Gene transfer-mediated pre-mRNA segmental trans-splicing as a strategy to deliver intracellular toxins for cancer therapy. Academic Article uri icon

Overview

MeSH

  • Alternative Splicing
  • Base Sequence
  • Carcinoma, Squamous Cell
  • Cell Line, Tumor
  • DNA Primers
  • DNA, Complementary
  • Exons
  • Gene Transfer Techniques
  • HeLa Cells
  • Humans
  • Lung Neoplasms
  • Reverse Transcriptase Polymerase Chain Reaction

MeSH Major

  • Neoplasms
  • RNA Precursors
  • RNA Splicing
  • Shiga Toxin
  • Toxins, Biological

abstract

  • Virus-mediated transfer of genes coding for intracellular toxins holds promise for cancer therapy, but the inherent toxicity of such vectors make them a risk to normal tissues and a challenge to produce due to the intrinsic dilemma that expression of toxin molecules kills producer cells. We employed pre-mRNA segmental trans-splicing (STS), in which two engineered DNA fragments coding for 5' "donor" and 3' "acceptor" segments of a toxin gene, respectively, are expressed by viral vectors. When co-delivered to target cells, the two vectors generate two toxin pre-mRNA fragments which are spliced by the target cell machinery to produce functional mRNA and toxin. To test this approach, we used an enzymatic fragment of Shigatoxin1A1 (STX1A1) known to provoke apoptotic cell death. Two adenovirus vectors, Shigatoxin1A1 donor (AdStx1A1Do) and Shigatoxin1A1 acceptor (AdStx1A1Ac), respectively, were used to deliver the Stx1A1 gene fragments. HeLa, HEp2, and A549 cells transfected with AdStx1A1Do and AdStx1A1Ac had a dose-dependent reduction in viability and inhibition of protein synthesis. Intratumoral injection of AdStx1A1Do and AdStx1A1Ac into preexisting HeLa, Hep2, and A549 tumors in immunodeficient mice revealed significant inhibition of tumor growth. There was no evidence of liver damage, suggesting that there was no leakage of vector or toxin from the site of injection following intratumoral injection of AdStx1A1Do and AdStx1A1Ac. These results suggest that the obstacles preventing gene transfer of intracellular toxins for local cancer therapy could be overcome by pre-mRNA segmental trans-splicing.

publication date

  • January 1, 2005

has subject area

  • Alternative Splicing
  • Base Sequence
  • Carcinoma, Squamous Cell
  • Cell Line, Tumor
  • DNA Primers
  • DNA, Complementary
  • Exons
  • Gene Transfer Techniques
  • HeLa Cells
  • Humans
  • Lung Neoplasms
  • Neoplasms
  • RNA Precursors
  • RNA Splicing
  • Reverse Transcriptase Polymerase Chain Reaction
  • Shiga Toxin
  • Toxins, Biological

Research

keywords

  • Journal Article

Identity

Language

  • eng

PubMed ID

  • 15665302

Additional Document Info

start page

  • 254

end page

  • 263

volume

  • 65

number

  • 1