Advances in Nucleic Acid Research: Exploring the Potential of Oligonucleotides for Therapeutic Applications and Biological Studies.

In recent years, nucleic acids have emerged as powerful biomaterials, revolutionizing the field of biomedicine. This review explores the multifaceted applications of nucleic acids, focusing on their pivotal role in various biomedical applications. Nucleic acids, including deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), possess unique properties such as molecular recognition ability, programmability, and ease of synthesis, making them versatile tools in biosensing and for gene regulation, drug delivery, and targeted therapy. Their compatibility with chemical modifications enhances their binding affinity and resistance to degradation, elevating their effectiveness in targeted applications. Additionally, nucleic acids have found utility as self-assembling building blocks, leading to the creation of nanostructures whose high order underpins their enhanced biological stability and affects the cellular uptake efficiency. Furthermore, this review delves into the significant role of oligonucleotides (ODNs) as indispensable tools for biological studies and biomarker discovery. ODNs, short sequences of nucleic acids, have been instrumental in unraveling complex biological mechanisms. They serve as probes for studying gene expression, protein interactions, and cellular pathways, providing invaluable insights into fundamental biological processes. By examining the synergistic interplay between nucleic acids as powerful biomaterials and ODNs as indispensable tools for biological studies and biomarkers, this review highlights the transformative impact of these molecules on biomedical research. Their versatile applications not only deepen our understanding of biological systems but also are the driving force for innovation in diagnostics and therapeutics, ultimately advancing the field of biomedicine.

Chlamydomonas reinhardtii: A Factory of Nutraceutical and Food Supplements for Human Health.

Chlamydomonas reinhardtii (C. reinhardtii) is one of the most well-studied microalgae organisms that revealed important information for the photosynthetic and metabolic processes of plants and eukaryotes. Numerous extensive studies have also underpinned its great potential as a biochemical factory, capable of producing various highly desired molecules with a direct impact on human health and longevity. Polysaccharides, lipids, functional proteins, pigments, hormones, vaccines, and antibodies are among the valuable biomolecules that are produced spontaneously or under well-defined conditions by C. reinhardtii and can be directly linked to human nutrition and diet. The aim of this review is to highlight the recent advances in the field focusing on the most relevant applications related to the production of important biomolecules for human health that are also linked with human nutrition and diet. The limitations and challenges are critically discussed along with the potential future applications of C. reinhardtii biomass and processed products in the field of nutraceuticals and food supplements. The increasing need for high-value and low-cost biomolecules produced in an environmentally and economy sustainable manner also underline the important role of C. reinhardtii.

Supported Gold Nanoparticle-Catalyzed Selective Reduction of Multifunctional, Aromatic Nitro Precursors into Amines and Synthesis of 3,4-Dihydroquinoxalin-2-Ones.

The synthesis of 3,4-dihydroquinoxalin-2-ones via the selective reduction of aromatic, multifunctional nitro precursors catalyzed by supported gold nanoparticles is reported. The reaction proceeds through the in situ formation of the corresponding amines under heterogeneous transfer hydrogenation of the initial nitro compounds catalyzed by the commercially available Au/TiO2-Et3SiH catalytic system, followed by an intramolecular C-N transamidation upon treatment with silica acting as a mild acid. Under the present conditions, the Au/TiO2-TMDS system was also found to catalyze efficiently the present selective reduction process. Both transfer hydrogenation processes showed very good functional-group tolerance and were successfully applied to access more structurally demanding products bearing other reducible moieties such as chloro, aldehyde or methyl ketone. An easily scalable (up to 1 mmol), low catalyst loading (0.6 mol%) synthetic protocol was realized, providing access to this important scaffold. Under these mild catalytic conditions, the desired products were isolated in good to high yields and with a TON of 130. A library analysis was also performed to demonstrate the usefulness of our synthetic strategy and the physicochemical profile of the derivatives.

Selective C-H Allylic Oxygenation of Cycloalkenes and Terpenoids Photosensitized by [Cu(Xantphos)(neoc)]BF4.

We present herein for the first time the use of the [Cu(Xantphos)(neoc)]BF4 as a photocatalyst for the selective C-H allylic oxygenation of cycloalkenes into the corresponding allylic hydroperoxides or alcohols in the presence of molecular oxygen. The proposed methodology affords the products at good yields and has also been applied successfully to several bioactive terpenoids, such as geraniol, linalool, ß-citronellol, and phytol. A mechanistic study involving also kinetic isotope effects (KIEs) supports the proposed singlet oxygen-mediated reaction. On the basis of the high chemoselectivity and yields and the fast and clean reaction processes observed, the present catalytic system, [Cu(Xantphos)(neoc)]BF4, has also been applied to the synthesis, at a laboratory scale, of the cis-Rose oxide, a well-known perfumery ingredient used in rose and geranium perfumes.

Selective Reduction of Nitroarenes to Arylamines by the Cooperative Action of Methylhydrazine and a Tris(N-heterocyclic thioamidate) Cobalt(III) Complex.

We report an efficient catalytic protocol that chemoselectively reduces nitroarenes to arylamines, by using methylhydrazine as a reducing agent in combination with the easily synthesized and robust catalyst tris(N-heterocyclic thioamidate) Co(III) complex [Co(κS,N-tfmp2S)3], tfmp2S = 4-(trifluoromethyl)-pyrimidine-2-thiolate. A series of arylamines and heterocyclic amines were formed in excellent yields and chemoselectivity. High conversion yields of nitroarenes into the corresponding amines were observed by using polar protic solvents, such as MeOH and iPrOH. Among several hydrogen donors that were examined, methylhydrazine demonstrated the best performance. Preliminary mechanistic investigations, supported by UV-vis and NMR spectroscopy, cyclic voltammetry, and high-resolution mass spectrometry, suggest a cooperative action of methylhydrazine and [Co(κS,N-tfmp2S)3] via a coordination activation pathway that leads to the formation of a reduced cobalt species, responsible for the catalytic transformation. In general, the corresponding N-arylhydroxylamines were identified as the sole intermediates. Nevertheless, the corresponding nitrosoarenes can also be formed as intermediates, which, however, are rapidly transformed into the desired arylamines in the presence of methylhydrazine through a noncatalytic path. On the basis of the observed high chemoselectivity and yields, and the fast and clean reaction processes, the present catalytic system [Co(κS,N-tfmp2S)3]/MeNHNH2 shows promise for the efficient synthesis of aromatic amines that could find various industrial applications.

Direct and Indirect Chemiluminescence: Reactions, Mechanisms and Challenges.

Emission of light by matter can occur through a variety of mechanisms. When it results from an electronically excited state of a species produced by a chemical reaction, it is called chemiluminescence (CL). The phenomenon can take place both in natural and artificial chemical systems and it has been utilized in a variety of applications. In this review, we aim to revisit some of the latest CL applications based on direct and indirect production modes. The characteristics of the chemical reactions and the underpinning CL mechanisms are thoroughly discussed in view of studies from the very recent bibliography. Different methodologies aiming at higher CL efficiencies are summarized and presented in detail, including CL type and scaffolds used in each study. The CL role in the development of efficient therapeutic platforms is also discussed in relation to the Reactive Oxygen Species (ROS) and singlet oxygen (1O2) produced, as final products. Moreover, recent research results from our team are included regarding the behavior of commonly used photosensitizers upon chemical activation under CL conditions. The CL prospects in imaging, biomimetic organic and radical chemistry, and therapeutics are critically presented in respect to the persisting challenges and limitations of the existing strategies to date.

Nucleotide Excision Repair and Impact of Site-Specific 5',8-Cyclopurine and Bulky DNA Lesions on the Physical Properties of Nucleosomes.

The nonbulky 5',8-cyclopurine DNA lesions (cP) and the bulky, benzo[ a]pyrene diol epoxide-derived stereoisomeric cis- and trans- N2-guanine adducts (BPDE-dG) are good substrates of the human nucleotide excision repair (NER) mechanism. These DNA lesions were embedded at the In or Out rotational settings near the dyad axis in nucleosome core particles reconstituted either with native histones extracted from HeLa cells (HeLa-NCP) or with recombinant histones (Rec-NCP). The cP lesions are completely resistant to NER in human HeLa cell extracts. The BPDE-dG adducts are also NER-resistant in Rec-NCPs but are good substrates of NER in HeLa-NCPs. The four BPDE-dG adduct samples are excised with different efficiencies in free DNA, but in HeLa-NCPs, the efficiencies are reduced by a common factor of 2.2 ± 0.2 relative to the NER efficiencies in free DNA. The NER response of the BPDE-dG adducts in HeLa-NCPs is not directly correlated with the observed differences in the thermodynamic destabilization of HeLa-NCPs, the Förster resonance energy transfer values, or hydroxyl radical footprint patterns and is weakly dependent on the rotational settings. These and other observations suggest that NER is initiated by the binding of the DNA damage-sensing NER factor XPC-RAD23B to a transiently opened BPDE-modified DNA sequence that corresponds to the known footprint of XPC-DNA-RAD23B complexes (≥30 bp). These observations are consistent with the hypothesis that post-translational modifications and the dimensions and properties of the DNA lesions are the major factors that have an impact on the dynamics and initiation of NER in nucleosomes.

New Insights into the Reaction Paths of Hydroxyl Radicals with Purine Moieties in DNA and Double-Stranded Oligodeoxynucleotides.

The reaction of hydroxyl radical (HO•) with DNA produces many primary reactive species and many lesions as final products. In this study, we have examined the optical spectra of intermediate species derived from the reaction of HO• with a variety of single- and double-stranded oligodeoxynucleotides and ct-DNA in the range of 1 µs to 1 ms by pulse radiolysis using an Intensified Charged Coupled Device (ICCD) camera. Moreover, we applied our published analytical protocol based on an LC-MS/MS system with isotopomeric internal standards to enable accurate and precise measurements of purine lesion formation. In particular, the simultaneous measurement of the four purine 5',8-cyclo-2'-deoxynucleosides (cPu) and two 8-oxo-7,8-dihydro-2'-deoxypurine (8-oxo-Pu) was obtained upon reaction of genetic material with HO• radicals generated either by γ-radiolysis or Fenton-type reactions. Our results contributed to the debate in the literature regarding absolute level of lesions, method of HO• radical generation, 5'R/5'S diastereomeric ratio in cPu, and relative abundance between cPu and 8-oxo-Pu.

A 5', 8-cyclo-2'-deoxypurine lesion induces trinucleotide repeat deletion via a unique lesion bypass by DNA polymerase ß.

5',8-cyclo-2'-deoxypurines (cdPus) are common forms of oxidized DNA lesions resulting from endogenous and environmental oxidative stress such as ionizing radiation. The lesions can only be repaired by nucleotide excision repair with a low efficiency. This results in their accumulation in the genome that leads to stalling of the replication DNA polymerases and poor lesion bypass by translesion DNA polymerases. Trinucleotide repeats (TNRs) consist of tandem repeats of Gs and As and therefore are hotspots of cdPus. In this study, we provided the first evidence that both (5'R)- and (5'S)-5',8-cyclo-2'-deoxyadenosine (cdA) in a CAG repeat tract caused CTG repeat deletion exclusively during DNA lagging strand maturation and base excision repair. We found that a cdA induced the formation of a CAG loop in the template strand, which was skipped over by DNA polymerase ß (pol ß) lesion bypass synthesis. This subsequently resulted in the formation of a long flap that was efficiently cleaved by flap endonuclease 1, thereby leading to repeat deletion. Our study indicates that accumulation of cdPus in the human genome can lead to TNR instability via a unique lesion bypass by pol ß.

Structural basis for the recognition of diastereomeric 5',8-cyclo-2'-deoxypurine lesions by the human nucleotide excision repair system.

The hydroxyl radical is a powerful oxidant that generates DNA lesions including the stereoisomeric R and S 5',8-cyclo-2'-deoxyadenosine (cdA) and 5',8-cyclo-2'-deoxyguanosine (cdG) pairs that have been detected in cellular DNA. Unlike some other oxidatively generated DNA lesions, cdG and cdA are repaired by the human nucleotide excision repair (NER) apparatus. The relative NER efficiencies of all four cyclopurines were measured and compared in identical human HeLa cell extracts for the first time under identical conditions, using identical sequence contexts. The cdA and cdG lesions were excised with similar efficiencies, but the efficiencies for both 5'R cyclopurines were greater by a factor of ∼2 than for the 5'S lesions. Molecular modeling and dynamics simulations have revealed structural and energetic origins of this difference in NER-incision efficiencies. These lesions cause greater DNA backbone distortions and dynamics relative to unmodified DNA in 5'R than in 5'S stereoisomers, producing greater impairment in van der Waals stacking interaction energies in the 5'R cases. The locally impaired stacking interaction energies correlate with relative NER incision efficiencies, and explain these results on a structural basis in terms of differences in dynamic perturbations of the DNA backbone imposed by the R and S covalent 5',8 bonds.

Differences in the Access of Lesions to the Nucleotide Excision Repair Machinery in Nucleosomes.

In nucleosomes, the access of DNA lesions to nucleotide excision repair is hindered by histone proteins. However, evidence that the nature of the DNA lesions may play a role in facilitating access is emerging, but these phenomena are not well-understood. We have used molecular dynamics simulations to elucidate the structural, dynamic, and energetic properties of the R and S 5'-8-cyclo-2'-dG and the (+)-cis-anti-B[a]P-dG lesions in a nucleosome. Our results show that the (+)-cis-anti-B[a]P-dG adduct is more dynamic and more destabilizing than the smaller and more constrained 5',8-cyclo-2'-dG lesions, suggesting more facile access to the more bulky (+)-cis-anti-B[a]P-dG lesion.

Protocol for the simultaneous quantification of oxidative purine lesions in DNA using LC-MS/MS analysis.

Most DNA damages induced through oxidative metabolism are single lesions which can accumulate in tissues. Here, we present a protocol for the simultaneous quantification of oxidative purine lesions (cPu and 8-oxo-Pu) in DNA. We describe steps for enzymatic digestion of DNA and sample pre-purification, followed by quantification through liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. We optimized this protocol in commercially available calf thymus DNA and used genomic and mitochondrial DNA extracted from cell cultures and animal and human tissues.

Synergy of redox-activity and hemilability in thioamidato cobalt(III) complexes for the chemoselective reduction of nitroarenes to anilines: catalytic and mechanistic investigation.

Reduction of nitro-compounds to amines is one of the most often employed and challenging catalytic processes in the fine and bulk chemical industry. Herein, we present two series of mononuclear homoleptic and heteroleptic Co(III) complexes, i.e., [Co(LNS)3] and [Co(LNS)2L1L2]x+, respectively (x = 0 or 1, LNS = pyrimidine- or pyridine-thioamidato, L1/L2 = thioamidato, phosphine or pyridine), which successfully catalyze the transformation of nitroarenes to anilines by methylhydrazine. The catalytic reaction can be accomplished for a range of electronically and sterically diverse nitroarenes, using mild experimental conditions and low catalyst loadings, resulting in the corresponding anilines in high yields, with high chemoselectivity, and no side-products. Electronic and steric properties of the ligands play pivotal role in the catalytic efficacy of the respective complexes. In particular, complexes bearing ligands of high hemilability/lability and being capable of stabilizing lower metal oxidation-states exhibit the highest catalytic activity. Mechanistic investigations suggest the participation of the Co(III) complexes in two parallel reaction pathways: (a) coordination-induced activation of methylhydrazine and (b) reduction of nitroarenes to anilines by methylhydrazine, through the formation of Co(I) and Co-hydride intermediates.

One-pot five-component synthesis of spirocyclopenta[b]chromene derivatives and their acid-catalyzed rearrangement.

The reaction of the zwitterionic intermediate, generated in situ from either tert-butylisocyanide or cyclohexylisocyanide and acetylenedicarboxylates, with 3-cyanochromones is described, whereupon spirochromenofuran derivatives 5 or 6 were obtained in good yields. The subsequent acid-catalyzed rearrangement of the isolated 2-imino-spirochromenofurans 5 to 2-amino-spirochromenofurans 7 has also been studied. Rational mechanistic schemes for the formation of compounds 5, 6, and 7 are proposed. The structure elucidation of the products was accomplished by 1D and 2D NMR experiments and confirmed by X-ray crystallographic analysis. Full assignment of all (1)H and (13)C NMR chemical shifts has been unambiguously achieved with the aid of DFT/GIAO calculations.

Purine 5',8-cyclonucleoside lesions: chemistry and biology.

5',8-Cyclo-2'-deoxyadenosine and 5',8-cyclo-2'-deoxyguanosine in their 5'R and 5'S diastereomeric forms are tandem-type lesions observed among the DNA modifications and identified in mammalian cellular DNA in vivo. These lesions result from the chemistry of the C5' radicals generated by the attack of HO˙ radicals to 2-deoxyribose units. Quantitative determination of these lesions in biological samples as biomarkers of free radical damage is a challenge. Results reported for irradiated samples of calf thymus DNA have been critically reviewed, underlining the need of further research for the potential involvement of these lesions in human health (76 references).

Biomimetic models of radical stress and related biomarkers.

The biological consequences of free radical production is the central subject of a very lively scientific debate, focusing on the estimation of the type and extent of damage, as well as the efficiency of the protective and repair systems. When studying free radical based chemical mechanisms, it is very important to establish biomimetic models, which allow the experiments to be performed in a simplified environment, but suitably designed to be in strict connection with cellular conditions. The biomimetic modeling approach has been coupled with physical organic chemistry methodologies and knowledge of free radical reactivity. Molecular basis of important processes have been identified, building up molecular libraries of products concerning unsaturated lipids, sulfur-containing proteins and nucleic acids, to be developed as biomarkers. Ongoing projects in our group deal with lipidomics, genomics and proteomics of free radical stress and some examples will be described.

One-pot synthesis of functionalized spirobenzofuranones via MCR involving 3-cyanochromones.

Another aspect concerning chromone chemistry leading to the one-pot synthesis of functionalized novel spirobenzofuranones has been described. The synthesis involves reaction of the zwitterionic intermediates formed by the 1:1 interaction between isocyanides and acetylenecarboxylates with 3-cyanochromones, whereupon through an unexpected and unprecedented reaction of the chromone moiety the isolated benzofuranones are formed. The regioselectivity of the reaction was investigated by DFT calculations. The geometries of the intermediates, transition structures, and intermediate products, leading to the final products, were optimized using the B3LYP functional with the 6-31G(d) basis set. The structures of the products were elucidated by 1D and 2D NMR experiments. Full assignment of all (1)H and (13)C NMR chemical shifts has been achieved. A plausible mechanistic rationale is proposed.

On the relevance of hydroxyl radical to purine DNA damage.

Hydroxyl radical (HO•) is the most reactive toward DNA among the reactive oxygen species (ROS) generated in aerobic organisms by cellular metabolisms. HO• is generated also by exogenous sources such as ionizing radiations. In this review we focus on the purine DNA damage by HO• radicals. In particular, emphasis is given on mechanistic aspects for the various lesion formation and their interconnections. Although the majority of the purine DNA lesions like 8-oxo-purine (8-oxo-Pu) are generated by various ROS (including HO•), the formation of 5',8-cyclopurine (cPu) lesions in vitro and in vivo relies exclusively on the HO• attack. Methodologies generally utilized for the purine lesions quantification in biological samples are reported and critically discussed. Recent results on cPu and 8-oxo-Pu lesions quantification in various types of biological specimens associated with the cellular repair efficiency as well as with distinct pathologies are presented, providing some insights on their biological significance.

Skeletally Tunable Seven-Membered-Ring Fused Pyrroles.

We describe a copper-mediated method that enables the synthesis of seven-membered-ring fused pyrroles (7-mrFPs). The protocol proceeds via an in situ spiro-intermediate ring expansion and tolerates a library of 7-mrFP derivatives with a broad range of functional groups in a simple step with tangible parameters and substrate adaptations. These rare 7-mrFPs are now accessible on a millimolar scale, and selected examples exhibit high antioxidant activity.

One-pot synthesis of chromenylfurandicarboxylates and cyclopenta[b]chromenedicarboxylates involving zwitterionic intermediates. A DFT investigation on the regioselectivity of the reaction.

The reaction of 1:1 zwitterionic intermediates generated in situ from either tert-butylisocyanide or cyclohexylisocyanide and acetylenedicarboxylates with 3-formylchromones is described, whereupon either chromenylfurandicarboxylates or cyclopenta[b]chromenedicarboxylates are formed, depending on the nature of the chromone 6-position substituent and also on the acetylene ester group. In addition, from the reaction with a 1:2 zwitterionic intermediate, cyclohepta[b]chromenetetracarboxylates are isolated. The regioselectivity of the reaction was also investigated by DFT calculations. The geometries of the reactants, intermediate zwitterions, transition structures, and intermediate products, leading to the final products, were optimized using the B3LYP functional with the 6-31G(d) basis set. The structures of the products were elucidated by 1D and 2D NMR experiments. Full assignment of all (1)H and (13)C NMR chemical shifts has been achieved. Plausible mechanistic schemes are provided.