S52 Development of protocols for mouse GLP-toxicology studies
IntroductionWe have developed a Simian Immunodeficiency Virus (SIV)-based lentiviral vector pseudotyped with the Sendai-virus envelope glycoproteins (F/HN) (rSIV.F/HN) that is effective at transducing pulmonary epithelium in vivo. To prepare for a first-in-man clinical trial, we have developed proto...
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| Veröffentlicht in: | Thorax 2021-02, Vol.76 (Suppl 1), p.A32-A32 |
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| creator | Sinadinos, A Sergijenko, A Meng, C Gamlen, T Hyde, S Gill, DR Griesenbach, U Alton, EWFW |
| description | IntroductionWe have developed a Simian Immunodeficiency Virus (SIV)-based lentiviral vector pseudotyped with the Sendai-virus envelope glycoproteins (F/HN) (rSIV.F/HN) that is effective at transducing pulmonary epithelium in vivo. To prepare for a first-in-man clinical trial, we have developed protocols that can be used in a mouse GLP-toxicology study to efficiently transduce nasal-tissue or lungs.MethodsReporter-imaging, molecular, and radiopharmaceutical tracing methods have been used to quantify intranasally administered lentiviral vector deposition and subsequent tissue-level transgene expression in mice.ResultsNose study. A standard 100 µL vector bolus administration (‘nasal sniffing’) was compared to slow perfusion via catheter (6.7µL/min) and to small volume pipette dosing (2 × 5µL). All animals received 1e7 TU/mouse of rSIV.F/HN expressing luciferase (n=6/group). 10–12 days after transduction, all methods led to similar gene expression in the nasal cavity. Catheter-based delivery led to significant (p |
| doi_str_mv | 10.1136/thorax-2020-BTSabstracts.57 |
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| fullrecord | <record><control><sourceid>proquest_bmj_p</sourceid><recordid>TN_cdi_proquest_journals_2479619473</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>2479619473</sourcerecordid><originalsourceid>FETCH-LOGICAL-b753-2f8c03006c094cbdded4f3bf5f61a4febb1414e79c7acded36caaecd5e8618ca3</originalsourceid><addsrcrecordid>eNpNkEFLAzEUhIMoWKv_YaHn1JdNNtk9iVatQkGhvYckm-gu3aYmWWlvXvyj_hK31IOngZlh3uNDaEJgSgjl1-ndB7XDOeSA71ZLpWMKyqQ4LcQJGhHGS0zzip-iEQADzKng5-gixhYASkLECN0si_zn6_veftq133Z2kzLvsm3wyRu_jpnzIet8H202X7zi5HfNYPu3fRZTXzc2XqIzp9bRXv3pGK0eH1azJ7x4mT_PbhdYi4Li3JUGKAA3UDGj69rWzFHtCseJYs5qTRhhVlRGKDOElBulrKkLW3JSGkXHaHKcHT776G1MsvV92AwXZc5ExUnFBB1a_NjSXSu3oelU2EsC8sBKHlnJAyv5n5UsBP0Fg7JlJQ</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>2479619473</pqid></control><display><type>article</type><title>S52 Development of protocols for mouse GLP-toxicology studies</title><source>Alma/SFX Local Collection</source><creator>Sinadinos, A ; Sergijenko, A ; Meng, C ; Gamlen, T ; Hyde, S ; Gill, DR ; Griesenbach, U ; Alton, EWFW</creator><creatorcontrib>Sinadinos, A ; Sergijenko, A ; Meng, C ; Gamlen, T ; Hyde, S ; Gill, DR ; Griesenbach, U ; Alton, EWFW</creatorcontrib><description>IntroductionWe have developed a Simian Immunodeficiency Virus (SIV)-based lentiviral vector pseudotyped with the Sendai-virus envelope glycoproteins (F/HN) (rSIV.F/HN) that is effective at transducing pulmonary epithelium in vivo. To prepare for a first-in-man clinical trial, we have developed protocols that can be used in a mouse GLP-toxicology study to efficiently transduce nasal-tissue or lungs.MethodsReporter-imaging, molecular, and radiopharmaceutical tracing methods have been used to quantify intranasally administered lentiviral vector deposition and subsequent tissue-level transgene expression in mice.ResultsNose study. A standard 100 µL vector bolus administration (‘nasal sniffing’) was compared to slow perfusion via catheter (6.7µL/min) and to small volume pipette dosing (2 × 5µL). All animals received 1e7 TU/mouse of rSIV.F/HN expressing luciferase (n=6/group). 10–12 days after transduction, all methods led to similar gene expression in the nasal cavity. Catheter-based delivery led to significant (p<0.05) spill-over into the lung and was discontinued. In contrast to nasal sniffing, pipetting of small volumes abrogated lung transgene expression. Technetium radiotracer (5MBq of 99mTc-DTPA/mouse) showed that 98±1% of the pipetted dose was retained in the head. In contrast, nasal sniffing retained only 36±12% in the head, with the rest dispersed into lungs (34±16%) and the remaining body (30±5%). To increase vector delivery to the nasal epithelium, 5 and 10 × 5µL doses (at 5 min intervals) were administered, up to 2.4e8 TU/mouse (n=5/group), and a dose-related increase in gene expression was observed (P<0.0001, r2=0.87). Importantly, even after 10 × 5µL doses, most radiotracer (90±3%) was retained in the head.Lung study. Comparing nasal sniffing to an oropharyngeal (OP) delivery method (1e7 TU/50µL dose for both techniques), there was no difference in gene expression between lungs. By technetium radiotracer (5MBq as before, n=5–6/group), there was no difference in dose distribution between lungs (sniffing: 40±18%, OP: 37±11%). However, nasal sniffing can deliver double the volume compared to OP.Abstract S52 Figure 1Development of protocols for mouse GLP-toxicology studies. A) A Simian Immunodeficiency Virus-based lentiviral vector (rSIV.F/HN) has been administered to mice via intranasal pipette delivery in either a single larger (100 µl) or multiple smaller (5 µl) volumes and the nose and lung vector distributions and expressions compared. B) Whole-body and individual lung luciferase-reporter transgene expression 10–12 days after dosing, showing robust signal in nose and lungs with a single larger volume, undetectable lung signal with a similar titre small volume administration, and an increase in nasal-tissue transgene expression with increasing multiple small volume doses, without significant lung spill-over. RLU = relative light unitsConclusionsTo optimise target organ vector delivery, we have developed protocols suitable for GLP toxicology studies in the murine nose and lung. By defining dose distribution, effective viral titres and overages can be better calculated and compared to proposed clinical doses in future toxicology studies.</description><identifier>ISSN: 0040-6376</identifier><identifier>EISSN: 1468-3296</identifier><identifier>DOI: 10.1136/thorax-2020-BTSabstracts.57</identifier><language>eng</language><publisher>London: BMJ Publishing Group LTD</publisher><subject>Catheters ; Gene expression ; Lungs ; Nose ; Toxicology</subject><ispartof>Thorax, 2021-02, Vol.76 (Suppl 1), p.A32-A32</ispartof><rights>Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.</rights><rights>2021 Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.</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>314,776,780,27903,27904</link.rule.ids></links><search><creatorcontrib>Sinadinos, A</creatorcontrib><creatorcontrib>Sergijenko, A</creatorcontrib><creatorcontrib>Meng, C</creatorcontrib><creatorcontrib>Gamlen, T</creatorcontrib><creatorcontrib>Hyde, S</creatorcontrib><creatorcontrib>Gill, DR</creatorcontrib><creatorcontrib>Griesenbach, U</creatorcontrib><creatorcontrib>Alton, EWFW</creatorcontrib><title>S52 Development of protocols for mouse GLP-toxicology studies</title><title>Thorax</title><description>IntroductionWe have developed a Simian Immunodeficiency Virus (SIV)-based lentiviral vector pseudotyped with the Sendai-virus envelope glycoproteins (F/HN) (rSIV.F/HN) that is effective at transducing pulmonary epithelium in vivo. To prepare for a first-in-man clinical trial, we have developed protocols that can be used in a mouse GLP-toxicology study to efficiently transduce nasal-tissue or lungs.MethodsReporter-imaging, molecular, and radiopharmaceutical tracing methods have been used to quantify intranasally administered lentiviral vector deposition and subsequent tissue-level transgene expression in mice.ResultsNose study. A standard 100 µL vector bolus administration (‘nasal sniffing’) was compared to slow perfusion via catheter (6.7µL/min) and to small volume pipette dosing (2 × 5µL). All animals received 1e7 TU/mouse of rSIV.F/HN expressing luciferase (n=6/group). 10–12 days after transduction, all methods led to similar gene expression in the nasal cavity. Catheter-based delivery led to significant (p<0.05) spill-over into the lung and was discontinued. In contrast to nasal sniffing, pipetting of small volumes abrogated lung transgene expression. Technetium radiotracer (5MBq of 99mTc-DTPA/mouse) showed that 98±1% of the pipetted dose was retained in the head. In contrast, nasal sniffing retained only 36±12% in the head, with the rest dispersed into lungs (34±16%) and the remaining body (30±5%). To increase vector delivery to the nasal epithelium, 5 and 10 × 5µL doses (at 5 min intervals) were administered, up to 2.4e8 TU/mouse (n=5/group), and a dose-related increase in gene expression was observed (P<0.0001, r2=0.87). Importantly, even after 10 × 5µL doses, most radiotracer (90±3%) was retained in the head.Lung study. Comparing nasal sniffing to an oropharyngeal (OP) delivery method (1e7 TU/50µL dose for both techniques), there was no difference in gene expression between lungs. By technetium radiotracer (5MBq as before, n=5–6/group), there was no difference in dose distribution between lungs (sniffing: 40±18%, OP: 37±11%). However, nasal sniffing can deliver double the volume compared to OP.Abstract S52 Figure 1Development of protocols for mouse GLP-toxicology studies. A) A Simian Immunodeficiency Virus-based lentiviral vector (rSIV.F/HN) has been administered to mice via intranasal pipette delivery in either a single larger (100 µl) or multiple smaller (5 µl) volumes and the nose and lung vector distributions and expressions compared. B) Whole-body and individual lung luciferase-reporter transgene expression 10–12 days after dosing, showing robust signal in nose and lungs with a single larger volume, undetectable lung signal with a similar titre small volume administration, and an increase in nasal-tissue transgene expression with increasing multiple small volume doses, without significant lung spill-over. RLU = relative light unitsConclusionsTo optimise target organ vector delivery, we have developed protocols suitable for GLP toxicology studies in the murine nose and lung. By defining dose distribution, effective viral titres and overages can be better calculated and compared to proposed clinical doses in future toxicology studies.</description><subject>Catheters</subject><subject>Gene expression</subject><subject>Lungs</subject><subject>Nose</subject><subject>Toxicology</subject><issn>0040-6376</issn><issn>1468-3296</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2021</creationdate><recordtype>article</recordtype><sourceid>BENPR</sourceid><recordid>eNpNkEFLAzEUhIMoWKv_YaHn1JdNNtk9iVatQkGhvYckm-gu3aYmWWlvXvyj_hK31IOngZlh3uNDaEJgSgjl1-ndB7XDOeSA71ZLpWMKyqQ4LcQJGhHGS0zzip-iEQADzKng5-gixhYASkLECN0si_zn6_veftq133Z2kzLvsm3wyRu_jpnzIet8H202X7zi5HfNYPu3fRZTXzc2XqIzp9bRXv3pGK0eH1azJ7x4mT_PbhdYi4Li3JUGKAA3UDGj69rWzFHtCseJYs5qTRhhVlRGKDOElBulrKkLW3JSGkXHaHKcHT776G1MsvV92AwXZc5ExUnFBB1a_NjSXSu3oelU2EsC8sBKHlnJAyv5n5UsBP0Fg7JlJQ</recordid><startdate>202102</startdate><enddate>202102</enddate><creator>Sinadinos, A</creator><creator>Sergijenko, A</creator><creator>Meng, C</creator><creator>Gamlen, T</creator><creator>Hyde, S</creator><creator>Gill, DR</creator><creator>Griesenbach, U</creator><creator>Alton, EWFW</creator><general>BMJ Publishing Group LTD</general><scope>3V.</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>BENPR</scope><scope>BTHHO</scope><scope>CCPQU</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>K9.</scope><scope>M0S</scope><scope>M1P</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope></search><sort><creationdate>202102</creationdate><title>S52 Development of protocols for mouse GLP-toxicology studies</title><author>Sinadinos, A ; Sergijenko, A ; Meng, C ; Gamlen, T ; Hyde, S ; Gill, DR ; Griesenbach, U ; Alton, EWFW</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-b753-2f8c03006c094cbdded4f3bf5f61a4febb1414e79c7acded36caaecd5e8618ca3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2021</creationdate><topic>Catheters</topic><topic>Gene expression</topic><topic>Lungs</topic><topic>Nose</topic><topic>Toxicology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Sinadinos, A</creatorcontrib><creatorcontrib>Sergijenko, A</creatorcontrib><creatorcontrib>Meng, C</creatorcontrib><creatorcontrib>Gamlen, T</creatorcontrib><creatorcontrib>Hyde, S</creatorcontrib><creatorcontrib>Gill, DR</creatorcontrib><creatorcontrib>Griesenbach, U</creatorcontrib><creatorcontrib>Alton, EWFW</creatorcontrib><collection>ProQuest Central (Corporate)</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central</collection><collection>BMJ Journals</collection><collection>ProQuest One Community College</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><jtitle>Thorax</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Sinadinos, A</au><au>Sergijenko, A</au><au>Meng, C</au><au>Gamlen, T</au><au>Hyde, S</au><au>Gill, DR</au><au>Griesenbach, U</au><au>Alton, EWFW</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>S52 Development of protocols for mouse GLP-toxicology studies</atitle><jtitle>Thorax</jtitle><date>2021-02</date><risdate>2021</risdate><volume>76</volume><issue>Suppl 1</issue><spage>A32</spage><epage>A32</epage><pages>A32-A32</pages><issn>0040-6376</issn><eissn>1468-3296</eissn><abstract>IntroductionWe have developed a Simian Immunodeficiency Virus (SIV)-based lentiviral vector pseudotyped with the Sendai-virus envelope glycoproteins (F/HN) (rSIV.F/HN) that is effective at transducing pulmonary epithelium in vivo. To prepare for a first-in-man clinical trial, we have developed protocols that can be used in a mouse GLP-toxicology study to efficiently transduce nasal-tissue or lungs.MethodsReporter-imaging, molecular, and radiopharmaceutical tracing methods have been used to quantify intranasally administered lentiviral vector deposition and subsequent tissue-level transgene expression in mice.ResultsNose study. A standard 100 µL vector bolus administration (‘nasal sniffing’) was compared to slow perfusion via catheter (6.7µL/min) and to small volume pipette dosing (2 × 5µL). All animals received 1e7 TU/mouse of rSIV.F/HN expressing luciferase (n=6/group). 10–12 days after transduction, all methods led to similar gene expression in the nasal cavity. Catheter-based delivery led to significant (p<0.05) spill-over into the lung and was discontinued. In contrast to nasal sniffing, pipetting of small volumes abrogated lung transgene expression. Technetium radiotracer (5MBq of 99mTc-DTPA/mouse) showed that 98±1% of the pipetted dose was retained in the head. In contrast, nasal sniffing retained only 36±12% in the head, with the rest dispersed into lungs (34±16%) and the remaining body (30±5%). To increase vector delivery to the nasal epithelium, 5 and 10 × 5µL doses (at 5 min intervals) were administered, up to 2.4e8 TU/mouse (n=5/group), and a dose-related increase in gene expression was observed (P<0.0001, r2=0.87). Importantly, even after 10 × 5µL doses, most radiotracer (90±3%) was retained in the head.Lung study. Comparing nasal sniffing to an oropharyngeal (OP) delivery method (1e7 TU/50µL dose for both techniques), there was no difference in gene expression between lungs. By technetium radiotracer (5MBq as before, n=5–6/group), there was no difference in dose distribution between lungs (sniffing: 40±18%, OP: 37±11%). However, nasal sniffing can deliver double the volume compared to OP.Abstract S52 Figure 1Development of protocols for mouse GLP-toxicology studies. A) A Simian Immunodeficiency Virus-based lentiviral vector (rSIV.F/HN) has been administered to mice via intranasal pipette delivery in either a single larger (100 µl) or multiple smaller (5 µl) volumes and the nose and lung vector distributions and expressions compared. B) Whole-body and individual lung luciferase-reporter transgene expression 10–12 days after dosing, showing robust signal in nose and lungs with a single larger volume, undetectable lung signal with a similar titre small volume administration, and an increase in nasal-tissue transgene expression with increasing multiple small volume doses, without significant lung spill-over. RLU = relative light unitsConclusionsTo optimise target organ vector delivery, we have developed protocols suitable for GLP toxicology studies in the murine nose and lung. By defining dose distribution, effective viral titres and overages can be better calculated and compared to proposed clinical doses in future toxicology studies.</abstract><cop>London</cop><pub>BMJ Publishing Group LTD</pub><doi>10.1136/thorax-2020-BTSabstracts.57</doi></addata></record> |
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| subjects | Catheters Gene expression Lungs Nose Toxicology |
| title | S52 Development of protocols for mouse GLP-toxicology studies |
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