The Conformational Changes of 5SrRNA from Lupin Seeds and tRNAPhe in Presence of Ca2+, Mn2+ Cations by DSC Method

The results of calorimetric studies of 5SrRNA isolated from Lupinus luteus and of tRNA^Phe both in the absence and in the presence of different concentrations of cations Ca²⁺, Mn²⁺ were reported. The temperature and the enthalpy of melting were determined. Using the deconvolution method the elementary transitions were distinguished and discussed. This revised version was published online in August 2006 with corrections to the Cover Date.     

Complexation of well-controlled low-molecular weight polyelectrolytes with antisense oligonucleotides

The influences of polymer-related properties such as molecular weight, charge density, counter ion, and hydrophilic block on the complexation of polyelectrolytes and a fluorescein-labeled oligonucleotide (ON) were investigated. A series of well-defined and well-controlled 2-(N,N-dimethylamino)ethyl methacrylate (DMAEMA) polymers and block copolymers were prepared using living anionic and radical polymerization methods. Fluorescence measurement was used to reveal the effects of polymer molecular weight, charge density, and counter ion type on the complexation. PolyDMAEMA samples having double molecular weights of the chosen oligonucleotide gave the optimal complexation performance. Kinetic studies showed that high-molecular weight/high-charge density polymer samples produced very stable complexes. The fully charged polyDMAEMA displayed the strongest binding with the ON. These complexes were therefore less sensitive to the changes in the environment. PolyDMAEMA–DMSQ samples had slightly higher complexation ability than polyDMAEMA–MCQ (DMSQ: dimethylsulfate quat; MCQ: methylchloride quat). Both poly(DMAEMA-b-HEMA) and poly(DMAEMA–MCQ-b-PEG) block copolymers showed good complexation ability and steric stability [HEMA: 2-hydroxyethyl methacrylate; PEG: poly(ethylene glycol)]. PEG, but not HEMA block, enhanced the effectiveness of polyDMAEMA–MCQ binding with the ON.     

Site-specific modification of the 5"-terminal fragment of PGY1/MDR1 gene mRNA by reactive conjugates of antisense oligonucleotides

The site-specific modification of the 5"-terminal fragment of PGY1/MDR1 mRNA by oligodeoxyribonucleotide conjugates bearing residues of bleomycin A₅ (Blm), cobalt(ii) tetracarboxyphthalocyanine (Phcn), 4-[N-(2-chloroethyl)-N-methylamino]benzylamine (RCl), or perfluoroarylazide (Az) was studied. Conjugates of oligonucleotides complementary to the RNA sequences 123—138 and 155—166 selectively modify RNA in the vicinity of these regions. The highest efficacy (up to 50%) was achieved in reactions with alkylating and perfluoroarylazide conjugates of oligonucleotides. Conjugates of perfluoroarylazide with 2"-O-modified oligonucleotides are much more efficient than analogous conjugates with oligodeoxyribonucleotides (extents of RNA modification are 40—50% and 20%, respectively).     

Biological efficacy of antisense oligonucleotides complementary to overlapping regions of the mRNA target

Phosphorothioate oligonucleotides complementary to target mRNA are stable in biological milieu and are capable of decreasing levels of this mRNA and the protein encoded by this mRNA (antisense knockdown). The results of our study are compared with the data published in the literature on the efficacy of three antisense 18—21-mer oligonucleotides, which are targeted to the start codon or nearby sequences of _2A-adrenoceptor mRNA, on receptor expression, and functions regulated by these receptors. The highest biological efficacy was shown by the oligonucleotide, which is complementary to the mRNA region and contains the largest number of unpaired bases in the theoretically calculated conformation corresponding to the free energy minimum. Targeting of both ends of the antisense on unpaired bases of the target also leads to the enhancement of its biological efficacy.     

A novel solid support for the synthesis of 3-aminoalkylated oligonucleotides

A novel improved controlled pore glass (CPG) support based on the 2-(hydroxymethyl)-6-nitrobenzoyl (HMNB) protecting group was developed for the synthesis of 3^′-aminoalkylated oligonucleotides. The release of oligonucleotides with free 3^′-amino groups from the support is complete within 2 h at 55 °C in concentrated ammonia.     

Introducing LNAzo: More Rigidity for Improved Photocontrol of Oligonucleotide Hybridization**

Oligonucleotide-based therapeutics have made rapid progress in clinical treatment of a variety of disease indications. Since most therapeutic oligonucleotides serve more than just one function and tend to have a prolonged lifetime, spatio-temporal control of these functions would be desirable. Photoswitches like azobenzene have proven themselves as useful tools in this matter. Upon irradiation, the photoisomerization of the azobenzene moiety causes destabilization in adjacent base pairs, leading to a decreased hybridization affinity. Since the way the azobenzene is incorporated in the oligonucleotide is of utmost importance, we synthesized locked azobenzene C-nucleosides and compared their photocontrol capabilities to established azobenzene C-nucleosides in oligonucleotide test-sequences by means of fluorescence-, UV/Vis-, and CD-spectroscopy.     

Simple reagent for the synthesis of oligonucleotides labeled with 3,3,3′,3′-tetramethyl-2,2′-indodicarbocyanine

Synthesis of a phosphoramidite reagent for the preparation of oligonucleotides labeled at the 5′-end with a fluorescent dye, 3,3,3′,3′-tetramethyl-2,2′-indodicarbocyanine, is described. The efficiency of this reagent is confirmed by the synthesis of several labeled oligonucleotides. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 1, pp. 154–158, January, 2006.     

Spectroscopic studies on the interaction of cilostazole with iodine and 2,3-dichloro-5,6-dicyanobenzoquinone

Lanthanide sensitized luminescence and chemiluminescence (CL) are of great importance because of the unique spectral properties, such as long lifetime, large Stokes shifts, and narrow emission bands characteristic to lanthanide ions (Ln³⁺). With the fluoroquinolone (FQ) compounds including enoxacin (ENX), norfloxacin (NFLX), lomefloxacin (LMFX), fleroxacin (FLRX), ofloxacin (OFLX), rufloxacin (RFX), gatifloxacin (GFLX) and sparfloxacin (SPFX), the luminescence and CL properties of Tb³⁺–FQ and Eu³⁺–FQ complexes have been investigated in this contribution. Ce⁴⁺–SO₃²⁻ in acidic conditions was taken as the CL system and sensitized CL intensities of Tb³⁺–FQ and Eu³⁺–FQ complexes were determined by flow-injection analysis. The luminescence and CL spectra of Tb³⁺–FQ complexes show characteristic peaks of Tb³⁺ at 490 nm, 545 nm, 585 nm and 620 nm. Complexes of Tb³⁺–ENX, Tb³⁺–NFLX, Tb³⁺–LMFX and Tb³⁺–FLRX display relatively strong emission intensity compared with Tb³⁺–OFLX, Tb³⁺–RFX, Tb³⁺–GFLX and Tb³⁺–SPFX. Quite weak peaks with unique characters of Eu³⁺ at 590 nm and 617 nm appear in the luminescence and CL spectra of Eu³⁺–ENX, but no notable sensitized luminescence and CL of Eu³⁺ could be observed when Eu³⁺ is added into other FQ. The distinct differences on emission intensity of Tb³⁺–FQ and Eu³⁺–FQ might originate from the different energy gap between the triplet levels of FQ and the excited levels of the Ln³⁺. The different sensitized luminescence and CL signals among Tb³⁺–FQ complexes could be attributed to different optical properties and substituents of these FQ compounds. The detailed mechanism involved in the luminescence and CL properties of Tb³⁺–FQ and Eu³⁺–FQ complexes has been investigated by analyzing the luminescence and CL spectra, quantum yields, and theoretical calculation results.     

Polymorphism of hydrogen bonding in the short double helixes of oligonucleotides: Semiempirical quantum chemical study

The origin of heterogeneity of nucleotide steps geometry in short double helixes is studied theoretically. By using the semiempirical MNDO/PM3 technique, the stability of “propeller‐like” and “step‐like” forms of base H‐pairing is examined in the structure of oligonucleotide duplexes of different types. The influence of end effects on the process of nucleotides packing, as well as the dependence of duplex curvature on the nature of bonded oligonucleotides, are examined. It is concluded that the structural polymorphism of base pairs most likely determines the unique packing of complementary pairs and their flexibility in DNA structure. © 2002 Wiley Periodicals, Inc. Int J Quantum Chem, 2002     

The MOE Modification of RNA: Origins and Widescale Impact on the Oligonucleotide Therapeutics Field

In an article published by Helvetica Chimica Acta in 1995, chemist P. Martin describes the synthesis of 2′-O-alkylated ribonucleosides for use in therapeutic antisense oligonucleotides (ASOs). This work was motivated by the need for a modified ribose structure that was compatible with solid-phase synthesis protocols and that, when incorporated into an oligonucleotide, would render it resistant to nucleases without attenuating its ability to hybridize to a complementary RNA target. Martin described a robust route to 2′-O-alkylribonucleosides in which the ribose 2′-OH group is substituted with 2′-ethylene glycol derivatives. Oligonucleotides containing these modifications displayed überraschende Eigenschaften – ‘surprising properties’ – notably, higher affinity and specificity for RNA substrates and greater stability to nucleases relative to their unmodified counterparts. Today, the 2′-ethylene glycol modification is universally known in the field as the 2′-O-methoxyethyl (MOE) modification. The chemistry features in four ASO drugs and many others in clinical trials. Here, we 1) summarize the synthesis of the MOE-modified ribose; 2) outline the properties of MOE-modified oligonucleotides as reported in Martin’s article; 3) highlight the first approved MOE-modified ASO drugs, mipomersen and nusinersen; and 4) survey MOE-modified ASOs in clinical development. In the outlook, we put these developments into context and consider future possibilities for the MOE modification.     

Chirality at phosphorus: hybrid duplexes of chimeric oligonucleotides containing methylphosphonothioate linkages with complementary DNA and RNA

Chimeric oligonucleotides with incorporated diastereomerically pure dinucleoside(3′,5′)-methylphosphonothioates and their oxo- and seleno- congeners of known absolute configuration are reported. The relation between stability of the hybrid duplexes with complementary DNA and RNA and their structure is analyzed in context of absolute configuration of the P-chiral internucleotide bonds.     

Hybridization of Nafion membranes with an acid functionalised polysiloxane: Effect of morphology on water sorption and proton conductivity

Polysiloxane-modified hybrid Nafion membranes were prepared by casting a mixture of Nafion solution and a precursor of acid functionalised polysiloxane based on tetraethoxysilane and a mercaptan-organoalkoxysilane.Scanning Electron Microscopy (SEM) and Atomic Force Microscopy analysis revealed that the functionalised polysiloxane was dispersed either as finely nanosized inclusions or as coarse domains depending on the rate of the solvent evaporation during the casting procedure. In particular the slower is the rate of solvent evaporation the more interpenetrated and homogenously dispersed at nanosized level is the polysiloxane inside the Nafion membrane.The hybridization process increases the thermal stability of the membranes of about 50 °C relatively to the unmodified Nafion. Small angle X-ray scattering (SAXS) analysis reveals that the hybrid membranes exhibited the typical morphology of Nafion consisting of distinct hydrophilic and hydrophobic domains.Water vapor sorption and proton conductivity were measured varying the temperature (up to 120 °C) and the water activity conditions (from 0.1 to 0.8). The polysiloxane network always increases the water vapor uptake of the membranes and increases significantly the proton conductivity at higher temperature depending on the type of morphology developed by the manufacturing method. In particular hybrid membranes exhibiting nanosized polysiloxane dispersion show a proton conductivity which is up to one-and-half time higher than Nafion recast membrane at high temperature and low water content.     

The statistical generation of SCS values and interaction factors for the13C NMR spectra of arenes

A new methodology which statistically determines substituent chemical shifts (SCS) and interaction factors of adjacent groups for use in the³C NMR spectroscopy of arenes is described. From a data base of 15,255 signals we have computed several hundred of thesestatistical SCS values (SSCS values), and interaction factors for many common pairs of groups. Some interaction factors for 1,2,3-substitution are also reported. When these SSCS values and interaction factors are used to predict the signal position they are found to give a correlation coefficient of predicted verses observed values of 0.994 with a standard deviation of 1.5 ppm.