1. Maintenance and gene electrotransfer efficiency of antibiotic resistance gene-free plasmids encoding mouse, canine and human interleukin-12 orthologuesUrška Kamenšek, Andrej Renčelj, Tanja Jesenko, Tinkara Remic, Gregor Serša, Maja Čemažar, 2022, original scientific article Abstract: 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. Keywords: electrotransfer, interleukin-12, immunotherapy, mammals Published in DiRROS: 23.09.2022; Views: 760; Downloads: 374
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2. Tumor cell-based vaccine contributes to local tumor irradiation by eliciting a tumor model-dependent systemic immune responseTinkara Remic, Gregor Serša, Kristina Levpušček, Urša Lampreht Tratar, Katja Uršič Valentinuzzi, Andrej Cör, Urška Kamenšek, 2022, original scientific article Abstract: Multimodal treatment approaches, such as radio-immunotherapy, necessitate regimen optimization and the investigation of the interactions of different modalities. The aim of this study was two-fold. Firstly, to select the most effective combination of irradiation and the previously developed tumor cell-based vaccine and then to provide insight into the immune response to the selected combinatorial treatment. The study was performed in immunologically different murine tumor models: B16F10 melanoma and CT26 colorectal carcinoma. The most effective combinatorial treatment was selected by comparing three different IR regimens and three different vaccination regimens. We determined the local immune response by investigating immune cell infiltration at the vaccination site and in tumors. Lastly, we determined the systemic immune response by investigating the amount of tumor-specific effector lymphocytes in draining lymph nodes. The selected most effective combinatorial treatment was 5× 5 Gy in combination with concomitant single-dose vaccination (B16F10) or with concomitant multi-dose vaccination (CT26). The combinatorial treatment successfully elicited a local immune response at the vaccination site and in tumors in both tumor models. It also resulted in the highest amount of tumor-specific effector lymphocytes in draining lymph nodes in the B16F10, but not in the CT26 tumor-bearing mice. However, the amount of tumor-specific effector lymphocytes was intrinsically higher in the CT26 than in the B16F10 tumor model. Upon the selection of the most effective combinatorial treatment, we demonstrated that the vaccine elicits an immune response and contributes to the antitumor efficacy of tumor irradiation. However, this interaction is multi-faceted and appears to be dependent on the tumor immunogenicity. Keywords: experimental oncology, multimodal treatment, radio-imunotherapy Published in DiRROS: 14.09.2022; Views: 746; Downloads: 398
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3. Razvoj avtologne tumorske vakcine pripravljene z obsevanjemTinkara Remic, Gregor Serša, Katja Uršič Valentinuzzi, Kristina Levpušček, Urša Lampreht Tratar, Maja Čemažar, Urška Kamenšek, 2022, published scientific conference contribution abstract Keywords: onkološka cepiva, tumorski antigeni, eksperimentalna onkologija Published in DiRROS: 17.06.2022; Views: 866; Downloads: 224
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4. Priprava in testiranje tumorske vakcine z obsevanjemTinkara Remic, Gregor Serša, Maja Čemažar, Katja Uršič Valentinuzzi, Kristina Levpušček, Urška Kamenšek, 2021, published scientific conference contribution Keywords: genski elektroprenos, obsevanje, tumorska vakcina Published in DiRROS: 01.04.2022; Views: 764; Downloads: 229
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