Soluble biodegradable polymer-based cytokine gene delivery for cancer treatment

Anurag Maheshwari, Ram I. Mahato, John McGregor, Sang Oh Han, Wolfram E. Samlowski, Jong Sang Park, Sung Wan Kim

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79 Scopus citations

Abstract

Transgene expression and tumor regression after direct injection of plasm id DNA encoding cytokine genes, such as mIL-12 and mIFN-γ, remain very low. The objective of this study is to develop nontoxic biodegradable polymer-based cytokine gene delivery systems, which should enhance mIL-12 expression, increasing the likelihood of complete tumor elimination. We synthesized poly[α-(4-aminobutyl)-L-glycolic acid] (PAGA), a biodegradable nontoxic polymer, by melting condensation. Plasmids used in this study encoded luciferase (pLuc) and murine interleukin-12 (pmIL-12) genes. PAGA/plasm id complexes were prepared at different (±) charge ratios and characterized in terms of particle size, zeta potential, osmolality, surface morphology, and cytotoxicity. Polyplexes prepared by complexing PAGA with pmIL-12 as well as pLuc were used for transfert ion into cultured CT-26 colon adenocarcinoma cells as well as into CT-26 tumor-bearing BALB/c mice. The in vitro and in vivo transfection efficiency was determined by luciferase assay (for pLuc), enzyme-linked immunosorbent assay (for mIL-12, p70, and p40), and reverse transcriptase-polymerase chain reaction (RT-PCR) (for Luc and mIL-12 p35). PAGA condensed and protected plasmids from nuclease degradation. The mean particle size and zeta potential of the polyplexes prepared in 5% (w/v) glucose at 3:1 (±) charge ratio were approximately 100 nm and 20 mV, respectively. The surface characterization of polyplexes as determined by atomic force microscopy showed complete condensation of DNA with an ellipsoidal structure in Z direction. The levels of mIL-12 p40, mIL-12 p70, and mIFN-γ were significantly higher for PAGA/pmIL-12 complexes compared to that of naked pmIL-12. This is in good agreement with RT-PCR data, which showed significant levels of mIL-12 p35 expression. The PAGA/pmIL-12 complexes did not induce any cytotoxicity in CT-26 cells as evidenced by 3-{4,5-dimethylthiazol-2-yl}-2,5-diphenylte-trazolium bromide assay and showed enhanced antitumor activity in vivo compared to naked pmIL-12. PAGA/pmIL-12 complexes are nontoxic and significantly enhance mIL-12 expression at mRNA and protein levels both in vitro and in vivo.

Original languageEnglish (US)
Pages (from-to)121-130
Number of pages10
JournalMolecular Therapy
Volume2
Issue number2
DOIs
Publication statusPublished - Aug 2000

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Keywords

  • Biodegradable cationic polymer
  • ELISA
  • Gene expression
  • Interleukin-12
  • Intratumoral delivery
  • RT-PCR

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Pharmacology
  • Drug Discovery

Cite this

Maheshwari, A., Mahato, R. I., McGregor, J., Han, S. O., Samlowski, W. E., Park, J. S., & Kim, S. W. (2000). Soluble biodegradable polymer-based cytokine gene delivery for cancer treatment. Molecular Therapy, 2(2), 121-130. https://doi.org/10.1006/mthe.2000.0105