Insertion of Retroviral Vectors in NOD/SCID Repopulating Human Peripheral Blood Progenitor Cells Occurs Preferentially in the Vicinity of Transcription Start Regions and in Introns
Reports on insertional "genotoxicity" in patients have created intense interest in characterizing retroviral vector integrations on the genomic level. The retroviral vector SF91m3 was used for transduction of human peripheral blood progenitor cells (PBPC). These PBPC were transplanted into...
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| Veröffentlicht in: | Molecular therapy 2004-11, Vol.10 (5), p.874-881 |
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| creator | Laufs, Stephanie Nagy, K Zsuzsanna Giordano, Frank A Hotz-Wagenblatt, Agnes Zeller, W Jens Fruehauf, Stefan |
| description | Reports on insertional "genotoxicity" in patients have created intense interest in characterizing retroviral vector integrations on the genomic level. The retroviral vector SF91m3 was used for transduction of human peripheral blood progenitor cells (PBPC). These PBPC were transplanted into nonobese diabetic/severe combined immunodeficient mice. A total of 186 retroviral vector integration sites were isolated by ligation-mediated PCR from chimeric mouse bone marrow of five PBPC donors, sequenced, and blasted against the human genome. Preferred integration near the transcription start regions, within CpG islands, and within Alu regions was observed. Detailed analysis of targeted RefSeq genes showed a favored integration within the first intron. Integrations were most common in genes coding for signaling proteins, transcription factors, and kinases. In all genes targeted independently multiple times the respective orientation of the provirus within the gene was identical, indicating integration hot spot regions and similar steric determinants for integration sites. Possible explanations for these findings could be nonrandom vector integration, clonal selection due to gene expression interference, or engraftment issues related to gene insertion in signaling and cell cycle genes. The low frequency of integrations in exons may be reassuring as to the safety of retroviral gene therapy with normal human PBPC. |
| doi_str_mv | 10.1016/j.ymthe.2004.08.001 |
| format | Article |
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The retroviral vector SF91m3 was used for transduction of human peripheral blood progenitor cells (PBPC). These PBPC were transplanted into nonobese diabetic/severe combined immunodeficient mice. A total of 186 retroviral vector integration sites were isolated by ligation-mediated PCR from chimeric mouse bone marrow of five PBPC donors, sequenced, and blasted against the human genome. Preferred integration near the transcription start regions, within CpG islands, and within Alu regions was observed. Detailed analysis of targeted RefSeq genes showed a favored integration within the first intron. Integrations were most common in genes coding for signaling proteins, transcription factors, and kinases. In all genes targeted independently multiple times the respective orientation of the provirus within the gene was identical, indicating integration hot spot regions and similar steric determinants for integration sites. Possible explanations for these findings could be nonrandom vector integration, clonal selection due to gene expression interference, or engraftment issues related to gene insertion in signaling and cell cycle genes. The low frequency of integrations in exons may be reassuring as to the safety of retroviral gene therapy with normal human PBPC.</description><identifier>ISSN: 1525-0016</identifier><identifier>EISSN: 1525-0024</identifier><identifier>DOI: 10.1016/j.ymthe.2004.08.001</identifier><identifier>PMID: 15509505</identifier><language>eng</language><publisher>United States: Elsevier Limited</publisher><subject>Alu Elements - genetics ; Animals ; Bone marrow ; Cancer research ; Cell cycle ; Chromosomes ; Chromosomes, Human - genetics ; Computer simulation ; CpG Islands - genetics ; Diabetes ; Gene therapy ; Gene Transfer Techniques ; Genetic Therapy ; Genetic Vectors - genetics ; Genomes ; Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells - virology ; Humans ; Introns - genetics ; Kinases ; Leukemia ; Medical research ; Mice ; Mice, Inbred NOD ; Mice, SCID ; Mutagenesis ; Mutagenesis, Insertional - genetics ; Retroviridae - genetics ; Transcription Initiation Site ; Vectors (Biology) ; Virus Integration - genetics</subject><ispartof>Molecular therapy, 2004-11, Vol.10 (5), p.874-881</ispartof><rights>Copyright Nature Publishing Group Nov 2004</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c465t-c112e6f1e8ec3f994d3c2f9b26359ca6a82cde4a6a1568b8473abf4a5d9f5fcb3</citedby><cites>FETCH-LOGICAL-c465t-c112e6f1e8ec3f994d3c2f9b26359ca6a82cde4a6a1568b8473abf4a5d9f5fcb3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.proquest.com/docview/1792811063?pq-origsite=primo$$EHTML$$P50$$Gproquest$$H</linktohtml><link.rule.ids>315,782,786,27931,27932,64392,64396,72476</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15509505$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Laufs, Stephanie</creatorcontrib><creatorcontrib>Nagy, K Zsuzsanna</creatorcontrib><creatorcontrib>Giordano, Frank A</creatorcontrib><creatorcontrib>Hotz-Wagenblatt, Agnes</creatorcontrib><creatorcontrib>Zeller, W Jens</creatorcontrib><creatorcontrib>Fruehauf, Stefan</creatorcontrib><title>Insertion of Retroviral Vectors in NOD/SCID Repopulating Human Peripheral Blood Progenitor Cells Occurs Preferentially in the Vicinity of Transcription Start Regions and in Introns</title><title>Molecular therapy</title><addtitle>Mol Ther</addtitle><description>Reports on insertional "genotoxicity" in patients have created intense interest in characterizing retroviral vector integrations on the genomic level. The retroviral vector SF91m3 was used for transduction of human peripheral blood progenitor cells (PBPC). These PBPC were transplanted into nonobese diabetic/severe combined immunodeficient mice. A total of 186 retroviral vector integration sites were isolated by ligation-mediated PCR from chimeric mouse bone marrow of five PBPC donors, sequenced, and blasted against the human genome. Preferred integration near the transcription start regions, within CpG islands, and within Alu regions was observed. Detailed analysis of targeted RefSeq genes showed a favored integration within the first intron. Integrations were most common in genes coding for signaling proteins, transcription factors, and kinases. In all genes targeted independently multiple times the respective orientation of the provirus within the gene was identical, indicating integration hot spot regions and similar steric determinants for integration sites. Possible explanations for these findings could be nonrandom vector integration, clonal selection due to gene expression interference, or engraftment issues related to gene insertion in signaling and cell cycle genes. The low frequency of integrations in exons may be reassuring as to the safety of retroviral gene therapy with normal human PBPC.</description><subject>Alu Elements - genetics</subject><subject>Animals</subject><subject>Bone marrow</subject><subject>Cancer research</subject><subject>Cell cycle</subject><subject>Chromosomes</subject><subject>Chromosomes, Human - genetics</subject><subject>Computer simulation</subject><subject>CpG Islands - genetics</subject><subject>Diabetes</subject><subject>Gene therapy</subject><subject>Gene Transfer Techniques</subject><subject>Genetic Therapy</subject><subject>Genetic Vectors - genetics</subject><subject>Genomes</subject><subject>Hematopoietic Stem Cell Transplantation</subject><subject>Hematopoietic Stem Cells - virology</subject><subject>Humans</subject><subject>Introns - genetics</subject><subject>Kinases</subject><subject>Leukemia</subject><subject>Medical research</subject><subject>Mice</subject><subject>Mice, Inbred NOD</subject><subject>Mice, SCID</subject><subject>Mutagenesis</subject><subject>Mutagenesis, Insertional - genetics</subject><subject>Retroviridae - genetics</subject><subject>Transcription Initiation Site</subject><subject>Vectors (Biology)</subject><subject>Virus Integration - genetics</subject><issn>1525-0016</issn><issn>1525-0024</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNpFUd1u2yAYtaZNbdf2CSZNSLuOC9hQc7ml3RqpWqL15xZh_JES2eABnpT32gMON1F3xUGcP3SK4hPBJcGEX-3K_ZBeoKQY1yVuSozJu-KMMMoWGNP6_Rsm_LT4GOMuI8IEPylOCWNYMMzOir8rFyEk6x3yBv2CFPwfG1SPnkEnHyKyDv1c31w9LFc3-Xn049SrZN0W3U2DcmgDwY4vMCu-9d53aBP8FpzNWrSEvo9orfWUfTYBDARwyaq-38-2uTt6ttpm8n4OfwzKRZ3tXts8JBVSTtzmS0TKdbNk5XI_Fy-KD0b1ES6P53nx9P32cXm3uF__WC2_3i90zVlaaEIocEOgAV0ZIequ0tSIlvKKCa24aqjuoM6AMN60TX1dqdbUinXCMKPb6rz4cvAdg_89QUxy56fgcqQk14I2hGBeZVZ1YOngY8zflGOwgwp7SbCcl5I7-bqUnJeSuJF5iKz6fPSe2gG6_5rjNJmADgSn0hTgjTCk2YZQUVf_ADApn5U</recordid><startdate>200411</startdate><enddate>200411</enddate><creator>Laufs, Stephanie</creator><creator>Nagy, K Zsuzsanna</creator><creator>Giordano, Frank A</creator><creator>Hotz-Wagenblatt, Agnes</creator><creator>Zeller, W Jens</creator><creator>Fruehauf, Stefan</creator><general>Elsevier Limited</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>3V.</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope></search><sort><creationdate>200411</creationdate><title>Insertion of Retroviral Vectors in NOD/SCID Repopulating Human Peripheral Blood Progenitor Cells Occurs Preferentially in the Vicinity of Transcription Start Regions and in Introns</title><author>Laufs, Stephanie ; 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The retroviral vector SF91m3 was used for transduction of human peripheral blood progenitor cells (PBPC). These PBPC were transplanted into nonobese diabetic/severe combined immunodeficient mice. A total of 186 retroviral vector integration sites were isolated by ligation-mediated PCR from chimeric mouse bone marrow of five PBPC donors, sequenced, and blasted against the human genome. Preferred integration near the transcription start regions, within CpG islands, and within Alu regions was observed. Detailed analysis of targeted RefSeq genes showed a favored integration within the first intron. Integrations were most common in genes coding for signaling proteins, transcription factors, and kinases. In all genes targeted independently multiple times the respective orientation of the provirus within the gene was identical, indicating integration hot spot regions and similar steric determinants for integration sites. Possible explanations for these findings could be nonrandom vector integration, clonal selection due to gene expression interference, or engraftment issues related to gene insertion in signaling and cell cycle genes. The low frequency of integrations in exons may be reassuring as to the safety of retroviral gene therapy with normal human PBPC.</abstract><cop>United States</cop><pub>Elsevier Limited</pub><pmid>15509505</pmid><doi>10.1016/j.ymthe.2004.08.001</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Alu Elements - genetics Animals Bone marrow Cancer research Cell cycle Chromosomes Chromosomes, Human - genetics Computer simulation CpG Islands - genetics Diabetes Gene therapy Gene Transfer Techniques Genetic Therapy Genetic Vectors - genetics Genomes Hematopoietic Stem Cell Transplantation Hematopoietic Stem Cells - virology Humans Introns - genetics Kinases Leukemia Medical research Mice Mice, Inbred NOD Mice, SCID Mutagenesis Mutagenesis, Insertional - genetics Retroviridae - genetics Transcription Initiation Site Vectors (Biology) Virus Integration - genetics |
| title | Insertion of Retroviral Vectors in NOD/SCID Repopulating Human Peripheral Blood Progenitor Cells Occurs Preferentially in the Vicinity of Transcription Start Regions and in Introns |
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