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Gene silencing and antitumoral effects of Eg5 or Ran siRNA oligoaminoamide polyplexes

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Abstract

Two antitumoral siRNAs (directed against target genes Eg5 and Ran) complexed with one of three sequence-defined cationic oligomers were compared in gene silencing in vitro and antitumoral in vivo efficacy upon intratumoral injection. Two lipo-oligomers (T-shape 49, i-shape 229) and the three-arm oligomer 386 were chosen because of their high efficiency in previous marker gene silencing screens. The oligomers showed very similar target-specific gene knockdown in murine neuroblastoma cells. Silencing persisted only for a short period (maximum on day 1 at mRNA and day 2 at protein level) triggering siRNA specific in vitro tumor cell killing. The fastest onset of protein knockdown and strongest antitumoral effect was mediated by oligomer 386. Tumor growth reduction in vivo was evaluated in the subcutaneous Neuro2A mouse model. Intratumoral injections of either Eg5 or Ran siRNA/oligomer 49 polyplexes led to reduced tumor growth and prolonged survival of mice compared to control siRNA and buffer treatment. Target knockdown was evidenced in tumors by mitotic Aster formation for Eg5 knockdown and apoptotic TUNEL stain for Ran knockdown. Ran siRNA displayed better antitumoral efficacy and was chosen for in vivo comparison of the oligomers. A very clear order of antitumoral activity (oligomer 386 > 49 > 229) was observed. In summary, the similar in vitro gene silencing efficiencies on mRNA level by the tested oligomers did not correlate with the observed therapeutic effects in vivo. Oligomer 386 with the fastest onset of protein knockdown and best in vitro cell killing mediated the best in vivo antitumoral efficacy.

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Acknowledgments

This work was supported by the DFG Cluster “Nanosystems Intitiative Munich”, a grant from Axolabs GmbH (formerly Roche Kulmbach GmbH), and the Biotech Cluster m4 T12. We thank Naresh Badgujar and Irene Martin for help with some of the oligomers synthesis.

Conflict of interest

Daniel Edinger, Raphaela Kläger, Christina Troiber, Christian Dohmen, and Ernst Wagner declare that they have no conflict of interest.

Integrity of research and reporting

I confirm that all experiments have been performed according to the German law and the rules of the German Research Foundation (DFG). All institutional and national guidelines for the care and use of laboratory animals were followed.

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Authors and Affiliations

  1. Department of Pharmacy, Pharmaceutical Biotechnology, Center for System-based Drug Research and Center for NanoScience (CeNS), Ludwig-Maximilians-University, Butenandtstrasse 5-13, 81377, Munich, Germany

    Daniel Edinger, Raphaela Kläger, Christina Troiber, Christian Dohmen & Ernst Wagner

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  1. Daniel Edinger
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  2. Raphaela Kläger
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  3. Christina Troiber
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  4. Christian Dohmen
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Correspondence to Daniel Edinger.

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Supporting 1

Intratumoral injection of oligomer 49 Eg5 siRNA polyplexes showed aster formation and confirmed Eg5 knockdown in Neuro2A tumor cells (JPEG 23 kb)

High-resolution image (TIFF 1799 kb)

Supporting 2

Intratumoral injection of oligomer 49 Ran siRNA polyplexes showed positive tunnel staining and confirmed that Ran knockdown led to apoptosis of Neuro2A tumor cells (JPEG 50 kb)

High-resolution image (TIFF 6922 kb)

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Edinger, D., Kläger, R., Troiber, C. et al. Gene silencing and antitumoral effects of Eg5 or Ran siRNA oligoaminoamide polyplexes. Drug Deliv. and Transl. Res. 4, 84–95 (2014). https://doi.org/10.1007/s13346-013-0146-y

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