Four Novel Splice-Switch Reporter Cell Lines: Distinct Impact of Oligonucleotide Chemistry and Delivery Vector on Biological Activity

Nucleic Acid Ther. 2016 Dec;26(6):381-391. doi: 10.1089/nat.2016.0631. Epub 2016 Sep 15.


New advances in oligonucleotide (ON) chemistry emerge continuously, and over the last few years, several aspects of ON delivery have been improved. However, clear knowledge regarding how certain chemistries behave alone, or in combination with various delivery vectors, is limited. Moreover, characterization is frequently limited to a single reporter cell line and, when different cell types are studied, experiments are commonly not carried out under similar conditions, hampering comparative analysis. To address this, we have developed a small "tissue" library of new, stable, pLuc/705 splice-switching reporter cell lines (named HuH7_705, U-2 OS_705, C2C12_705, and Neuro-2a_705). Our data show that, indeed, the cell type used in activity screenings influences the efficiency of ONs of different chemistry (phosphorothioate with locked nucleic acid or 2'-O-methyl with or without N,N-diethyl-4-(4-nitronaphthalen-1-ylazo)-phenylamine). Likewise, the delivery method, Lipofectamine® 2000, PepFect14 nanoparticles, or "naked" uptake, also demonstrates cell-type-dependent outcomes. Taken together, these cell lines can potentially become useful tools for future in vitro evaluation of new nucleic acid-based oligomers as well as delivery compounds for splice-switching approaches and cell-specific therapies.

Keywords: delivery vectors; gymnotic delivery; oligonucleotide chemistries; pLuc 705; reporter cell lines; splice-switching oligonucleotides.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Cell Line
  • Cell-Penetrating Peptides / pharmacology
  • Genes, Reporter
  • Genetic Vectors / chemistry
  • Genetic Vectors / metabolism
  • HeLa Cells
  • Hepatocytes / cytology
  • Hepatocytes / metabolism*
  • Humans
  • Lipids / pharmacology
  • Lipopeptides / pharmacology
  • Luciferases / genetics
  • Luciferases / metabolism
  • Mice
  • Myoblasts / cytology
  • Myoblasts / metabolism*
  • Neuroglia / cytology
  • Neuroglia / metabolism*
  • Oligonucleotides / chemical synthesis
  • Oligonucleotides / metabolism*
  • Organ Specificity
  • Osteoblasts / cytology
  • Osteoblasts / metabolism*
  • Phosphorothioate Oligonucleotides / chemical synthesis
  • Phosphorothioate Oligonucleotides / metabolism*
  • Transfection / methods*
  • Transfection / standards


  • Cell-Penetrating Peptides
  • Lipids
  • Lipofectamine
  • Lipopeptides
  • Oligonucleotides
  • PepFect14 peptide
  • Phosphorothioate Oligonucleotides
  • locked nucleic acid
  • Luciferases