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A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides

We have developed a non-cationic transfection vector in the form of bottlebrush polymer-antisense oligonucleotide (ASO) conjugates. Termed pacDNA (polymer-assisted compaction of DNA), these agents show improved biopharmaceutical characteristics and antisense potency in vivo while suppressing non-ant...

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Autores principales: Zhang, Lei, Wang, Yuyan, Chen, Peiru, Wang, Dali, Sun, Tingyu, Zhang, Zheyu, Wang, Ruimeng, Kang, Xi, Fang, Yang, Lu, Hao, Cai, Jiansong, Ren, Mengqi, Dong, Sijia S., Zhang, Ke
Formato: Online Artículo Texto
Lenguaje:English
Publicado: RSC 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9906284/
https://www.ncbi.nlm.nih.gov/pubmed/36794022
http://dx.doi.org/10.1039/d2cb00149g
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author Zhang, Lei
Wang, Yuyan
Chen, Peiru
Wang, Dali
Sun, Tingyu
Zhang, Zheyu
Wang, Ruimeng
Kang, Xi
Fang, Yang
Lu, Hao
Cai, Jiansong
Ren, Mengqi
Dong, Sijia S.
Zhang, Ke
author_facet Zhang, Lei
Wang, Yuyan
Chen, Peiru
Wang, Dali
Sun, Tingyu
Zhang, Zheyu
Wang, Ruimeng
Kang, Xi
Fang, Yang
Lu, Hao
Cai, Jiansong
Ren, Mengqi
Dong, Sijia S.
Zhang, Ke
author_sort Zhang, Lei
collection PubMed
description We have developed a non-cationic transfection vector in the form of bottlebrush polymer-antisense oligonucleotide (ASO) conjugates. Termed pacDNA (polymer-assisted compaction of DNA), these agents show improved biopharmaceutical characteristics and antisense potency in vivo while suppressing non-antisense side effects. Nonetheless, there still is a lack of the mechanistic understanding of the cellular uptake, subcellular trafficking, and gene knockdown with pacDNA. Here, we show that the pacDNA enters human non-small cell lung cancer cells (NCI-H358) predominantly by scavenger receptor-mediated endocytosis and macropinocytosis and trafficks via the endolysosomal pathway within the cell. The pacDNA significantly reduces a target gene expression (KRAS) in the protein level but not in the mRNA level, despite that the transfection of certain free ASOs causes ribonuclease H1 (RNase H)-dependent degradation of KRAS mRNA. In addition, the antisense activity of pacDNA is independent of ASO chemical modification, suggesting that the pacDNA always functions as a steric blocker.
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spelling pubmed-99062842023-02-14 A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides Zhang, Lei Wang, Yuyan Chen, Peiru Wang, Dali Sun, Tingyu Zhang, Zheyu Wang, Ruimeng Kang, Xi Fang, Yang Lu, Hao Cai, Jiansong Ren, Mengqi Dong, Sijia S. Zhang, Ke RSC Chem Biol Chemistry We have developed a non-cationic transfection vector in the form of bottlebrush polymer-antisense oligonucleotide (ASO) conjugates. Termed pacDNA (polymer-assisted compaction of DNA), these agents show improved biopharmaceutical characteristics and antisense potency in vivo while suppressing non-antisense side effects. Nonetheless, there still is a lack of the mechanistic understanding of the cellular uptake, subcellular trafficking, and gene knockdown with pacDNA. Here, we show that the pacDNA enters human non-small cell lung cancer cells (NCI-H358) predominantly by scavenger receptor-mediated endocytosis and macropinocytosis and trafficks via the endolysosomal pathway within the cell. The pacDNA significantly reduces a target gene expression (KRAS) in the protein level but not in the mRNA level, despite that the transfection of certain free ASOs causes ribonuclease H1 (RNase H)-dependent degradation of KRAS mRNA. In addition, the antisense activity of pacDNA is independent of ASO chemical modification, suggesting that the pacDNA always functions as a steric blocker. RSC 2022-11-08 /pmc/articles/PMC9906284/ /pubmed/36794022 http://dx.doi.org/10.1039/d2cb00149g Text en This journal is © The Royal Society of Chemistry https://creativecommons.org/licenses/by/3.0/
spellingShingle Chemistry
Zhang, Lei
Wang, Yuyan
Chen, Peiru
Wang, Dali
Sun, Tingyu
Zhang, Zheyu
Wang, Ruimeng
Kang, Xi
Fang, Yang
Lu, Hao
Cai, Jiansong
Ren, Mengqi
Dong, Sijia S.
Zhang, Ke
A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title_full A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title_fullStr A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title_full_unstemmed A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title_short A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
title_sort mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9906284/
https://www.ncbi.nlm.nih.gov/pubmed/36794022
http://dx.doi.org/10.1039/d2cb00149g
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