PMID- 27138041 OWN - NLM STAT- MEDLINE DCOM- 20170606 LR - 20181113 IS - 1525-0024 (Electronic) IS - 1525-0016 (Print) IS - 1525-0016 (Linking) VI - 24 IP - 7 DP - 2016 Aug TI - Generation of Genetically Engineered Precursor T-Cells From Human Umbilical Cord Blood Using an Optimized Alpharetroviral Vector Platform. PG - 1216-26 LID - 10.1038/mt.2016.89 [doi] AB - Retroviral engineering of hematopoietic stem cell-derived precursor T-cells (preTs) opens the possibility of targeted T-cell transfer across human leukocyte antigen (HLA)-barriers. Alpharetroviral vectors exhibit a more neutral integration pattern thereby reducing the risk of insertional mutagenesis. Cord blood-derived CD34+ cells were transduced and differentiated into preTs in vitro. Two promoters, elongation-factor-1-short-form, and a myeloproliferative sarcoma virus variant in combination with two commonly used envelopes were comparatively assessed choosing enhanced green fluorescent protein or a third-generation chimeric antigen receptor (CAR) against CD123 as gene of interest. Furthermore, the inducible suicide gene iCaspase 9 has been validated. Combining the sarcoma virus-derived promoter with a modified feline endogenous retrovirus envelope glycoprotein yielded in superior transgene expression and transduction rates. Fresh and previously frozen CD34+ cells showed similar transduction and expansion rates. Transgene-positive cells did neither show proliferative impairment nor alteration in their lymphoid differentiation profile. The sarcoma virus-derived promoter only could express sufficient levels of iCaspase 9 to mediate dimerizer-induced apoptosis. Finally, the CD123 CAR was efficiently expressed in CD34+ cells and proved to be functional when expressed on differentiated T-cells. Therefore, the transduction of CD34+ cells with alpharetroviral vectors represents a feasible and potentially safer approach for stem cell-based immunotherapies for cancer. FAU - Hubner, Juwita AU - Hubner J AD - Department of Pediatric Hematology and Oncology, Hannover Medical School, Hannover, Germany. AD - Integrated Research and Treatment Center Transplantation (IFB-Tx), Hannover Medical School, Hannover, Germany. FAU - Hoseini, Shahabuddin S AU - Hoseini SS AD - Department of Pediatric Hematology and Oncology, Hannover Medical School, Hannover, Germany. FAU - Suerth, Julia D AU - Suerth JD AD - Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany. FAU - Hoffmann, Dirk AU - Hoffmann D AD - Integrated Research and Treatment Center Transplantation (IFB-Tx), Hannover Medical School, Hannover, Germany. AD - Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany. FAU - Maluski, Marcel AU - Maluski M AD - Department of Pediatric Hematology and Oncology, Hannover Medical School, Hannover, Germany. FAU - Herbst, Jessica AU - Herbst J AD - Department of Pediatric Hematology and Oncology, Hannover Medical School, Hannover, Germany. FAU - Maul, Holger AU - Maul H AD - Department of Gynecology and Obstetrics, Marienkrankenhaus, Hamburg, Germany. FAU - Ghosh, Arnab AU - Ghosh A AD - Department of Immunology and Medicine, Memorial Sloan-Kettering Cancer Center, New York, USA. FAU - Eiz-Vesper, Britta AU - Eiz-Vesper B AD - Institute for Transfusion Medicine, Hannover Medical School, Hannover, Germany. FAU - Yuan, Qinggong AU - Yuan Q AD - Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany. FAU - Ott, Michael AU - Ott M AD - Department of Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Hannover, Germany. FAU - Heuser, Michael AU - Heuser M AD - Integrated Research and Treatment Center Transplantation (IFB-Tx), Hannover Medical School, Hannover, Germany. AD - Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Hannover, Germany. FAU - Schambach, Axel AU - Schambach A AD - Integrated Research and Treatment Center Transplantation (IFB-Tx), Hannover Medical School, Hannover, Germany. AD - Institute of Experimental Hematology, Hannover Medical School, Hannover, Germany. FAU - Sauer, Martin G AU - Sauer MG AD - Department of Pediatric Hematology and Oncology, Hannover Medical School, Hannover, Germany. AD - Integrated Research and Treatment Center Transplantation (IFB-Tx), Hannover Medical School, Hannover, Germany. LA - eng GR - P30 CA008748/CA/NCI NIH HHS/United States PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20160503 PL - United States TA - Mol Ther JT - Molecular therapy : the journal of the American Society of Gene Therapy JID - 100890581 RN - 0 (Antigens, CD34) RN - 0 (Bacterial Outer Membrane Proteins) RN - 0 (Biomarkers) RN - 0 (Interleukin-3 Receptor alpha Subunit) RN - 0 (Receptors, Antigen, T-Cell) RN - 0 (membrane protein, 75kDa) SB - IM MH - Alpharetrovirus/*genetics MH - Antigens, CD34/metabolism MH - Apoptosis MH - Bacterial Outer Membrane Proteins MH - Biomarkers MH - Cell Differentiation MH - Fetal Blood/*cytology MH - Gene Expression MH - Gene Transfer Techniques MH - Genes, Reporter MH - *Genetic Engineering MH - Genetic Vectors/*genetics MH - Hematopoietic Stem Cells/cytology/metabolism MH - Humans MH - Interleukin-3 Receptor alpha Subunit/immunology MH - Phenotype MH - Precursor Cells, T-Lymphoid/*cytology/*metabolism MH - Promoter Regions, Genetic MH - Receptors, Antigen, T-Cell/genetics/immunology MH - T-Lymphocytes/cytology/immunology/metabolism MH - Transduction, Genetic MH - Transgenes PMC - PMC5088766 EDAT- 2016/05/04 06:00 MHDA- 2017/06/07 06:00 PMCR- 2017/08/01 CRDT- 2016/05/04 06:00 PHST- 2015/07/16 00:00 [received] PHST- 2016/04/13 00:00 [accepted] PHST- 2016/05/04 06:00 [entrez] PHST- 2016/05/04 06:00 [pubmed] PHST- 2017/06/07 06:00 [medline] PHST- 2017/08/01 00:00 [pmc-release] AID - S1525-0016(16)30354-9 [pii] AID - 10.1038/mt.2016.89 [doi] PST - ppublish SO - Mol Ther. 2016 Aug;24(7):1216-26. doi: 10.1038/mt.2016.89. Epub 2016 May 3.