity.</abstract><cop>Philadelphia, PA</cop><pub>American Association for Cancer Research</pub><pmid>11559542</pmid><tpages>7</tpages></addata></record> |
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subjects | Alleles Analytical, structural and metabolic biochemistry Biological and medical sciences Breast Neoplasms - enzymology Breast Neoplasms - genetics Catechol O-Methyltransferase - genetics Catechol O-Methyltransferase - metabolism Cloning, Molecular Enzyme Activation Enzyme Stability Enzyme-Linked Immunosorbent Assay Estrogens, Catechol - metabolism Fundamental and applied biological sciences. Psychology Gas Chromatography-Mass Spectrometry Gynecology. Andrology. Obstetrics Humans Isoenzymes - genetics Isoenzymes - metabolism Mammary gland diseases Medical sciences Methylation Other biological molecules Polymerase Chain Reaction Recombinant Proteins - genetics Recombinant Proteins - metabolism Terpenes, steroids. Hormones Tumor Cells, Cultured Tumors |
title | Catechol-O-methyltransferase (COMT)-mediated metabolism of catechol estrogens : Comparison of wild-type and variant COMT isoforms |
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