Efficient GLP-1 gene delivery using two-step transcription amplification plasmid system with a secretion signal peptide and arginine-grafted bioreducible polymer

• Published in: Journal of controlled release 2012-01, Vol.157 (2), p.243-248
• Main Authors: Kim, Tae-il, Lee, Minhyung, Kim, Sung Wan
• Format: Article
• Language: English
• Subjects: Animals; Arginine - chemistry; Arginine-grafted bioreducible polymer; Biological and medical sciences; Cell Line, Tumor; Controlled release; Cytotoxicity; Diabetes mellitus; Enzyme-linked immunosorbent assay; Gene delivery; Gene transfer; General pharmacology; genes; Genetic Therapy - methods; Glucagon-Like Peptide 1 - genetics; Glucagon-Like Peptide 1 - metabolism; Glucagon-like peptide-1; Insulin; Insulin - metabolism; Insulin Secretion; Insulinoma; Medical sciences; Mice; NIH 3T3 Cells; noninsulin-dependent diabetes mellitus; Pharmaceutical technology. Pharmaceutical industry; Pharmacology. Drug treatments; Plasmids; Plasmids - genetics; Polyamines - chemistry; Polymerase chain reaction; polymers; Protein Sorting Signals; reverse transcriptase polymerase chain reaction; Secretion; Secretion signal peptide; secretion signals; signal peptide; Signal peptides; Substance P; therapeutics; Transcription; transcription (genetics); Transcription, Genetic; Transfection; Transfection - methods; Two-step transcription amplification; zeta potential
• ISSN: 0168-3659; 1873-4995
• DOI: 10.1016/j.jconrel.2011.09.072

Glucagon-like peptide (GLP-1) encoding dual plasmid (pDNA) system (TSTA (SP-GLP-1)) which is composed of pβ-Gal4-p65 and pUAS-SP-GLP-1 was constructed to improve the production and secretion of expressed GLP-1 by combining the advantages of signal peptide (SP) and two-step transcription amplification (TSTA) system. Its potential for GLP-1 gene delivery system was investigated with employment of arginine-grafted bioreducible polymer (ABP) as a gene carrier. Their polyplexes have about 140nm-sizes and 20mV Zeta-potential values. ABP showed no cytotoxicity contrary to PEI25k. It was found in RT-PCR experiments that TSTA-SP pDNA systems showed increased GLP-1 gene transcription level in comparison with mono pDNA system (pβ-GLP-1). It was also observed in GLP-1 ELISA that GLP-1 secretion level of TSTA (SP-GLP-1) pDNA system was 2.7–3.4 times higher than those of pβ-GLP-1 and 1.5–1.7 times than TSTA (GLP-1). Additionally, 2.5–3.5 folds increased level of GLP-1 secretion was found in ABP gene carrier system in comparison with PEI25k. When transfection medium containing secreted GLP-1 was transferred to NIT-1 insulinoma cells, the highest secretion level of insulin was induced in ABP/TSTA (SP-GLP-1) polyplex medium-treated cells. Therefore, this novel system could be utilized as a safe and efficient GLP-1 gene delivery system for type 2 diabetes therapy. [Display omitted]