Palmitic acid conjugation enhances potency of tricyclo-DNA splice switching oligonucleotides.

Tricyclo-DNA (tcDNA) is a conformationally constrained oligonucleotide analog that has demonstrated great therapeutic potential as antisense oligonucleotide (ASO) for several diseases. Like most ASOs in clinical development, tcDNA were modified with phosphorothioate (PS) backbone for therapeutic purposes in order to improve their biodistribution by enhancing association with plasma and cell protein. Despite the advantageous protein binding properties, systemic delivery of PS-ASO remains limited and PS modifications can result in dose limiting toxicities in the clinic.

Identifying and Avoiding tcDNA-ASO Sequence-Specific Toxicity for the Development of DMD Exon 51 Skipping Therapy.

Tricyclo-DNA (tcDNA) antisense oligonucleotides (ASOs) hold promise for therapeutic splice-switching applications and the treatment of Duchenne muscular dystrophy (DMD) in particular. We have previously reported the therapeutic potential of tcDNA-ASO in mouse models of DMD, highlighting their unique pharmaceutical properties and unprecedented uptake in many tissues after systemic delivery, including the heart and central nervous system. Following these encouraging results, we developed phosphorothioate (PS)-modified tcDNA-ASOs targeting the human dystrophin exon 51 (H51).

Overcoming Endocytosis Deficiency by Cubosome Nanocarriers.

The use of lipid-based nanoparticles for the delivery of biomacromolecules has attracted considerable attention due to the current interest in protein-based therapeutics. Cubosomes protect the incorporated therapeutics, which are susceptible to degradation by enzymes, thereby improving their bioavailability, and concomitantly enhance cellular uptake. The cubosome nanoparticles presented herein were loaded with bovine serum albumin (BSA) and characterized by small-angle X-ray scattering and dynamic light scattering techniques, while the BSA encapsulation and its release were evaluated .

Evaluating the Impact of Variable Phosphorothioate Content in Tricyclo-DNA Antisense Oligonucleotides in a Duchenne Muscular Dystrophy Mouse Model.

Antisense oligonucleotides (ASOs) hold promise for therapeutic splice switching correction for genetic diseases, in particular for Duchenne muscular dystrophy (DMD), for which ASO-exon skipping represents one of the most advanced therapeutic strategies. We have previously reported the therapeutic potential of tricyclo-DNA (tcDNA) in mouse models of DMD, highlighting the unique pharmaceutical properties and unprecedented uptake in many tissues after systemic delivery, including the heart and central nervous system. TcDNA-ASOs demonstrate an encouraging safety profile and no particular class-related toxicity, however, when administered in high doses for several months, mild renal toxicity is observed secondary to predictable phosphorothioate (PS)-ASO accumulation in kidneys.

Tailored host-guest lipidic cubic phases: a protocell model exhibiting nucleic acid recognition.

A classical conundrum in origin-of-life studies relates to the nature of the first chemical system: was it a carrier of genetic information or a facilitator of cellular compartmentalization? Here we present a system composed of tailor-made nucleolipids and hydrated monoolein, which assemble at ambient temperatures to form host-guest lipidic cubic phase (LCP) materials that are stable in bulk water and can perform both functions. As such, they may represent a molecular model for a protocell in origin-of-life studies. Nucleolipids within the lipidic material sequester and bind selectively complementary oligonucleotide sequences from solution by virtue of base-pairing; noncomplementary sequences diffuse freely between the LCP material and the bulk aqueous environment.

Role of damage-specific DNA polymerases in M13 phage mutagenesis induced by a major lipid peroxidation product trans-4-hydroxy-2-nonenal.

One of the major lipid peroxidation products trans-4-hydroxy-2-nonenal (HNE), forms cyclic propano- or ethenoadducts bearing six- or seven-carbon atom side chains to G>C≫A>T. To specify the role of SOS DNA polymerases in HNE-induced mutations, we tested survival and mutation spectra in the lacZα gene of M13mp18 phage, whose DNA was treated in vitro with HNE, and which was grown in uvrA(-)Escherichia coli strains, carrying one, two or all three SOS DNA polymerases. When Pol IV was the only DNA SOS polymerase in the bacterial host, survival of HNE-treated M13 DNA was similar to, but mutation frequency was lower than in the strain containing all SOS DNA polymerases.

Nucleotide excision repair and recombination are engaged in repair of trans-4-hydroxy-2-nonenal adducts to DNA bases in Escherichia coli.

One of the major products of lipid peroxidation is trans-4-hydroxy-2-nonenal (HNE). HNE forms highly mutagenic and genotoxic adducts to all DNA bases. Using M13 phage lacZ system, we studied the mutagenesis and repair of HNE treated phage DNA in E.

Cockayne syndrome group B protein is engaged in processing of DNA adducts of lipid peroxidation product trans-4-hydroxy-2-nonenal.

Cockayne syndrome complementation group B (CSB) protein is engaged in transcription-coupled repair (TCR) of UV induced DNA damage and its deficiency leads to progressive multisystem degeneration and premature aging. Here, we show that human CSB-deficient cells are hypersensitive to physiological concentrations (1-10 microM) of a lipid peroxidation product, trans-4-hydroxy-2-nonenal (HNE), and in response to HNE they develop a higher level of sister chromatid exchanges (SCEs) in comparison to the wild-type cells. HNE-DNA adducts block in vitro transcription by T7 RNA polymerase, as well as by HeLa cell-free extracts.

Practical highly enantioselective synthesis of (R)- and (S)-(E)-4-hydroxynon-2-enal.

Oxidative stress enhances lipid peroxidation (LPO) implicated in cancer promotion and progression. (E)-4-Hydroxynon-2-enal 1 (trans-4-hydroxy-2-nonenal, HNE) is one of the most abundant products of LPO. Reactions of HNE with DNA and proteins are responsible for its mutagenic and toxic effects.

Long-chain adducts of trans-4-hydroxy-2-nonenal to DNA bases cause recombination, base substitutions and frameshift mutations in M13 phage.

Oxidative stress enhances lipid peroxidation (LPO) implicated in the promotion and progression of carcinogenesis. One of the major LPO products is trans-4-hydroxy-2-nonenal (HNE), which was shown to react with guanosine and under peroxidizing conditions also with adenosine. We show here that all four DNA bases are targets for HNE, although displaying different reactivity: dG > dC > dA approximately equal to dT.