Electroporation is a standard laboratory technique for transfection of bacteria, eukaryotic cells, and tissues in vivo. Our laboratories were among the first to develop in vivo plasmid electroporation for therapeutic applications by delivery to preclinical tumors, muscle, and skin and the first that conducted veterinary and clinical gene therapy trials. In vivo electroporation is becoming a well-accepted technique for clinical molecular delivery. Currently, nearly 100 clinical trials are registered in the NIH's clinicaltrials.gov database for electroporation of drugs and nucleic acids. We observed that complete tumor regression can occur when control backbone pDNA is electroporated into different tumors types using various electroporation protocols. Regression is preceded by the production of proinflammatory mRNAs and proteins. Our preliminary results implicate the activation of DNA-specific pattern recognition receptors (PRRs), which normally function for defense against pathogen invasion, in the induction of these proinflammatory proteins. These sensors are found in both immune and non-immune cell types. Activation of these sensors in non-immune cells, such as tumor cells, may contribute to the inflammation and regression that we observed in our previous research and may influence gene therapy efficacy. For example, inflammation may enhance the efficacy of an anti-tumor therapy or an infectious disease vaccine, but interfere with a protein replacement therapy targeting a healthy tissue. Our hypothesis is that DNA-specific PRR activation in non-immune cell types may influence the therapeutic potential of DNA electroporation. Therefore, in this proposal, we will elucidate the activation of PRRs in non-immune cells following different electroporation protocols in several normal cell types and in tumor cells, examine potential activation in a 3-dimensional microenvironment, and relate this pattern to subsequent in vivo effects. Our long-term goal is to be able to predict, based on putative PRR activation and in vivo effects, what electroporation parameters should be used for a specific tissue and therapeutic application. To test our hypothesis, we will conduct our experiments within the following specific aims:
Specific Aim 1. Determine the DNA sensor activation pattern in response to plasmid DNA electroporation using various protocols in non-immune cell types Specific Aim 2. Determine effect of 3-dimensional culture and co-cultures of tumor and normal cells on the DNA sensor activation pattern in response to plasmid DNA electroporation Specific Aim 3. Determine the in vivo DNA sensor activation pattern after plasmid DNA electroporation and its potential influence on gene therapies delivered to tumor, muscle, and skin tissues.

Public Health Relevance

In vivo electroporation is becoming a well-accepted technique for delivery of therapeutic molecules in clinical practice. Our preliminary results suggest that DNA-specific pattern recognition receptors (PRR) are activated after DNA electroporation, producing inflammation that may help or hinder the efficacy of gene therapies. Our long-term goal is to be able to predict, based on putative PRR activation and in vivo effects, what electroporation parameters should be used for a specific tissue and therapeutic application.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
7R01CA196796-05
Application #
10075676
Study Section
Gene and Drug Delivery Systems Study Section (GDD)
Program Officer
Welch, Anthony R
Project Start
2016-02-01
Project End
2021-01-31
Budget Start
2020-02-01
Budget End
2021-01-31
Support Year
5
Fiscal Year
2020
Total Cost
Indirect Cost
Name
University of South Florida
Department
Engineering (All Types)
Type
Biomed Engr/Col Engr/Engr Sta
DUNS #
069687242
City
Tampa
State
FL
Country
United States
Zip Code
33617
Bosnjak, Masa; Kamensek, Urska; Sersa, Gregor et al. (2018) Inhibition of the Innate Immune Receptors for Foreign DNA Sensing Improves Transfection Efficiency of Gene Electrotransfer in Melanoma B16F10 Cells. J Membr Biol 251:179-185
Znidar, Katarina; Bosnjak, Masa; Semenova, Nina et al. (2018) Tumor cell death after electrotransfer of plasmid DNA is associated with cytosolic DNA sensor upregulation. Oncotarget 9:18665-18681
Bosnjak, Masa; Jesenko, Tanja; Kamensek, Urska et al. (2018) Electrotransfer of Different Control Plasmids Elicits Different Antitumor Effectiveness in B16.F10 Melanoma. Cancers (Basel) 10:
Bosnjak, Masa; Kamensek, Urska; Sersa, Gregor et al. (2018) Erratum to: Inhibition of the Innate Immune Receptors for Foreign DNA Sensing Improves Transfection Efficiency of Gene Electrotransfer in Melanoma B16F10 Cells. J Membr Biol 251:187
Znidar, Katarina; Bosnjak, Masa; Jesenko, Tanja et al. (2018) Upregulation of DNA Sensors in B16.F10 Melanoma Spheroid Cells After Electrotransfer of pDNA. Technol Cancer Res Treat 17:1533033818780088
Znidar, Katarina; Bosnjak, Masa; Cemazar, Maja et al. (2016) Cytosolic DNA Sensor Upregulation Accompanies DNA Electrotransfer in B16.F10 Melanoma Cells. Mol Ther Nucleic Acids 5:e322