An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing

Konrad Bienk, Michael Lykke Hvam, Malgorzata Maria Pakula, Frederik Dagnæs-Hansen, Jesper Wengel, Birgitte Mølholm Malle, Ulrich Kragh-Hansen, Jason Cameron, Jens Thostrup Bukrinski, Kenneth A Howard

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd > 1 × 10- 7 M) dependent on number of modifications. The rHSA/siRNA complex exhibited reduced nuclease degradation and reduced induction of TNF-α production by human peripheral blood mononuclear cells. The increased solubility of heavily cholesteryl modified siRNA in the presence of rHSA facilitated duplex annealing and consequent interaction that allowed in vivo studies using multiple cholesteryl modifications. A structural-activity-based screen of in vitro EGFP-silencing was used to select optimal siRNA designs containing cholesteryl modifications within the sense strand that were used for in vivo studies. We demonstrate plasma half-life extension in NMRI mice from t1/2 12 min (naked) to t1/2 45 min (single cholesteryl) and t1/2 71 min (double cholesteryl) using fluorescent live bioimaging. The biodistribution showed increased accumulation in the liver for the double cholesteryl modified siRNA that correlated with an increase in hepatic Factor VII gene silencing of 28% (rHSA/siRNA) compared to 4% (naked siRNA) 6 days post-injection. This work presents a novel albumin-mediated cholesteryl design-based strategy for tuning pharmacokinetics and systemic gene silencing.

Original languageEnglish
JournalJournal of Controlled Release
Volume232
Pages (from-to)143-151
ISSN0168-3659
DOIs
Publication statusPublished - 2016

Fingerprint

RNA Interference
Small Interfering RNA
Albumins
Pharmacokinetics
Half-Life
Liver
Life Expectancy

Keywords

  • Albumin
  • Biodistribution
  • Drug delivery
  • Factor VII
  • Half-life extension
  • siRNA

Cite this

Bienk, Konrad ; Hvam, Michael Lykke ; Pakula, Malgorzata Maria ; Dagnæs-Hansen, Frederik ; Wengel, Jesper ; Malle, Birgitte Mølholm ; Kragh-Hansen, Ulrich ; Cameron, Jason ; Bukrinski, Jens Thostrup ; Howard, Kenneth A. / An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing. In: Journal of Controlled Release. 2016 ; Vol. 232. pp. 143-151.
@article{e5bab0a9ebf24fc39bbe7f8e48eb4694,
title = "An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing",
abstract = "Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd > 1 × 10- 7 M) dependent on number of modifications. The rHSA/siRNA complex exhibited reduced nuclease degradation and reduced induction of TNF-α production by human peripheral blood mononuclear cells. The increased solubility of heavily cholesteryl modified siRNA in the presence of rHSA facilitated duplex annealing and consequent interaction that allowed in vivo studies using multiple cholesteryl modifications. A structural-activity-based screen of in vitro EGFP-silencing was used to select optimal siRNA designs containing cholesteryl modifications within the sense strand that were used for in vivo studies. We demonstrate plasma half-life extension in NMRI mice from t1/2 12 min (naked) to t1/2 45 min (single cholesteryl) and t1/2 71 min (double cholesteryl) using fluorescent live bioimaging. The biodistribution showed increased accumulation in the liver for the double cholesteryl modified siRNA that correlated with an increase in hepatic Factor VII gene silencing of 28{\%} (rHSA/siRNA) compared to 4{\%} (naked siRNA) 6 days post-injection. This work presents a novel albumin-mediated cholesteryl design-based strategy for tuning pharmacokinetics and systemic gene silencing.",
keywords = "Albumin, Biodistribution, Drug delivery, Factor VII, Half-life extension, siRNA",
author = "Konrad Bienk and Hvam, {Michael Lykke} and Pakula, {Malgorzata Maria} and Frederik Dagn{\ae}s-Hansen and Jesper Wengel and Malle, {Birgitte M{\o}lholm} and Ulrich Kragh-Hansen and Jason Cameron and Bukrinski, {Jens Thostrup} and Howard, {Kenneth A}",
year = "2016",
doi = "10.1016/j.jconrel.2016.04.013",
language = "English",
volume = "232",
pages = "143--151",
journal = "Journal of Controlled Release",
issn = "0168-3659",
publisher = "Elsevier",

}

Bienk, K, Hvam, ML, Pakula, MM, Dagnæs-Hansen, F, Wengel, J, Malle, BM, Kragh-Hansen, U, Cameron, J, Bukrinski, JT & Howard, KA 2016, 'An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing', Journal of Controlled Release, vol. 232, pp. 143-151. https://doi.org/10.1016/j.jconrel.2016.04.013

An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing. / Bienk, Konrad; Hvam, Michael Lykke; Pakula, Malgorzata Maria; Dagnæs-Hansen, Frederik; Wengel, Jesper; Malle, Birgitte Mølholm; Kragh-Hansen, Ulrich; Cameron, Jason; Bukrinski, Jens Thostrup; Howard, Kenneth A.

In: Journal of Controlled Release, Vol. 232, 2016, p. 143-151.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing

AU - Bienk, Konrad

AU - Hvam, Michael Lykke

AU - Pakula, Malgorzata Maria

AU - Dagnæs-Hansen, Frederik

AU - Wengel, Jesper

AU - Malle, Birgitte Mølholm

AU - Kragh-Hansen, Ulrich

AU - Cameron, Jason

AU - Bukrinski, Jens Thostrup

AU - Howard, Kenneth A

PY - 2016

Y1 - 2016

N2 - Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd > 1 × 10- 7 M) dependent on number of modifications. The rHSA/siRNA complex exhibited reduced nuclease degradation and reduced induction of TNF-α production by human peripheral blood mononuclear cells. The increased solubility of heavily cholesteryl modified siRNA in the presence of rHSA facilitated duplex annealing and consequent interaction that allowed in vivo studies using multiple cholesteryl modifications. A structural-activity-based screen of in vitro EGFP-silencing was used to select optimal siRNA designs containing cholesteryl modifications within the sense strand that were used for in vivo studies. We demonstrate plasma half-life extension in NMRI mice from t1/2 12 min (naked) to t1/2 45 min (single cholesteryl) and t1/2 71 min (double cholesteryl) using fluorescent live bioimaging. The biodistribution showed increased accumulation in the liver for the double cholesteryl modified siRNA that correlated with an increase in hepatic Factor VII gene silencing of 28% (rHSA/siRNA) compared to 4% (naked siRNA) 6 days post-injection. This work presents a novel albumin-mediated cholesteryl design-based strategy for tuning pharmacokinetics and systemic gene silencing.

AB - Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd > 1 × 10- 7 M) dependent on number of modifications. The rHSA/siRNA complex exhibited reduced nuclease degradation and reduced induction of TNF-α production by human peripheral blood mononuclear cells. The increased solubility of heavily cholesteryl modified siRNA in the presence of rHSA facilitated duplex annealing and consequent interaction that allowed in vivo studies using multiple cholesteryl modifications. A structural-activity-based screen of in vitro EGFP-silencing was used to select optimal siRNA designs containing cholesteryl modifications within the sense strand that were used for in vivo studies. We demonstrate plasma half-life extension in NMRI mice from t1/2 12 min (naked) to t1/2 45 min (single cholesteryl) and t1/2 71 min (double cholesteryl) using fluorescent live bioimaging. The biodistribution showed increased accumulation in the liver for the double cholesteryl modified siRNA that correlated with an increase in hepatic Factor VII gene silencing of 28% (rHSA/siRNA) compared to 4% (naked siRNA) 6 days post-injection. This work presents a novel albumin-mediated cholesteryl design-based strategy for tuning pharmacokinetics and systemic gene silencing.

KW - Albumin

KW - Biodistribution

KW - Drug delivery

KW - Factor VII

KW - Half-life extension

KW - siRNA

U2 - 10.1016/j.jconrel.2016.04.013

DO - 10.1016/j.jconrel.2016.04.013

M3 - Journal article

C2 - 27084489

AN - SCOPUS:84964632116

VL - 232

SP - 143

EP - 151

JO - Journal of Controlled Release

JF - Journal of Controlled Release

SN - 0168-3659

ER -