Cellular delivery of polyheteroaromate-peptide nucleic acid conjugates mediated by cationic lipids

Takehiko Shiraishi; Nadia Bendifallah; Peter E. Nielsen

doi:10.1021/bc050246z

Cellular delivery of polyheteroaromate-peptide nucleic acid conjugates mediated by cationic lipids

Takehiko Shiraishi, Nadia Bendifallah, Peter E. Nielsen

Transcription, RNA, and Gene Medicine Program

Abstract

In the search of facile and efficient methods for PNA cellular delivery, we have tested a series of PNA conjugates based on (hetero) aromatic, lipophilic compounds such as 9-aminoacridine, benzimidazoles, carbazole, anthraquinone, porphyrine, psoralen, pyrene, and phenyl-bis-benzimidazole ("Hoechst"). These chemically modified PNAs were delivered to cultured pLuc705HeLa cells mediated by cationic liposomes (LipofectAMINE or LiofectAMINE2000), and their nuclear delivery was inferred from induced luciferase activity as a consequence of pre-mRNA splicing correction by the antisense-PNA. PNAs modified with 9-aminoacridine, "Hoechst", or acetyl-"Hoechst" showed highest antisense activities (while unmodified PNA failed to show any significant antisense activity). In particular, bis-acridine-conjugated PNA showed nearly 60% splicing correction at 250 nM concentration in combination with LipofectAMINE2000. Interestingly, relative differences between the derivatives were observed when LipofectAMINE was used as compared to LipofectAMINE2000, but in general the latter yielded the higher antisense activity. The most active modifications of these PNA constructs were further tested for antisense down-regulation of luciferase in p53R cells in order to evaluate the cytoplasmic activity (uptake) of the PNAs. A dose-dependent down regulation of luciferase was demonstrated also in this system. The PNA conjugated to acetyl-Hoechst caused a reduction of luciferase activity to less than 40% of the control at a concentration of 1 muM. These results indicate that conjugation of (hetero) polyaromatic compounds to PNA can dramatically improve liposome-mediated cellular delivery both to cytoplasm as well as to the nucleus. However, no clear structure/activity relations are apparent from the present results, except that both 9-aminoacridine and "Hoechst" are also nucleic acid binding ligands.

Originalsprog	Engelsk
Tidsskrift	Bioconjugate Chemistry
Vol/bind	17
Udgave nummer	1
Sider (fra-til)	189-94
Antal sider	6
ISSN	1043-1802
DOI	https://doi.org/10.1021/bc050246z
Status	Udgivet - 19 jan. 2006

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10.1021/bc050246z

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abstract = "In the search of facile and efficient methods for PNA cellular delivery, we have tested a series of PNA conjugates based on (hetero) aromatic, lipophilic compounds such as 9-aminoacridine, benzimidazoles, carbazole, anthraquinone, porphyrine, psoralen, pyrene, and phenyl-bis-benzimidazole ({"}Hoechst{"}). These chemically modified PNAs were delivered to cultured pLuc705HeLa cells mediated by cationic liposomes (LipofectAMINE or LiofectAMINE2000), and their nuclear delivery was inferred from induced luciferase activity as a consequence of pre-mRNA splicing correction by the antisense-PNA. PNAs modified with 9-aminoacridine, {"}Hoechst{"}, or acetyl-{"}Hoechst{"} showed highest antisense activities (while unmodified PNA failed to show any significant antisense activity). In particular, bis-acridine-conjugated PNA showed nearly 60% splicing correction at 250 nM concentration in combination with LipofectAMINE2000. Interestingly, relative differences between the derivatives were observed when LipofectAMINE was used as compared to LipofectAMINE2000, but in general the latter yielded the higher antisense activity. The most active modifications of these PNA constructs were further tested for antisense down-regulation of luciferase in p53R cells in order to evaluate the cytoplasmic activity (uptake) of the PNAs. A dose-dependent down regulation of luciferase was demonstrated also in this system. The PNA conjugated to acetyl-Hoechst caused a reduction of luciferase activity to less than 40% of the control at a concentration of 1 muM. These results indicate that conjugation of (hetero) polyaromatic compounds to PNA can dramatically improve liposome-mediated cellular delivery both to cytoplasm as well as to the nucleus. However, no clear structure/activity relations are apparent from the present results, except that both 9-aminoacridine and {"}Hoechst{"} are also nucleic acid binding ligands.",

keywords = "Antisense Elements (Genetics), DNA/administration & dosage, Drug Carriers, HeLa Cells, Humans, Lipids/administration & dosage, Liposomes, Peptide Nucleic Acids/administration & dosage, Transfection",

author = "Takehiko Shiraishi and Nadia Bendifallah and Nielsen, {Peter E.}",

year = "2006",

month = jan,

day = "19",

doi = "10.1021/bc050246z",

language = "English",

volume = "17",

pages = "189--94",

journal = "Bioconjugate Chemistry",

issn = "1043-1802",

publisher = "American Chemical Society",

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TY - JOUR

T1 - Cellular delivery of polyheteroaromate-peptide nucleic acid conjugates mediated by cationic lipids

AU - Shiraishi, Takehiko

AU - Bendifallah, Nadia

AU - Nielsen, Peter E.

PY - 2006/1/19

Y1 - 2006/1/19

N2 - In the search of facile and efficient methods for PNA cellular delivery, we have tested a series of PNA conjugates based on (hetero) aromatic, lipophilic compounds such as 9-aminoacridine, benzimidazoles, carbazole, anthraquinone, porphyrine, psoralen, pyrene, and phenyl-bis-benzimidazole ("Hoechst"). These chemically modified PNAs were delivered to cultured pLuc705HeLa cells mediated by cationic liposomes (LipofectAMINE or LiofectAMINE2000), and their nuclear delivery was inferred from induced luciferase activity as a consequence of pre-mRNA splicing correction by the antisense-PNA. PNAs modified with 9-aminoacridine, "Hoechst", or acetyl-"Hoechst" showed highest antisense activities (while unmodified PNA failed to show any significant antisense activity). In particular, bis-acridine-conjugated PNA showed nearly 60% splicing correction at 250 nM concentration in combination with LipofectAMINE2000. Interestingly, relative differences between the derivatives were observed when LipofectAMINE was used as compared to LipofectAMINE2000, but in general the latter yielded the higher antisense activity. The most active modifications of these PNA constructs were further tested for antisense down-regulation of luciferase in p53R cells in order to evaluate the cytoplasmic activity (uptake) of the PNAs. A dose-dependent down regulation of luciferase was demonstrated also in this system. The PNA conjugated to acetyl-Hoechst caused a reduction of luciferase activity to less than 40% of the control at a concentration of 1 muM. These results indicate that conjugation of (hetero) polyaromatic compounds to PNA can dramatically improve liposome-mediated cellular delivery both to cytoplasm as well as to the nucleus. However, no clear structure/activity relations are apparent from the present results, except that both 9-aminoacridine and "Hoechst" are also nucleic acid binding ligands.

AB - In the search of facile and efficient methods for PNA cellular delivery, we have tested a series of PNA conjugates based on (hetero) aromatic, lipophilic compounds such as 9-aminoacridine, benzimidazoles, carbazole, anthraquinone, porphyrine, psoralen, pyrene, and phenyl-bis-benzimidazole ("Hoechst"). These chemically modified PNAs were delivered to cultured pLuc705HeLa cells mediated by cationic liposomes (LipofectAMINE or LiofectAMINE2000), and their nuclear delivery was inferred from induced luciferase activity as a consequence of pre-mRNA splicing correction by the antisense-PNA. PNAs modified with 9-aminoacridine, "Hoechst", or acetyl-"Hoechst" showed highest antisense activities (while unmodified PNA failed to show any significant antisense activity). In particular, bis-acridine-conjugated PNA showed nearly 60% splicing correction at 250 nM concentration in combination with LipofectAMINE2000. Interestingly, relative differences between the derivatives were observed when LipofectAMINE was used as compared to LipofectAMINE2000, but in general the latter yielded the higher antisense activity. The most active modifications of these PNA constructs were further tested for antisense down-regulation of luciferase in p53R cells in order to evaluate the cytoplasmic activity (uptake) of the PNAs. A dose-dependent down regulation of luciferase was demonstrated also in this system. The PNA conjugated to acetyl-Hoechst caused a reduction of luciferase activity to less than 40% of the control at a concentration of 1 muM. These results indicate that conjugation of (hetero) polyaromatic compounds to PNA can dramatically improve liposome-mediated cellular delivery both to cytoplasm as well as to the nucleus. However, no clear structure/activity relations are apparent from the present results, except that both 9-aminoacridine and "Hoechst" are also nucleic acid binding ligands.

KW - Antisense Elements (Genetics)

KW - DNA/administration & dosage

KW - Drug Carriers

KW - HeLa Cells

KW - Humans

KW - Lipids/administration & dosage

KW - Liposomes

KW - Peptide Nucleic Acids/administration & dosage

KW - Transfection

U2 - 10.1021/bc050246z

DO - 10.1021/bc050246z

M3 - Journal article

C2 - 16417268

SN - 1043-1802

VL - 17

SP - 189

EP - 194

JO - Bioconjugate Chemistry

JF - Bioconjugate Chemistry

IS - 1

ER -

Cellular delivery of polyheteroaromate-peptide nucleic acid conjugates mediated by cationic lipids

Abstract

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