20.500.12556/DiRROS-15587 Maintenance and gene electrotransfer efficiency of antibiotic resistance gene-free plasmids encoding mouse, canine and human interleukin-12 orthologues Interleukin 12 (IL-12) is a cytokine used as a therapeutic molecule in cancer immunotherapy. Gene electrotransfer mediated delivery of IL-12 gene has reached clinical evaluation in the USA using a plasmid that in addition to IL- 12 gene also carry an antibiotic resistance gene needed for its production in bacteria. In Europe however, Eu- ropean Medicines Agency recommends against the use of antibiotics during the production of clinical grade plasmids. We have prepared several antibiotic resistance gene-free plasmids using an antibiotic-free selection strategy called operator-repressor titration, including plasmids encoding mouse, canine and human IL-12 orthologues. The aim of this study was to evaluate the maintenance of these plasmids in bacterial culture and test their transfection efficiency using gene electrotransfer. Plasmid maintenance was evaluated by determining plasmid yields and topologies after subculturing transformed bacteria. Transfection efficiency was evaluated by determining the plasmid copy number, expression and cytotoxicity after gene electrotransfer to mouse, canine and human melanoma cells. The results demonstrated that our IL-12 plasmids without an antibiotic resistance gene are stably maintained in bacteria and provide sufficient IL-12 expression after in vitro gene electrotransfer; therefore, they have the potential to proceed to further in vivo evaluation studies. electrotransfer interleukin-12 immunotherapy mammals elektroprenos genov interlevkin 12 imunoterapija sesalci true false true Elsevier Angleški jezik Slovenski jezik by Authors Neznano 2022-09-23 03:34:36 2022-09-23 13:13:16 2022-09-24 03:44:04 0000-00-00 00:00:00 2022 0 0 Nasl. z nasl. zaslona; Opis vira z dne 7. 2. 2022; str. 1-08879-10-08879 no. 2 Vol. 8 Feb. 2022 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 2022-02-01 602.6/.7 2405-8440 10.1016/j.heliyon.2022.e08879 96531715 https://doi.org/10.1016/j.heliyon.2022.e08879 1 1a302e45-3acf-11ed-bea5-001a4af901a5 https://dirros.openscience.si/Dokument.php?lang=slv&id=20611 RAZ_Kamensek_Urska_2022.pdf RAZ_Kamensek_Urska_2022.pdf 1 BC8F0E9397F9B6850113EC4CC62CB37B f1505b41320a2847043c9a28c37553b7f5d007373cd2088b30afecca6256c91c 442d85fb-3b20-11ed-bea5-001a4af901a5 https://dirros.openscience.si/Dokument.php?lang=slv&id=20617 Onkološki inštitut Ljubljana 0 0 0