利用腺病毒技术和靶向超声成像验证受控受体表达

Reshu Saini, J. Warram, A. Sorace, H. Umphrey, K. Zinn, K. Hoyt
{"title":"利用腺病毒技术和靶向超声成像验证受控受体表达","authors":"Reshu Saini, J. Warram, A. Sorace, H. Umphrey, K. Zinn, K. Hoyt","doi":"10.1109/ULTSYM.2010.5935763","DOIUrl":null,"url":null,"abstract":"This paper details a model system for evaluating targeted ultrasound (US) contrast agents using adenoviral (Ad) vectors for controlled receptor expression. Breast cancer cell receptor density in vitro was modulated by varying the multiplicity of infection (MOI) from 0 to 100. Target receptors were induced using a GFP-positive Ad vector for gene transfer and expression of the human somatostatin receptor subtype 2a (hSSTr2) with a hemagglutinin (HA) tag. Subsequently, receptor expression and anti-HA antibody (Ab) binding was examined with flow cytometry. Targeted US contrast agents (MB) were created by conjugating either biotinylated anti-HA or isotype control Ab to the surface of biotin coated MBs via a streptavidin bridge. Targeted MBs were incubated with Ad infected cells with to test in vitro MB binding. An in vivo study was performed in tumor-bearing nude athymic mice with induced HA-hSSTr2-GFP receptor expression by intratumoral injection of the Ad vector (MOI of 100). Mice were sorted and injected via the tail vein with the two MB groups followed by US imaging. 24 hrs later mice groups were switched and the MB study repeated. Experimental in vitro results found GFP expression to be directly correlated with Ad MOI (R2 = 0.96). Increasing the Ad MOI produced a corresponding increase in binding and accumulation of anti-HA Ab on the cell surface (P < 0.01). However, no differences was found between Cy5.5-labeled anti-HA Ab exposed cell groups at an MOI of 0 (P > 0.29) or in the control Ab group (P > 0.44) indicating minimal nonspecific binding. No difference was found between cells groups incubated with control MBs (P > 0.42) regardless of receptor density. However, cells exposed to targeted MBs showed increased levels of cell binding proportional to receptor expression levels (P < 0.02). Images taken from in vivo experiments were analyzed by two blinded reviewers in consensus to compare intratumoral MB accumulation. It was concluded that 78% (7 of 9) of US images from the targeted MB group exhibited increased local intratumoral accumulation compared to the control group. Overall, this study demonstrates use of an Ad vector for selectively controlling cellular expression and modulation of the receptor density. Furthermore, results demonstrate targeted MB accumulation at the receptor site.","PeriodicalId":6437,"journal":{"name":"2010 IEEE International Ultrasonics Symposium","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2010-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Validation of controlled receptor expression using adenoviral techniques and targeted ultrasound imaging\",\"authors\":\"Reshu Saini, J. Warram, A. Sorace, H. Umphrey, K. Zinn, K. Hoyt\",\"doi\":\"10.1109/ULTSYM.2010.5935763\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"This paper details a model system for evaluating targeted ultrasound (US) contrast agents using adenoviral (Ad) vectors for controlled receptor expression. Breast cancer cell receptor density in vitro was modulated by varying the multiplicity of infection (MOI) from 0 to 100. Target receptors were induced using a GFP-positive Ad vector for gene transfer and expression of the human somatostatin receptor subtype 2a (hSSTr2) with a hemagglutinin (HA) tag. Subsequently, receptor expression and anti-HA antibody (Ab) binding was examined with flow cytometry. Targeted US contrast agents (MB) were created by conjugating either biotinylated anti-HA or isotype control Ab to the surface of biotin coated MBs via a streptavidin bridge. Targeted MBs were incubated with Ad infected cells with to test in vitro MB binding. An in vivo study was performed in tumor-bearing nude athymic mice with induced HA-hSSTr2-GFP receptor expression by intratumoral injection of the Ad vector (MOI of 100). Mice were sorted and injected via the tail vein with the two MB groups followed by US imaging. 24 hrs later mice groups were switched and the MB study repeated. Experimental in vitro results found GFP expression to be directly correlated with Ad MOI (R2 = 0.96). Increasing the Ad MOI produced a corresponding increase in binding and accumulation of anti-HA Ab on the cell surface (P < 0.01). However, no differences was found between Cy5.5-labeled anti-HA Ab exposed cell groups at an MOI of 0 (P > 0.29) or in the control Ab group (P > 0.44) indicating minimal nonspecific binding. No difference was found between cells groups incubated with control MBs (P > 0.42) regardless of receptor density. However, cells exposed to targeted MBs showed increased levels of cell binding proportional to receptor expression levels (P < 0.02). Images taken from in vivo experiments were analyzed by two blinded reviewers in consensus to compare intratumoral MB accumulation. It was concluded that 78% (7 of 9) of US images from the targeted MB group exhibited increased local intratumoral accumulation compared to the control group. Overall, this study demonstrates use of an Ad vector for selectively controlling cellular expression and modulation of the receptor density. Furthermore, results demonstrate targeted MB accumulation at the receptor site.\",\"PeriodicalId\":6437,\"journal\":{\"name\":\"2010 IEEE International Ultrasonics Symposium\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2010-10-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"2010 IEEE International Ultrasonics Symposium\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.1109/ULTSYM.2010.5935763\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"2010 IEEE International Ultrasonics Symposium","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.1109/ULTSYM.2010.5935763","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

摘要

本文详细介绍了一种利用腺病毒(Ad)载体控制受体表达来评估靶向超声(US)造影剂的模型系统。通过改变感染的多重性(MOI)从0到100来调节体外乳腺癌细胞受体密度。使用gfp阳性的Ad载体诱导靶受体进行基因转移,并表达带有血凝素(HA)标签的人生长抑素受体亚型2a (hSSTr2)。随后用流式细胞术检测受体表达和抗ha抗体(Ab)结合。通过链亲和素桥将生物素化的抗ha或同型对照Ab偶联到生物素包被的MB表面,形成靶向US对比剂(MB)。将靶向MB与Ad感染的细胞孵育,检测MB的体外结合。通过瘤内注射Ad载体(MOI为100)诱导HA-hSSTr2-GFP受体在荷瘤裸鼠体内的表达研究。小鼠分选后经尾静脉注射两组MB,并行超声成像。24小时后切换小鼠组,重复MB研究。体外实验结果显示,GFP表达与Ad MOI直接相关(R2 = 0.96)。随着Ad MOI的增加,抗ha抗体在细胞表面的结合和积累也相应增加(P < 0.01)。然而,在MOI为0时,cy5.5标记的抗ha Ab暴露细胞组(P > 0.29)和对照Ab组(P > 0.44)之间没有发现差异,表明非特异性结合最小。无论受体密度如何,与对照MBs孵育的细胞组间无差异(P > 0.42)。然而,暴露于靶向MBs的细胞显示出与受体表达水平成正比的细胞结合水平增加(P < 0.02)。从体内实验中获得的图像由两位盲法评论者一致分析,以比较肿瘤内MB的积累。结论是,与对照组相比,靶向MB组78%(7 / 9)的US图像显示局部瘤内积聚增加。总的来说,本研究证明了使用Ad载体选择性地控制细胞表达和受体密度的调节。此外,结果表明靶向MB积累在受体部位。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Validation of controlled receptor expression using adenoviral techniques and targeted ultrasound imaging
This paper details a model system for evaluating targeted ultrasound (US) contrast agents using adenoviral (Ad) vectors for controlled receptor expression. Breast cancer cell receptor density in vitro was modulated by varying the multiplicity of infection (MOI) from 0 to 100. Target receptors were induced using a GFP-positive Ad vector for gene transfer and expression of the human somatostatin receptor subtype 2a (hSSTr2) with a hemagglutinin (HA) tag. Subsequently, receptor expression and anti-HA antibody (Ab) binding was examined with flow cytometry. Targeted US contrast agents (MB) were created by conjugating either biotinylated anti-HA or isotype control Ab to the surface of biotin coated MBs via a streptavidin bridge. Targeted MBs were incubated with Ad infected cells with to test in vitro MB binding. An in vivo study was performed in tumor-bearing nude athymic mice with induced HA-hSSTr2-GFP receptor expression by intratumoral injection of the Ad vector (MOI of 100). Mice were sorted and injected via the tail vein with the two MB groups followed by US imaging. 24 hrs later mice groups were switched and the MB study repeated. Experimental in vitro results found GFP expression to be directly correlated with Ad MOI (R2 = 0.96). Increasing the Ad MOI produced a corresponding increase in binding and accumulation of anti-HA Ab on the cell surface (P < 0.01). However, no differences was found between Cy5.5-labeled anti-HA Ab exposed cell groups at an MOI of 0 (P > 0.29) or in the control Ab group (P > 0.44) indicating minimal nonspecific binding. No difference was found between cells groups incubated with control MBs (P > 0.42) regardless of receptor density. However, cells exposed to targeted MBs showed increased levels of cell binding proportional to receptor expression levels (P < 0.02). Images taken from in vivo experiments were analyzed by two blinded reviewers in consensus to compare intratumoral MB accumulation. It was concluded that 78% (7 of 9) of US images from the targeted MB group exhibited increased local intratumoral accumulation compared to the control group. Overall, this study demonstrates use of an Ad vector for selectively controlling cellular expression and modulation of the receptor density. Furthermore, results demonstrate targeted MB accumulation at the receptor site.
求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信