Phosphoramidate Azole Oligonucleotides for Single Nucleotide Polymorphism Detection by PCR.

Detection of the Kirsten rat sarcoma gene (KRAS) mutational status is an important factor for the treatment of various malignancies. The most common KRAS-activating mutations are caused by single-nucleotide mutations, which are usually determined by using PCR, using allele-specific DNA primers. Oligonucleotide primers with uncharged or partially charged internucleotide phosphate modification have proved their ability to increase the sensitivity and specificity of various single nucleotide mutation detection. To enhance the specificity of single nucleotide mutation detection, the novel oligonucleotides with four types of uncharged and partially charged internucleotide phosphates modification, phosphoramide benzoazole (PABA) oligonucleotides (PABAO), was used to prove the concept on the KRAS mutation model. The molecular effects of different types of site-specific PABA modification in a primer or a template on a synthesis of full-length elongation product and PCR efficiency were evaluated. The allele-specific PCR (AS-PCR) on plasmid templates showed a significant increase in analysis specificity without changes in Cq values compared with unmodified primer. PABA modification is a universal mismatch-like disturbance, which can be used for single nucleotide polymorphism discrimination for various applications. The molecular insights of the PABA site-specific modification in a primer and a template affect PCR, structural features of four types of PABAO in connection with AS-PCR results, and improvements of AS-PCR specificity support the further design of novel PCR platforms for various biological targets testing.

Synthesis of Oligonucleotides Carrying Inter-nucleotide N-(Benzoazole)-phosphoramide Moieties.

In this work, we present new oligonucleotide derivatives containing inter-nucleotide N-benzimidazole, N-benzoxazole, N-benzothiazole, and 1,3-dimethyl-N-benzimidazole (benzoazoles) phosphoramide groups. These modifications were introduced via the Staudinger reaction with appropriate azides during standard automated solid-phase oligonucleotide synthesis. The principal structural difference between the new azido modifiers and those already known is that they are bulk heterocyclic structures, similar to purine nucleoside bases. Modified oligonucleotides with one and two modifications at different positions and multiple modified heteronucleotide sequences were obtained with high yields. The possibility of multiple modifications in the process of automatic DNA synthesis is fundamental and critical for further application of our oligonucleotide derivatives. Initial studies on the properties of new oligonucleotides were carried out. The stability of the oligodeoxyribonucleotide duplex containing phosphoramide groups of N-benzoazoles with complementary DNA or RNA is slightly lower than that of native complexes.

Synthesis of the new nucleoside 5'-alpha-iminophosphates using Staudinger reaction.

Efficient protocols were developed for the synthesis of a new compounds - nucleoside 5'-α-iminophosphates using the Staudinger reaction. These substances are alpha-phosphate mimetic nucleotide in which an oxygen atom is replaced by a corresponding imino (=N-R)-group. Various 5'-iminomonophosphates of nucleosides were obtained. A chemical method for the synthesis of triphosphate derivatives based on the iminomonophosphates has been designed. Thymidine 5'-(1,3-dimethylimidazolidin-2-ylidene)-triphosphate (ppp(DMI)T) was synthesized, its hydrolytic stability and substrate properties in relation to some DNA polymerases was firstly studied. It was shown that ppp(DMI)T can serve as substrate for enzyme catalyzed template-independent DNA synthesis by human terminal deoxynucleotidyltransferase TdT.

Data set on the synthesis and properties of 2',3'-dideoxyuridine triphosphate conjugated to SiO2 nanoparticles.

SiO2 nanoparticles were used as a transport system for cellular delivery of phosphorylated 2',3'-dideoxyuridine to increase its anticancer potency. This data set is related to the research article entitled "2',3'-Dideoxyuridine triphosphate conjugated to SiO2 nanoparticles: synthesis and evaluation of antiproliferative activity" (Vasilyeva et al., 2018) [1]. It includes a protocol for the synthesis of 2',3'-dideoxyuridine-5'-{N-[4-(prop-2-yn-1-yloxy)butyl]-γ-amino}-triphosphate, its identification by NMR, IR and ESI-MS, experimental procedure of covalent attachment to SiO2 nanoparticles with via Cu-catalyzed click-chemistry, experimental data on chemical stability of the conjugate at different pH values and cytotoxicity assessment of antiproliferative effect of the conjugate.

2',3'-Dideoxyuridine triphosphate conjugated to SiO2 nanoparticles: Synthesis and evaluation of antiproliferative activity.

A conjugate of triphosphorylated 2',3'-dideoxyuridine (ddU) with SiO2 nanoparticles was obtained via the CuAAC click chemistry between a γ-alkynyl ddU triphosphate and azido-modified SiO2 nanoparticles. Assessment of cytotoxicity in human breast adenocarcinoma MCF7 cells demonstrated that ddU triphosphate conjugated to SiO2 nanoparticles exhibited a 50% decrease in cancer cell growth at a concentration of 183 ±â€¯57 µg/mL, which corresponds to 22 ±â€¯7 µM of the parent nucleotide, whereas the parent nucleoside, nucleotide and alkynyl triphosphate precursor do not show any cytotoxicity. The data provide an example of remarkable potential of novel conjugates of SiO2 nanoparticles with phosphorylated nucleoside analogues, even those, which have not been used previously as therapeutics, for application as new anticancer agents.

DNA fluorescent labeling with naphtho[1,2,3-cd]indol-6(2H)-one for investigation of protein-DNA interactions.

Fluorescently labeled DNA to study protein-DNA interactions was synthesized using the Cu(I)-catalysed cycloaddition (CuAAC) reaction. For this purpose, a new azido-containing fluorophore based on the naphtho[1,2,3-cd]indol-6(2H)-one derivative was obtained. The fluorescent properties of naphtho[1,2,3-cd]indol-6(2H)-one derivatives and labeled DNA were studied. The new fluorescent DNA conjugate was shown to be a useful tool to study complex mechanisms of protein-DNA interactions.

Conjugates of phosphorylated zalcitabine and lamivudine with SiO2 nanoparticles: Synthesis by CuAAC click chemistry and preliminary assessment of anti-HIV and antiproliferative activity.

Conjugates of phosphorylated dideoxynucleoside antiviral drugs dideoxycytidine (zalcitabine) and lamivudine with SiO2 nanoparticles were obtained via the copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) click chemistry between a nucleoside triphosphate containing an alkynyl group at the γ-phosphate or azidothymidine triphosphate and SiO2 nanoparticles containing alkyl azide or alkynyl groups, respectively. 4-(Prop-2-yn-1-yloxy)butylamino group has been attached to the γ-phosphate group of dideoxycytidine (zalcitabine) and lamivudine 5'-triphosphates via the phosphoramidate linkage. New compounds were shown to be potent killers of human colon carcinoma cells. Anti-HIV activity of the conjugates was demonstrated as well. The conjugates of phosphorylated lamivudine and dideoxycytidine (zalcitabine) showed higher potency than the parent nucleosides. The conjugate of phosphorylated azidothymidine was less active against HIV-1 than the parent nucleoside probably because of the replacement of its 3'-azido group by 1,2,3-triazole ring. These results show an opportunity for using SiO2 nanoparticles as a transport for delivering phosphorylated nucleosides to cells in order to increase their efficiency as antiviral and anticancer drugs.

Application of Cu(I)-catalyzed azide-alkyne cycloaddition for the design and synthesis of sequence specific probes targeting double-stranded DNA.

Efficient protocols based on Cu(I)-catalyzed azide-alkyne cycloaddition were developed for the synthesis of conjugates of pyrrole-imidazole polyamide minor groove binders (MGB) with fluorophores and with triplex-forming oligonucleotides (TFOs). Diverse bifunctional linkers were synthesized and used for the insertion of terminal azides or alkynes into TFOs and MGBs. The formation of stable triple helices by TFO-MGB conjugates was evaluated by gel-shift experiments. The presence of MGB in these conjugates did not affect the binding parameters (affinity and triplex stability) of the parent TFOs.

On Field-Effect Photovoltaics: Gate Enhancement of the Power Conversion Efficiency in a Nanotube/Silicon-Nanowire Solar Cell.

Recent years have seen a resurgence of interest in crystalline silicon Schottky junction solar cells distinguished by the use of low density of electronic states (DOS) nanocarbons (nanotubes, graphene) as the metal contacting the Si. Recently, unprecedented modulation of the power conversion efficiency in a single material system has been demonstrated in such cells by the use of electronic gating. The gate field induced Fermi level shift in the low-DOS carbon serves to enhance the junction built-in potential, while a gate field induced inversion layer at the Si surface, in regions remote from the junction, keeps the photocarriers well separated there, avoiding recombination at surface traps and defects (a key loss mechanism). Here, we extend these results into the third dimension of a vertical Si nanowire array solar cell. A single wall carbon nanotube layer engineered to contact virtually each n-Si nanowire tip extracts the minority carriers, while an ionic liquid electrolytic gate drives the nanowire body into inversion. The enhanced light absorption of the vertical forest cell, at 100 mW/cm(2) AM1.5G illumination, results in a short-circuit current density of 35 mA/cm(2) and associated power conversion efficiency of 15%. These results highlight the use of local fields as opposed to surface passivation as a means of avoiding front surface recombination. A deleterious electrochemical reaction of the silicon due to the electrolyte gating is shown to be caused by oxygen/water entrained in the ionic liquid electrolyte. While encapsulation can avoid the issue, a nonencapsulation-based approach is also implemented.

SiO2 nanoparticles as platform for delivery of 3'-triazole analogues of AZT-triphosphate into cells.

A system for delivery of analogues of AZT-triphosphates (AZT*TP) based on SiO2 nanoparticles was proposed. For this purpose, a simple and versatile method was developed for the preparation of SiO2∼dNTP conjugates using the 'click'-reaction between AZTTP and premodified nanoparticles containing the alkyne groups. The substrate properties of SiO2∼AZT*TP were tested using Klenow fragment and HIV reverse transcriptase. The 3'-triazole derivatives of thymidine triphosphate being a part of the SiO2∼AZT*TP nanocomposites were shown to be incorporated into the growing DNA chain. It was shown by confocal microscopy that the proposed SiO2∼AZT*TP nanocomposites penetrate into cells. These nanocomposites were shown to inhibit the reproduction of POX and Herpes viruses at nontoxic concentrations.

Novel fluorescent pyrimidine nucleosides containing 2,1,3-benzoxadiazole and naphtho-[1,2,3-CD] Indole-6 (2H)-one fragments.

A series of novel fluorescent pyrimidine nucleosides containing 2,1,3-benzoxadiazole or naphtho[1,2,3-cd]indole-6 (2h)-one fragments was designed and synthesized. Introduction of fluorescent fragments into the position 5 of the uridine or cytidine heterocycle was carried out in two ways: by Sonogashira Coupling Reaction and CuI-catalyzed cycloaddition ("click" reaction). The obtained nucleoside derivatives became fluorescent due to the inserted fragments. The excitation wavelength (440-450 nm) was outside the absorption band of many biomolecules and significantly differed from the emission wavelength (560-600 nm). In addition, the intended nucleoside analogs were shown to kill cultured human tumor cells at submicromolar concentrations.

Extraordinary hydrogen evolution and oxidation reaction activity from carbon nanotubes and graphitic carbons.

The hydrogen evolution reaction, 2H(+) + 2e(­) → H2, and its converse, the hydrogen oxidation reaction, H2 → 2H(+) + 2e(­), are central to any realization of a hydrogen economy. Various forms of carbon have been used for decades as the precious metal catalyst support in these reactions. Here we report the unexpected result that single-wall carbon nanotubes and some graphitic carbons, activated by brief exposure to electrochemical potentials that induce hydrogen evolution in intercalating acids combined with extended soak times in such acids, acquire an activity for these reactions that exceeds that of known nonprecious metal catalysts.

Oligonucleotide functionalization by a novel alkyne-modified nonnucleosidic reagent obtained by versatile building block chemistry.

A convenient synthetic strategy has been designed to prepare an alkyne-modified synthon for automated DNA synthesis. It is based on the key O-DMTr-protected 4-(2-hydroxyethyl)morpholin-2,3-dione and building blocks obtained by its functionalization by various aliphatic amines. A respective nonnucleosidic phosphoramidite monomer containing a terminal alkyne in the side-chain was synthesized, and corresponding oligothymidylates incorporating the modification in various positions were prepared. The presence of the alkyne group was confirmed by Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) between the functionalized oligonucleotide and an azide derivative of 7-nitro-2,1,3-benzoxadiazole.

SiO2 nanoparticles as platform for delivery of nucleoside triphosphate analogues into cells.

A system for delivery of analogues of 2'-deoxyribonucleoside triphosphate (dNTP) based on SiO(2) nanoparticles was proposed. A simple and versatile method was developed for the preparation of SiO(2)-dNTP conjugates using the 'click'-reaction between premodified nanoparticles containing the azido groups and dNTP containing the alkyne-modified γ-phosphate group. The substrate properties of SiO(2)-dNTP were tested using Klenow fragment and HIV reverse transcriptase. Nucleoside triphosphates being a part of the SiO(2)-dNTP nanocomposites were shown to be incorporated into the growing DNA chain. The rate of polymerization with the use of SiO(2)-dNTP or common dNTP in case of HIV reverse transcriptase differed insignificantly. It was shown by confocal microscopy that the proposed SiO(2)-dNTP nanocomposites bearing the fluorescent label penetrate into cells and even into cellular nuclei.

Synthesis of novel nucleoside 5'-triphosphates and phosphoramidites containing alkyne or amino groups for the postsynthetic functionalization of nucleic acids.

A series of novel nucleoside 5'-triphosphates and phosphoramidites containing alkyne or amino groups for the postsynthetic functionalization of nucleic acids were designed and synthesized. For this purpose, the new 3-aminopropoxypropynyl linker group was used. It contains two alternative functional capabilities: an amino group for the reaction of amino-alkynyl-modified oligonucleotides with corresponding activated esters and an alkyne group for the copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reaction. It was shown that a variety of methods of the attachment of the new linker can be used to synthesize a diversity of modified pyrimidine nucleosides.

Material strategies for black-to-transmissive window-type polymer electrochromic devices.

Black-to-transmissive switching polymer electrochromic devices (ECDs) were designed using a set of spray-processable cathodically coloring polymers, a non-color-changing electroactive polymer poly(2,2,6,6-tetramethylpiperidinyloxy-4-yl methacrylate) (PTMA) as the charge-compensating counter electrode, and a highly conducting gel electrolyte (6.5 mS cm(-1)). The color "black" was obtained by utilizing (1) individual copolymers absorbing across the visible spectrum, and (2) blends and bilayers of several polymer electrochromes with complementary spectral absorption. Neutral-state black and ink-like dark purple-blue (or "ink-black") donor-acceptor (DA) copolymers composed of the electron-donor 3,4-propylenedioxythiophene (ProDOT) and the electron-acceptor 2,1,3-benzothiadiazole (BTD) building units, which possess relatively homogeneous absorption profiles across the visible spectrum, were chosen for their propensity to switch to transmissive states upon electrochemical oxidation. A blend of magenta and cyan polymers (PProDOT-(CH(2)OEtHx)(2) and P(ProDOT-BTD-ProDOT), respectively) was produced with the goal of generating the same dark purple-blue color as that obtained with the "ink-black" DA copolymer. While the multi-polymer ECDs demonstrate high contrasts (up to 50%T), and switch from a saturated purple-blue color (L*=32, a*=13, b*=-46) to a light green-blue transmissive state (L*=83, a*=-3, b*=-6), devices made with the DA electrochromic copolymers switch more than two times faster (0.7 s to attain 95% of the full optical change) than those involving the polymer blends (1.6 s), and exhibit more neutral achromatic colors (L*=38, a*=5, b*=-25 for the colored state and L*=87, a*=-3, b*=-2 for the bleached state, correspondingly). The results obtained suggest that these materials should prove to be applicable in both transmissive- (window-type) and reflective-type ECDs.

Color purity in polymer electrochromic window devices on indium-tin oxide and single-walled carbon nanotube electrodes.

Dual polymer absorptive/transmissive electrochromic (EC) window devices have been assembled using the solution-processable and high-EC-contrast polymer PProDOT-(CH(2)OEtHx)(2) as the EC material, along with a non-color-changing electroactive polymer, poly(2,2,6,6-tetramethylpiperidinyloxy-4-yl methacrylate) (PTMA), as the counter electrode material. Indium-tin oxide (ITO) and highly transmissive single-walled carbon nanotube (SWNT) film coated glass electrodes are used as electrode substrates. The use of the EC/non-color-changing polymer combination allowed us to construct window devices that rapidly switch between magenta and highly transmissive (>95% T for ITO and approximately 79% T for SWNT) states with large optical modulation (>71% DeltaT for ITO and 66% DeltaT for SWNT). The devices showed effective coloration and bleaching: the lightness parameter (L*) changing from 67 to 95 for ITO (approximately 50-92 for SWNT), essentially reaching a diffuse white upon oxidation. The color modulates from highly pure magenta with a* = 28 (red hue) and b* = -28 (blue chroma) for ITO (a* = 40 and b* = -36 for SWNT) to nearly colorless with a* = 1 and b* = -1 for ITO (a* = -2 and b* = -3 for SWNT) devices. Increasing the switching voltage from 2.55 V up to 3.5 V resulted in faster SWNT-based window device performance.

Metallocene-containing conjugated polymers.

The paper gives a review of publications on polymers with conjugated matrices (PPy, PTh, PAni, hydrocarbon or mixed chains...) which incorporate metallocene complexes (Fe, Ru, Co; Ni, Ti, Zr, Ta) with two cyclopentadienyl ligands (Cp) and their derivatives, in particular with methylated cyclopentadienyl rings (Cp*), as well as hemi-metallocene complexes (Fe, Ru, Co, Mn), as pendant groups or inside the principal chain (part B). The information on related short-chain systems, monomers and oligomers, is also included. In part A, a brief overview of various conjugated polymer materials is presented, with their classification in accordance with the conductivity mechanism (ionic, electronic or mixed conductors) or with the structural type (linear-chain organic or mixed polymers, derivatization, metallopolymers, multi-dimensional structures, alternating and block copolymers with organic or mixed units, hybrid materials with a mixture of conjugated and inert polymers, polymers inside a solid matrix, conjugated polymers with incorporated nanoelements of transition metals, carbon, semiconductors etc.

Synthesis of monomers bearing at the 2'-position cyanomethoxycarbonyl group for phosphoramidite oligonucleotide synthesis.

A novel series of phosphoroamidites for the synthesis of 2-modified oligonucleotides was designed and synthesized on the base of 2'-amino uridine and 2'-amino arabinoadenosine. The amino groups in these compounds were acidified by bis-cyanomethyl esters of different dicarbonic acids. Generated reactive linker groups containing cyanomethoxycarbonyl groups are stable under conditions of oligonucleotide synthesis but could be easily functionalised in post-synthetic stage by treatment with compounds bearing primary amino groups.