A Modular Albumin-Oligonucleotide Biomolecular Assembly for Delivery of Antisense Therapeutics: Molecular Pharmaceutics

TY - JOUR

T1 - A Modular Albumin-Oligonucleotide Biomolecular Assembly for Delivery of Antisense Therapeutics

T2 - Molecular Pharmaceutics

AU - Elkhashab, Marwa

AU - Dilek, Yeter

AU - Foss, Morten

AU - Creemers, Laura B.

AU - Howard, Kenneth A.

PY - 2024/2

Y1 - 2024/2

N2 - Antisense nucleic acid drugs are susceptible to nuclease degradation, rapid renal clearance, and short circulatory half-life. In this work, we introduce a modular-based recombinant human albumin-oligonucleotide (rHA-cODN) biomolecular assembly that allows incorporation of a chemically stabilized therapeutic gapmer antisense oligonucleotide (ASO) and FcRn-driven endothelial cellular recycling. A phosphodiester ODN linker (cODN) was conjugated to recombinant human albumin (rHA) using maleimide chemistry, after which a complementary gapmer ASO, targeting ADAMTS5 involved in osteoarthritis pathogenesis, was annealed. The rHA-cODN/ASO biomolecular assembly production, fluorescence labeling, and purity were confirmed using polyacrylamide gel electrophoresis. ASO release was triggered by DNase-mediated degradation of the linker strand, reaching 40% in serum after 72 h, with complete release observed following 30 min of incubation with DNase. Cellular internalization and trafficking of the biomolecular assembly using confocal microscopy in C28/I2 cells showed higher uptake and endosomal localization by increasing incubation time from 4 to 24 h. FcRn-mediated cellular recycling of the assembly was demonstrated in FcRn-expressing human microvascular endothelial cells. ADAMTS5 in vitro silencing efficiency reached 40%, which was comparable to free gapmer after 72 h incubation with human osteoarthritis patients’ chondrocytes. This work introduces a versatile biomolecular modular-based “Plug-and-Play” platform potentially applicable for albumin-mediated half-life extension for a range of different types of ODN therapeutics.

AB - Antisense nucleic acid drugs are susceptible to nuclease degradation, rapid renal clearance, and short circulatory half-life. In this work, we introduce a modular-based recombinant human albumin-oligonucleotide (rHA-cODN) biomolecular assembly that allows incorporation of a chemically stabilized therapeutic gapmer antisense oligonucleotide (ASO) and FcRn-driven endothelial cellular recycling. A phosphodiester ODN linker (cODN) was conjugated to recombinant human albumin (rHA) using maleimide chemistry, after which a complementary gapmer ASO, targeting ADAMTS5 involved in osteoarthritis pathogenesis, was annealed. The rHA-cODN/ASO biomolecular assembly production, fluorescence labeling, and purity were confirmed using polyacrylamide gel electrophoresis. ASO release was triggered by DNase-mediated degradation of the linker strand, reaching 40% in serum after 72 h, with complete release observed following 30 min of incubation with DNase. Cellular internalization and trafficking of the biomolecular assembly using confocal microscopy in C28/I2 cells showed higher uptake and endosomal localization by increasing incubation time from 4 to 24 h. FcRn-mediated cellular recycling of the assembly was demonstrated in FcRn-expressing human microvascular endothelial cells. ADAMTS5 in vitro silencing efficiency reached 40%, which was comparable to free gapmer after 72 h incubation with human osteoarthritis patients’ chondrocytes. This work introduces a versatile biomolecular modular-based “Plug-and-Play” platform potentially applicable for albumin-mediated half-life extension for a range of different types of ODN therapeutics.

KW - Antisense oligonucleotide

KW - Albumin

KW - Biomolecular assembly

KW - FcRn

KW - ADAMTS5

KW - osteoarthritis

KW - biomolecular assembly, FcRn

KW - antisense oligonucleotide

KW - albumin

KW - Oligonucleotides, Antisense/chemistry

KW - Humans

KW - Oligonucleotides/chemistry

KW - Deoxyribonucleases

KW - Albumins

KW - Endothelial Cells/metabolism

KW - Serum Albumin, Human/metabolism

KW - Osteoarthritis

U2 - 10.1021/acs.molpharmaceut.3c00561

DO - 10.1021/acs.molpharmaceut.3c00561

M3 - Journal article

C2 - 38214218

SN - 1543-8384

VL - 21

SP - 491

EP - 500

JO - Molecular Pharmaceutics

JF - Molecular Pharmaceutics

IS - 2

ER -