An Efficient Biodelivery System for Antisense Polyamide Nucleic Acid (PNA)

Abstract

With the aim of developing a general and straightforward procedure for the intracellular delivery of naked peptide nucleic acids (PNAs), we designed an intracellularly biodegradable triphenylphosphonium (TPP) cation based transporter system. In this system, TPP is linked, via a biolabile disul de bridge, to an activated mercaptoethoxycarbonyl moiety, allowing its direct coupling to the N-terminal extremity of a free PNA through a carbamate bond. We found that such TPP-PNA-carbamate conjugates were highly stable in a cell culture medium containing fetal calf serum. In a glutathione-containing medium mimicking the cytosol, the conjugates were rapidly degraded into an unstable intermediate, which spontaneously decomposed, releasing the free PNA. Using a uorescence-labeled PNA-TPP conjugate, we demonstrated that conjugates were taken up by cells. Ef cient cellular uptake and release of the PNA into the cytosol was further con rmed by the anti-HIV activity measured for the TPP-conjugate of a 16-mer PNA targeting the TAR region of the HIV-1 genome. This conjugate exhibited an IC 50 value of 1 µM, while the free 16-mer PNA did not inhibit replication of HIV in the same cellular test.

miRNA Therapeutics: Delivery and Biological Activity of Peptide Nucleic Acids Targeting miRNAs

“…Therefore, a major intracellular barrier for therapeutic applications of macromolecules having intracellular targets of action is their degradation in endocytic vesicles after uptake by endocytosis. Mehiri et al [97] described an interesting method for the development of a general and efficient procedure for the intracellular delivery of bioavailable naked PNAs. An intracellularly biodegradable triphenylphosphonium cation based transporter system was designed and linked, via a biolabile disulfide bridge, to an activated mercaptoethoxycarbonyl moiety, allowing its direct coupling to the N-terminal extremity of a free PNA through a carbamate bond [97].…”

“…Mehiri et al [97] described an interesting method for the development of a general and efficient procedure for the intracellular delivery of bioavailable naked PNAs. An intracellularly biodegradable triphenylphosphonium cation based transporter system was designed and linked, via a biolabile disulfide bridge, to an activated mercaptoethoxycarbonyl moiety, allowing its direct coupling to the N-terminal extremity of a free PNA through a carbamate bond [97]. PNA-triphenylphosphonium conjugates are taken up by cells and released into the cytosol following rapid degradation into an unstable intermediate, which spontaneously decomposed, releasing the free PNA [97].…”

“…An intracellularly biodegradable triphenylphosphonium cation based transporter system was designed and linked, via a biolabile disulfide bridge, to an activated mercaptoethoxycarbonyl moiety, allowing its direct coupling to the N-terminal extremity of a free PNA through a carbamate bond [97]. PNA-triphenylphosphonium conjugates are taken up by cells and released into the cytosol following rapid degradation into an unstable intermediate, which spontaneously decomposed, releasing the free PNA [97]. Another set of studies and reviews presented a second approach, photochemical internalization, as a novel technology for the release of endocytosed macromolecules into the cytosol [98].…”

Immunological Response to Peptide Nucleic Acid and its Peptide Conjugate Targeted to Transactivation Response (TAR) Region of HIV-1 RNA Genome

“…Several approaches have been used to improve the biodelivery and effi cacy of PNAs including their covalent conjugation to carriers such as neutral or cationic lipophilic molecules (Muratovska et al, 2001;Filipovska et al, 2004;Mehiri et al, 2008), cell-penetrating peptides (CPPs) (Kaushik et al, 2002a), and cell-specifi c receptor ligands or encapsulation in autologous erythrocytes (Boffa et al, 2000;Koppelhus and Nielsen, 2003;Chiarantini et al, 2005). We have shown that anti-HIV-1 PNA conjugated with neamine is not only effi ciently taken up by the cells but also acquires a unique nuclease-like property that specifi cally cleaves the target RNA (Riguet et al, 2004;Chaubey et al, 2007).…”

Prospects for antisense peptide nucleic acid (PNA) therapies for HIV

“…A number of reviews on CPP and their therapeutic application have recently been published [49-52] . A triphenyl-phosphonium-tagged PNA targeted to HIV-1 TAR was found to be efficiently taken up by cells and to block HIV-1 replication in cell culture [53]. In one unique study, PNA complementary to the HIV-1 TAR region was conjugated to neamine, a polycationic aminoglycoside.…”