Homocystamide Conjugates of Human Serum Albumin as a Platform to Prepare Bimodal Multidrug Delivery Systems for Boron Neutron Capture Therapy.

Boron neutron capture therapy is a unique form of adjuvant cancer therapy for various malignancies including malignant gliomas. The conjugation of boron compounds and human serum albumin (HSA)-a carrier protein with a long plasma half-life-is expected to extend systemic circulation of the boron compounds and increase their accumulation in human glioma cells. We report on the synthesis of fluorophore-labeled homocystamide conjugates of human serum albumin and their use in thiol-'click' chemistry to prepare novel multimodal boronated albumin-based theranostic agents, which could be accumulated in tumor cells. The novelty of this work involves the development of the synthesis methodology of albumin conjugates for the imaging-guided boron neutron capture therapy combination. Herein, we suggest using thenoyltrifluoroacetone as a part of an anticancer theranostic construct: approximately 5.4 molecules of thenoyltrifluoroacetone were bound to each albumin. Along with its beneficial properties as a chemotherapeutic agent, thenoyltrifluoroacetone is a promising magnetic resonance imaging agent. The conjugation of bimodal HSA with undecahydro-closo-dodecaborate only slightly reduced human glioma cell line viability in the absence of irradiation (~30 µM of boronated albumin) but allowed for neutron capture and decreased tumor cell survival under epithermal neutron flux. The simultaneous presence of undecahydro-closo-dodecaborate and labeled amino acid residues (fluorophore dye and fluorine atoms) in the obtained HSA conjugate makes it a promising candidate for the combination imaging-guided boron neutron capture therapy.

New synthetic substrates of mammalian nucleotide excision repair system.

DNA probes for the studies of damaged strand excision during the nucleotide excision repair (NER) have been designed using the novel non-nucleosidic phosphoramidite reagents that contain N-[6-(9-antracenylcarbamoyl)hexanoyl]-3-amino-1,2-propandiol (nAnt) and N-[6-(5(6)-fluoresceinylcarbamoyl)hexanoyl]-3-amino-1,2-propandiol (nFlu) moieties. New lesion-imitating adducts being inserted into DNA show good substrate properties in NER process. Modified extended linear nFlu- and nAntr-DNA are suitable for estimation of specific excision activity catalysed with mammalian whole-cell extracts. The following substrate activity range was revealed for the model 137-bp linear double-stranded DNA: nAnt-DNA ≈ nFlu-DNA > Chol-DNA (Chol-DNA--legitimate NER substrate that contains non-nucleoside fragment bearing cholesterol residue). In vitro assay shows that modified DNA can be a useful tool to study NER activity in whole-cell extracts. The developed approach should be of general use for the incorporation of NER-sensitive distortions into model DNAs. The new synthetic extended linear DNA containing bulky non-nucleoside modifications will be useful for NER mechanism study and for applications.

Ligand-directed acid-sensitive amidophosphate 5-trifluoromethyl-2'-deoxyuridine conjugate as a potential theranostic agent.

Herein, we report a novel strategy to engineer an acid-sensitive anticancer theranostic agent using a vector-drug ensemble. The ensemble was synthesized by directly conjugating the linoleic acid (LA)-modified branched polyethyleneimine with a chemotherapeutic drug trifluorothymidine. Linoleic acid residues were grafted onto 25 kDa polyethyleneimine (PEI) by treating PEI with linoleic acid chloroanhydride. 5-Trifluoromethyl-2'-deoxyuridine (trifluorothymidine, TFT) was introduced into LA-PEI conjugate by phosphorylating the conjugate with amidophosphate of trifluorothymidine 5'-monophosphate (pTFT), which had been activated by its conversion into the N,N-dimethylaminopyridine derivative. The extent of mononucleotide analog incorporation in the polymer was regulated by the ratio of pTFT to the polymer during the synthesis. Samples containing 20-70 TFT residues per PEI molecule were obtained. The cytotoxicity of PEI-LA-pTFT conjugates decreased with increasing nucleotide content, as examined using the MTT method. Due to the presence of fluorine atoms, TFT-based conjugates could be detected directly in the animals by (19)F magnetic resonance imaging. In addition, the presence of the amidophosphate group in PEI-LA-pTFT conjugates allowed their detection by in vivo(31)P NMR spectroscopy. Indeed, the (31)P NMR signal of a phosphoramide (δ ~ 12 ppm) was observed in the mouse muscle tissue treated with PEI-LA-pTFT conjugate along with the signals from endogenous phosphorus-containing compounds. At the same time, the use of PEI-LA-pTFT conjugate for chemotherapeutic drug delivery is limited due to the low release of pTFT from the carrier. To enhance the release of the drug from the conjugate in the endosomes, PEI-LA polymer was coupled with urocanic acid (UA), which bears imidazole ring and thus can form an acid-labile P-N bond with pTFT. The PEI-LA-UA-pTFT conjugate containing 30 residues of UA and 40 residues of pTFT was tested against the murine Krebs-II ascites carcinoma, grown as an ascetic tumor. The intraperitoneal injection of the conjugates resulted in prolongation of the animals' life and to the complete disappearance of the tumor after three injections.

Inactivation of the tick-borne encephalitis virus by RNA-cleaving compounds.

The tick-borne encephalitis virus (TBEV) is an RNA-containing enveloped virus, which poses a major threat to the well-being and health of humans. In this study, we describe an approach to the inactivation of TBEV, which involves the degradation of viral RNA by artificial ribonucleases (aRNases, small organic compounds that exhibit ribonuclease activity in vitro). We demonstrate that the incubation of TBEV with aRNases lead to the total inactivation of the virus as indicated by the plaque formation assay data, but retain the viral immunogenic properties, as shown by the ELISA data. We propose that a possible mechanism of TBEV inactivation with aRNase, which includes: i) formation of local breaks in the lipid membrane of the virus caused by aRNase, ii) penetration of aRNase into the viral capsid, iii) degradation of genomic RNA by aRNase. These data suggest that the proposed approach can be used in the production of killed-virus vaccine.

Per aspera ad astra: application of Simplex QSAR approach in antiviral research.

This review explores the application of the Simplex representation of molecular structure (SiRMS) QSAR approach in antiviral research. We provide an introduction to and description of SiRMS, its application in antiviral research and future directions of development of the Simplex approach and the whole QSAR field. In the Simplex approach every molecule is represented as a system of different simplexes (tetratomic fragments with fixed composition, structure, chirality and symmetry). The main advantages of SiRMS are consideration of the different physical-chemical properties of atoms, high adequacy and good interpretability of models obtained and clear procedures for molecular design. The reliability of developed QSAR models as predictive virtual screening tools and their ability to serve as the basis of directed drug design was validated by subsequent synthetic and biological experiments. The SiRMS approach is realized as the complex of the computer program 'HiT QSAR', which is available on request.

Design and synthesis of dinucleotide 5'-triphosphates with expanded functionality.

We propose the new approach to the synthesis of 5'-triphosphate derivatives of natural and modified dinucleotides with expanded functionality. Our strategy includes the combination of the solution phase synthesis of necessary dimers using the wide range of nucleic acids chemistry methods and the subsequent introduction of the triphosphate residue. A number of the new potential substrates for the template dependent synthesis of nucleic acids with expanded functionality are obtained, namely, 5'-triphosphates of dinucleotides containing the functionally active groups in heterocyclic bases, in carbohydrate-phosphate backbone, and the groups mimicking the residues of natural amino acids. The abilities of the proposed synthetic route are also demonstrated by the synthesis of 5'-triphosphates of dinucleotides with modified carbohydrate-phosphate backbone.

Design and synthesis of peptide-oligonucleotide conjugates as potential artificial ribonucleases.

A series of peptide-oligonucleotide conjugates was synthesized. The peptide fragment was attached to the middle part of DNA sequence. For this purpose the new amidophosphite dC with linker at C5-position was synthesized.

Design and synthesis of metal-free artificial ribonucleases.

The present review is aimed at giving a general overview of our results in the field of designing and synthesizing simple peptide-like molecules that mimic structural and functional aspects of natural ribonucleases, as well as designing oligonucleotide-based artificial ribonucleases.