[{"data":1,"prerenderedAt":-1},["ShallowReactive",2],{"doc-seo-378833-105":3,"detail-sidebar-cat-0-en-105":81,"doc-detail-378833-en":130},{"code":4,"msg":5,"data":6},0,"ok",{"site_id":7,"language":8,"slug":9,"title":10,"keywords":11,"description":12,"schema_data":13,"social_meta":74,"head_meta":76,"extra_data":78,"updated_unix":80},105,"en","reference-gene-selection-for-clinical-chimeric-antigen-receptor-t-cell-product-vector-copy-number-assays","Reference gene selection for clinical chimeric antigen receptor T-cell product vector copy number assays","","Reference genes underpin clinical assays such as droplet digital PCR (ddPCR) used to quantify vector copy number integrated into genetically engineered cells and to track plasmid loss in reprogrammed cells for clinical cell therapies. Variability across individuals and between diseased and healthy cells can create thousands of copy-number differences across genomic segments. This study identifies reference genes suitable for ddPCR-based vector copy number variation and plasmid-loss analysis in transduced CAR-T cells and induced pluripotent stem cells.",{"@graph":14,"@context":73},[15,34,56],{"@type":16,"itemListElement":17},"BreadcrumbList",[18,23,27,31],{"item":19,"name":20,"@type":21,"position":22},"https://docshare.wps.com","Home","ListItem",1,{"item":24,"name":25,"@type":21,"position":26},"https://docshare.wps.com/document/","Document",2,{"item":28,"name":29,"@type":21,"position":30},"https://docshare.wps.com/document/healthcare/","Healthcare",3,{"item":32,"name":10,"@type":21,"position":33},"https://docshare.wps.com/document/reference-gene-selection-for-clinical-chimeric-antigen-receptor-t-cell-product-vector-copy-number-assays/378833/",4,{"url":32,"name":10,"@type":35,"image":36,"author":41,"headline":10,"publisher":44,"fileFormat":47,"inLanguage":8,"description":12,"dateModified":48,"datePublished":49,"encodingFormat":47,"isAccessibleForFree":50,"interactionStatistic":51},"DigitalDocument",{"url":37,"@type":38,"width":39,"height":40},"https://docshare.wps.com/thumbnails/reference-gene-selection-for-clinical-chimeric-antigen-receptor-t-cell-product-vector-copy-number-assays/378833.png","ImageObject",300,407,{"name":42,"@type":43},"Oliver","Person",{"url":19,"name":45,"@type":46},"DocShare","Organization","application/pdf","2026-11-02","2026-09-24",true,{"@type":52,"interactionType":53,"userInteractionCount":55},"InteractionCounter",{"@type":54},"ViewAction",8,{"@type":57,"mainEntity":58},"FAQPage",[59,65,69],{"name":60,"@type":61,"acceptedAnswer":62},"Why is selecting reference genes important for ddPCR vector copy number assays?","Question",{"text":63,"@type":64},"Reference genes are essential for clinical ddPCR assays that measure vector copy number and plasmid loss. Selection must account for extensive copy-number variation across individuals and within diseased versus healthy cells.","Answer",{"name":66,"@type":61,"acceptedAnswer":67},"How were candidate reference genes evaluated in this study?",{"text":68,"@type":64},"Candidate genes were assessed using The Cancer Genome Atlas (TCGA) to identify low copy number variance in cancer. ddPCR then measured copy numbers in healthy subjects, cancer cell lines, and patients with multiple cancer types.",{"name":70,"@type":61,"acceptedAnswer":71},"Which genes were concluded to be suitable reference genes for CAR-T vector copy number assays?",{"text":72,"@type":64},"The study concludes that AP3B1, MKL2, and rPP30 are suitable reference genes for vector copy number assays in CAR-T cells from patients with acute leukemia, lymphoma, multiple myeloma, and HPV-associated cancers, with continued evaluation of AGO1.","https://schema.org",{"og:url":32,"og:type":75,"og:title":10,"og:site_name":45,"og:description":12},"article",{"robots":77,"canonical":32},"index,follow",{"doc_id":79,"site_id":7},378833,1790309165,{"code":4,"msg":82,"data":83},"success",[84,88,92,96,101,106,110,114,119,122,126],{"id":22,"doc_module":4,"doc_module_name":25,"category_name":85,"show_sort_weight":86,"slug":87},"Story & Novel",90,"story-novel",{"id":26,"doc_module":4,"doc_module_name":25,"category_name":89,"show_sort_weight":90,"slug":91},"Literature",80,"literature",{"id":33,"doc_module":4,"doc_module_name":25,"category_name":93,"show_sort_weight":94,"slug":95},"Exam",70,"exam",{"id":97,"doc_module":4,"doc_module_name":25,"category_name":98,"show_sort_weight":99,"slug":100},5,"Comic",60,"comic",{"id":102,"doc_module":4,"doc_module_name":25,"category_name":103,"show_sort_weight":104,"slug":105},6,"Technology",50,"technology",{"id":107,"doc_module":4,"doc_module_name":25,"category_name":29,"show_sort_weight":108,"slug":109},7,40,"healthcare",{"id":55,"doc_module":4,"doc_module_name":25,"category_name":111,"show_sort_weight":112,"slug":113},"Research & Report",30,"research-report",{"id":115,"doc_module":4,"doc_module_name":25,"category_name":116,"show_sort_weight":117,"slug":118},9,"Religion & Spirituality",20,"religion-spirituality",{"id":117,"doc_module":4,"doc_module_name":25,"category_name":120,"show_sort_weight":117,"slug":121},"World Cup","world-cup",{"id":123,"doc_module":4,"doc_module_name":25,"category_name":124,"show_sort_weight":123,"slug":125},10,"Lifestyle","lifestyle",{"id":127,"doc_module":4,"doc_module_name":25,"category_name":128,"show_sort_weight":97,"slug":129},19,"General","general",{"code":4,"msg":82,"data":131},{"doc_id":79,"user_id":132,"nickname":42,"user_avatar":133,"doc_module":4,"category_id":107,"category_name":29,"doc_title":10,"doc_description":12,"doc_content":134,"file_id":135,"file_url":136,"file_type":137,"file_size":138,"view_count":55,"is_deleted":4,"is_public":22,"is_downloadable":22,"audit_status":22,"page_count":127,"language":139,"language_code":8,"site_id":7,"html_lang":8,"table_of_contents":140,"faqs":141,"seo_title":142,"seo_description":12,"update_tm":143,"read_time":144},8796095461610,"https://ap-avatar.wpscdn.com/davatar_276721f389ce27ea32af1340a28f341c","Author Manuscript Author Manuscript Author Manuscript Author Manuscript  \n\n| | HHS Public Access\u003Cbr>Author manuscript\u003Cbr>Cytotherapy. Author manuscript; available in PMC 2023 June 01. |\n| --- | --- |\n\nPublished in final edited form as:  \nCytotherapy. 2023 June ; 25(6): 598–604. doi:10.1016/j.jcyt.2023.02.010 .  \nReference gene selection for clinical chimeric antigen receptor T-cell product vector copy number assays  \nJinxia Ma 1 , Lipei Shao 1 , Tatyana Fuksenko 1 , Hui Liu 1 , Rongye Shi 1 , Anh Dinh 1 , Steven L. Highfill 1 , Nan Zhang 1 , Sandhya R. Panch2 , Robert P. Somerville 1 , David F. Stroncek 1 , Ping Jin 1,*  \n1Center for Cellular Engineering, Department of Transfusion Medicine and Center for Cellular Engineering, National Instates of Health Clinical Center, Bethesda, Maryland, USA  \n2 Department of Medicine (Hematology Division), University of Washington/Fred Hutchinson Cancer Center, Seattle, Washington, USA  \nAbstract  \nBackground aims: Reference genes are an essential part of clinical assays such as droplet digital polymerase chain reaction (ddPCR), which measure the number of copies of vector integrated into genetically engineered cells and the loss of plasmids in reprogrammed cells used in clinical cell therapies. Care should be taken to select reference genes, because it has been discovered that there may be thousands of variations in copy number from genomic segments among different individuals. In addition, within the same person in the context of cancer and other proliferative disorders, substantial parts of the genome also can differ in copy number between cells from diseased and healthy people. The purpose of this study was to identify reference genes that could be used for copy number variation analysis of transduced chimeric antigen receptor T cells and for plasmid loss analysis in induced pluripotent stem cells using ddPCR.  \nMethods: We used The Cancer Genome Atlas (TCGA) to evaluate candidate reference genes. If TCGA found a candidate gene to have low copy number variance in cancer, ddPCR was used to measure the copy numbers of the potential reference gene in cells from healthy subjects, cancer cell lines and patients with acute lymphocytic leukemia, lymphoma, multiple myeloma and human papillomavirus–associated cancers.  \nResults: In addition to the rPP30 gene, which we have has been using in our copy number assays, three other candidate reference genes were evaluated using TCGA, and this analysis found that none of the four gene regions (AGO1, AP3B1, MKL2 and rPP30) were amplified or deleted in all of the cancer cell types that are currently being treated with cellular therapies by our facility.  \nThis is an open access article under the CC BY-NC-ND license ([http://creativecommons.org/licenses/by-nc-nd/4.0/](http://creativecommons.org/licenses/by-nc-nd/4.0/))  \n*Correspondence: Pin Jin, Center for Cellular Engineering, Department of Transfusion Medicine and Center for Cellular Engineering, National Instates of Health Clinical Center, 10 Center Drive, Building 10, Room 3N317B, Bethesda, MD 20892. [PJin@mail.cc.nih.gov](PJin@mail.cc.nih.gov) (P. Jin).  \nAuthor Contributions  \nConception and design of the study: JM, PJ and DS. Acquisition of data: JM, TF, HL, JM, TF, RS, AD, SP and NZ. Analysis and interpretation of data: JM, LS, RPS and SH. Drafting or revising the manuscript: AD, SP, NZ, RPS and SH. All authors have approved the final article.  \nDeclaration of Competing Interest  \nThe authors have no commercial, proprietary or financial interest in the products or companies described in this article.  \nAuthor Manuscript Author Manuscript Author Manuscript Author Manuscript  \nMa et al. Page 2  \nThe number of copies of the genes AP3B1, AGO1, rPP30 and MKL2 measured by ddPCR was similar among cells from healthy subjects. We found that AGO1 had copy number alteration in some of the clinical samples, and the number of copies of the genes AP3B1, MKL2 and rPP30 measured by ddPCR was similar among cells ","cbCaigEUnReIP3bS","https://ap.wps.com/l/cbCaigEUnReIP3bS","pdf",824978,"English","# Abstract\n## Background aims\n## Methods\n## Results\n## Conclusions\n# Introduction\n# Reference gene evaluation and ddPCR copy number findings\n# Clinical implications and future evaluation","[{\"question\":\"Why is selecting reference genes important for ddPCR vector copy number assays?\",\"answer\":\"Reference genes are essential for clinical ddPCR assays that measure vector copy number and plasmid loss. Selection must account for extensive copy-number variation across individuals and within diseased versus healthy cells.\"},{\"question\":\"How were candidate reference genes evaluated in this study?\",\"answer\":\"Candidate genes were assessed using The Cancer Genome Atlas (TCGA) to identify low copy number variance in cancer. ddPCR then measured copy numbers in healthy subjects, cancer cell lines, and patients with multiple cancer types.\"},{\"question\":\"Which genes were concluded to be suitable reference genes for CAR-T vector copy number assays?\",\"answer\":\"The study concludes that AP3B1, MKL2, and rPP30 are suitable reference genes for vector copy number assays in CAR-T cells from patients with acute leukemia, lymphoma, multiple myeloma, and HPV-associated cancers, with continued evaluation of AGO1.\"}]","Reference gene selection for clinical chimeric antigen receptor T-cell product vector copy number assays | PDF",1790232608,48]