Design, synthesis, and anti-respiratory syncytial virus potential of novel 3-(1,2,3-triazol-1-yl)furoxazine-fused benzimidazole derivatives.

Respiratory syncytial virus (RSV) is a major cause of serious lower respiratory tract infections in infants, children, and older persons. Currently, the only approved anti-viral chemotherapeutic drug for RSV treatment is ribavirin aerosol; however, its significant toxicity has led to restricted clinical use. In a previous study, we developed various benzimidazole derivatives against RSV. In this study, we synthesised 3-azide substituted furoxazine-fused benzimidazole derivatives by sulfonylation and azide substitution of the 3-hydroxyl group of the furoxazine-fused benzimidazole derivatives. Subsequently, a series of 3-(1,2,3-triazol-1-yl)-substituted furoxazine-fused benzimidazole derivatives were synthesised using the classical click reaction. Biological evaluations of the target compounds indicated that compound 4a-2 had higher activity against RSV (EC50 = 12.17 µM) and lower cytotoxicity (CC50 = 390.64 µM). Compound 4a-2 exerted anti-viral effects against the RSV Long strain by inhibiting apoptosis and the elevation of reactive oxygen species (ROS) and inflammatory factors caused by viral infection in vitro. Additionally, the clinical symptoms of the virus-infected mice were markedly relieved, and the viral load in the lung tissues was dramatically decreased. The biosafety profile of compound 4a-2 was also favourable, showing no detectable adverse effects on any of the major organs in vivo. These findings underscore the potential of compound 4a-2 as a valuable therapeutic option for combating RSV infections while also laying the foundation for further research and development in the field.

Design, synthesis and antitumour activity of novel 5(6)-amino-benzimidazolequinones containing a fused morpholine.

Based on the previous synthesis of tetracyclic and tricyclic benzimidazoles starting from 1,4:3,6-dianhydro-d-fructose and o-phenylenediamines, a series of 5(6)-amino substituted tetracyclic and tricyclic benzimidazolequinones were obtained through the oxidation of 4,7-dimethoxy-benzimidazole analogues with bis(trifluoroacetoxy)iodobenzene (PIFA) and subsequent substitution with various aliphatic and aromatic amines. Biological evaluations of the target benzimidazolequinones indicated that all the arylamino-substituted benzimidazolequinones possess potent antitumour activity against human gastric cancer cells (MGC-803), especially compound a21-2. Furthermore, compound a21-2 inhibits gastric cancer cells proliferation and cell colony formation. Mechanistic investigations showed that compound a21-2 induces ROS production, which subsequently causes DNA damage and activation of ATM/Chk2, leading to G2/M phase arrest. ROS activates the c-Jun N-terminal kinase (JNK) pathway to induce mitochondrial-mediated apoptosis. In vivo studies showed that compound a21-2 inhibits the growth of tumours in nude mice without significant systemic toxicity. These findings suggest that compound a21-2 represents a promising candidate antitumour drug.

Design, synthesis, in vitro and in vivo anti-respiratory syncytial virus (RSV) activity of novel oxizine fused benzimidazole derivatives.

Respiratory syncytial virus (RSV) causes serious lower respiratory tract infections. Currently, the only clinical anti-RSV drug is ribavirin, but ribavirin has serious toxic side effect and can only be used by critically ill patients. A series of benzimidazole derivatives were synthesized starting from 1,4:3,6-dianhydro-d-fructose and a variety of o-phenylenediamines. Evaluation of their antiviral activity showed that compound a27 had the highest antiviral activity with a half maximal effective concentration (EC50) of 9.49 µM. Investigation of the antiviral mechanism of compound a27 indicated that it can inhibit the replication of RSV by inhibiting apoptosis and autophagy pathways. Retinoic acid-inducible gene (RIG)-I, TNF receptor associated factor (TRAF)-3, TANK binding kinase (TBK)-1, interferon regulatory factor (IRF)-3, nuclear factor Kappa-B (NF-κB), interferon (IFN)-ß, Toll-like receptor (TLR)-3, interleukin (IL)-6 were suppressed at the cellular level. Mouse lung tissue was subjected to hematoxylin and eosin (HE) staining and immunohistochemistry, which showed that RSV antigen and M gene expression could be reduced by compound a27. Decreased expression of RIG-I, IRF-3, IFN-ß, TLR-3, IL-6, interleukin (IL)-8, interleukin (IL)-10, inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF)-α was also found in vivo.

Synthesis and in vitro antitumour activity of 4(R)-methyl-3-O-phosphonomethyl-α-l-threose nucleosides.

A series of novel α-l-threose nucleoside phosphonate analogs, 4(R)-methyl-3-O-phosphonomethyl-α-l-threose nucleosides, were synthesized in multistep sequences starting from d-xylose. The synthetic sequence consisted of the following key stages: (i) the multistep synthesis of 1,2-O-isopropylidenyl-4(R)-methyl-3-O-phosphonomethyl-l-threose, (ii) the transformation of 1,2-O-isopropylidenyl sugar into suitable 1,2-di-O-acyl l-threose precursor, and (iii) the construction of target α-l-threose nucleoside phosphonate analogs by Vorbrüggen glycosidation reaction, deprotection of acyl group, and hydrolysis of diethyl group on phosphonate. The target nucleoside phosphonates were evaluated for their antitumour activities in cell culture-based assays. Compound 8g, 2-fluroadenosine phosphonate, showed remarkable activity against human breast cancer cell lines (MCF-7 and MDA-MB-231) with IC50 values of 0.476 and 0.391 µM, corresponding to 41- and 47-fold higher potency than the reference compound 5-FU, respectively. Subsequent investigations found that the compound 8g can inhibit the proliferation of breast cancer cells and cell cloning. The mechanistic studies indicated that compound 8g could cause DNA damage to breast cancer cells through the ATM-Chk1/Chk2-cdc25c pathway, leading to blockage of the G2/M phase cycle of breast cancer cells, which ultimately led to apoptosis. Moreover, 8g could inhibit the PI3K/AKT signaling pathway and induce apoptosis. These results indicate that compound 8g holds promising potential as an antitumour agent.

Crystal structures of thrombin in complex with chemically modified thrombin DNA aptamers reveal the origins of enhanced affinity.

Thrombin-binding aptamer (TBA) is a DNA 15-mer of sequence 5'-GGT TGG TGT GGT TGG-3' that folds into a G-quadruplex structure linked by two T-T loops located on one side and a T-G-T loop on the other. These loops are critical for post-SELEX modification to improve TBA target affinity. With this goal in mind we synthesized a T analog, 5-(indolyl-3-acetyl-3-amino-1-propenyl)-2'-deoxyuridine (W) to substitute one T or a pair of Ts. Subsequently, the affinity for each analog was determined by biolayer interferometry. An aptamer with W at position 4 exhibited about 3-fold increased binding affinity, and replacing both T4 and T12 with W afforded an almost 10-fold enhancement compared to native TBA. To better understand the role of the substituent's aromatic moiety, an aptamer with 5-(methyl-3-acetyl-3-amino-1-propenyl)-2'-deoxyuridine (K; W without the indole moiety) in place of T4 was also synthesized. This K4 aptamer was found to improve affinity 7-fold relative to native TBA. Crystal structures of aptamers with T4 replaced by either W or K bound to thrombin provide insight into the origins of the increased affinities. Our work demonstrates that facile chemical modification of a simple DNA aptamer can be used to significantly improve its binding affinity for a well-established pharmacological target protein.

Origins of the enhanced affinity of RNA-protein interactions triggered by RNA phosphorodithioate backbone modification.

The well-characterized interaction between the MS2 coat protein and its cognate RNA hairpin was used to evaluate changes in affinity as a result of phosphorodithioate (PS2) replacing phosphate by biolayer interferometry (BLI). A structure-based analysis of the data provides insights into the origins of the enhanced affinity of RNA-protein interactions triggered by the PS2 moiety.

Evoking picomolar binding in RNA by a single phosphorodithioate linkage.

RNA aptamers are synthetic oligonucleotide-based affinity molecules that utilize unique three-dimensional structures for their affinity and specificity to a target such as a protein. They hold the promise of numerous advantages over biologically produced antibodies; however, the binding affinity and specificity of RNA aptamers are often insufficient for successful implementation in diagnostic assays or as therapeutic agents. Strong binding affinity is important to improve the downstream applications. We report here the use of the phosphorodithioate (PS2) substitution on a single nucleotide of RNA aptamers to dramatically improve target binding affinity by ∼1000-fold (from nanomolar to picomolar). An X-ray co-crystal structure of the α-thrombin:PS2-aptamer complex reveals a localized induced-fit rearrangement of the PS2-containing nucleotide which leads to enhanced target interaction. High-level quantum mechanical calculations for model systems that mimic the PS2 moiety and phenylalanine demonstrate that an edge-on interaction between sulfur and the aromatic ring is quite favorable, and also confirm that the sulfur analogs are much more polarizable than the corresponding phosphates. This favorable interaction involving the sulfur atom is likely even more significant in the full aptamer-protein complexes than in the model systems.

Synthesis and in vitro antiviral activities of [(dihydrofuran-2-yl)oxy]methyl-phosphonate nucleosides with 2-substituted adenine as base.

The synthesis of [(2',5'-dihydrofuran-2-yl)oxy]methyl-phosphonate nucleosides with a 2-substituted adenine base moiety starting from 2-deoxy-3,5-bis-O-(4-methylbenzoyl)-α-L-ribofuranosyl chloride and 2,6-dichloropurine is described. The key step is the regiospecific and stereoselective introduction of a phosphonate synthon at C(2) of the furan ring. None of the synthesized compounds showed significant in vitro activity against HIV, BVDV, and HBV.

The Henry reaction of (1R)-(1,4:3,6-dianhydro-D-mannitol-2-yl)-1,4:3,6-dianhydro-D-fructose 5,5'-dinitrate. Different reactive features of nitromethane to nitroethane.

Henry reactions of a novel higher sugar derivative, (1R)-(1,4:3,6-dianhydro-D-mannitol-2-yl)-1,4:3,6-dianhydro-D-fructose 5,5'-dinitrate (Alternate nomenclature: (1R)-(isomannid-2-yl)-1,4:3,6-dianhydro-D-fructose 5,5'-dinitrate), with nitromethane and nitroethane were studied. The kinetic and thermodynamic reactions with nitromethane under different conditions were carried out to afford (2S)- and (2R)-beta-nitroalcohols, respectively. But when using nitroethane the reaction gave a (2S)-beta-nitroalcohol with an inverted configuration at vicinal carbon C-1. Two stereogenic centers were generated, and one was altered in the reaction.

Synthesis of 2'-cyclohexenylnucleosides and corresponding CeNA building blocks.

A cyclohexene ring has similar structural properties and conformational behavior to a saturated five-membered furanose ring. In particular, it has good hydrolytic stability. Cyclohexenylnucleosides have been utilized in antiviral drug design, and some nucleosides and oligonucleotides based on the cyclohexene system have been developed. For further investigation of these modified nucleosides and oligonucleotides, synthesis of the chiral cyclohexenylnucleosides in high enantiomeric excess and in bulk quantities is necessary. This unit describes the complete synthesis of four enantiomerically pure 5'-hydroxy-4'-hydroxymethyl-2'-cyclohexenylnucleosides (thymine, cytosine, guanine, and adenine) and the four corresponding N-protected 4'-(monomethoxytrityl)oxymethyl cyclohexenyl nucleic acids (CeNA) building blocks. The chirality of these compounds is 1'S, 4'R, and 5'S.

1-(2,3-Di-O-acetyl-4-chloro-4-de-oxy-6-O-tosyl-ß-d-galactopyranos-yl)propan-2-one methanol 0.25-solvate.

The asymmetric unit of the title solvate, C(20)H(25)ClO(9)S·0.25CH(3)OH, contains one galactopyranosyl derivative and one-quarter of a methanol solvent mol-ecule. The galactopyran-ose ring is in the usual (4)C(1) conformation, and the anomeric center of the sugar has a ß configuration. The value of θ (3.44°) and the range of torsion angles [or 53.1 (5)-63.0 (5)°] reflect a slight distortion of the (4)C(1) pyran-ose ring. A minor orientational disorder affects a carbonyl group, which was modeled with two sites for the O atom having occupancies of 0.79 (5) and 0.21 (5). The crystal studied exhibited inversion twinning.

Synthesis of furanosyl alpha-C-glycosides derived from 4-chloro-4-deoxy-alpha-D-galactose and their cytotoxic activities.

Condensation of a new unnatural sugar 1 with 1,3-dicarbonyl compounds in the presence of anhydrous zinc chloride gave the polyhydroxyalkyl-furans in excellent yields. Further modification afforded the corresponding furanosyl alpha-C-glycoside derivatives. The absolute configuration of 3-acetyl-2-methyl-5-(2'-chloro-D-galacto-tetritol-1-yl)-furan was confirmed by single-crystal X-ray analysis. The in vitro cytotoxic activities of these furanosyl C-glycosides were also investigated.

An efficient synthesis of quinoxaline derivatives from 4-chloro-4-deoxy-alpha-D-galactose and their cytotoxic activities.

A novel and efficient method for the synthesis of quinoxaline derivatives has been developed. Isopropylidenation of 4-chloro-4-deoxy-alpha-D-galactose with 2,2-dimethoxypropane, followed by selective hydrolysis, afforded 2,3-O-isopropylidene-4-chloro-4-deoxy-D-galactose di-methyl acetal (3) as a sole product. Oxidation of compound 3 with (Bu3Sn)2O-Br2 gave corresponding hex-5-ulose derivative in high yields. The hex-5-ulose derivative reacted with o-phenylenediamines under neutral conditions to afford quinoxaline derivatives in reasonable yields. The in vitro cytotoxic activities of these quinoxaline derivatives were investigated.

Studies on the novel alpha-glucosidase inhibitory activity and structure-activity relationships for andrographolide analogues.

A series of analogues of andrographolide were synthesized and evaluated as novel alpha-glucosidase inhibitors. Among them compound 23, 15-p-methoxylbenzylidene 14-deoxy-11,12-didehydroandrographolide, was a potent inhibitor against alpha-glucosidase whose IC(50) value was 16microM. The structure-activity relationships were also discussed.

Highly stereoselective synthesis and structural characterization of new amino sugar derivatives.

2-C-Nitroalkyl-1,4:3,6-dianhydromannitols were synthesized via a Henry reaction of nitroalkyls with 1,4:3,6-dianhydrofructose. Catalytic hydrogenation then afforded the corresponding vicinal amino alcohols. Oximation of 1,4:3,6-dianhydrofructose with hydroxylamine, followed by hydrogenation, gave 2-amino-1,4:3,6-dianhydro-2-deoxymannitol. All compounds were elucidated by their HRMS, 1H NMR, 13C NMR, and IR spectra. The absolute configurations of the amino sugar derivatives were confirmed by single-crystal X-ray analysis or NOESY spectral studies. The possible mechanism for hydrogenation of the nitro 2-C-nitroalkyl sugar is proposed. The conformations of the fused furan rings of nitro and amino sugar derivatives are presented.

Chlorination of 3beta-hydroxyl-5-Delta steroids with anhydrous ferric chloride.

Treatment of 3beta-hydroxyl-5-Delta steroids with anhydrous FeCl(3) in CH(2)Cl(2) afforded reasonable yields of the corresponding alkyl chlorides with a retention of configurations. The structures of the chlorine-exchanging products were determined by NMR and HRMS spectra. The absolute configurations were confirmed by X-ray crystal analysis of 3beta-chloro-androst-5-en-17-one. The generality and scope of the reaction were also investigated.

Asymmetric synthesis of novel tetrahydroquinoline derivatives with a sugar building block and their bioactivities.

Some novel spiro-tetrahydroquinolines were stereoselectively synthesized by using keto-sugar derived from sucrose as a building block in one pot under mild conditions. The in vitro immunobiological activity and cytotoxicity of these novel tetrahydroquinolines were investigated. The results implied that these spiro-compounds have obvious bioactivity and may be structurally modified to improve bioactivity further.

Preparation of alpha and beta anomers of 1,2,3,6-tetra-O-acetyl-4-chloro-4-deoxy-D-galactopyranose based upon anomerization and kinetic acetylation.

4-Chloro-4-deoxy-alpha-d-galactopyranose, 1,2,3,6-tetra-O-acetyl-4-chloro-4-deoxy-alpha-d-galactopyranose and 1,2,3,6-tetra-O-acetyl-4-chloro-4-deoxy-beta-d-galactopyranose were readily prepared from 1,4:3,6-dianhydro-beta-d-fructofuranosyl 4-chloro-4-deoxy-alpha-d-galactopyranoside. In the study, we found an interesting anomerization phenomenon of 4-chloro-4-deoxy-d-galactose. The molar ratio of alpha and beta anomers in solution is about 1:2 when the anomerization reaches a dynamic equilibrium, and the beta anomer could completely convert to the alpha anomer in the process of crystallization and precipitation. The acetylation of 4-chloro-4-deoxy-d-galactopyranose is kinetically controlled, and the configuration of the starting galactose determines the configuration of the resulting acetates. The influence of the chloro group at C-4 and the O-acetyl group at the anomeric carbon on the galactopyranose ring conformations is discussed, based upon the crystallographic data for the alpha and beta anomers of 1,2,3,6-tetra-O-acetyl-4-chloro-4-deoxy-d-galactopyranose.

A facile approach to anhydrogalactosucrose derivatives from chlorinated sucrose.

Three new anhydrosucrose derivatives: 1,4:3,6-dianhydro-beta-D-fructofuranosyl 4-chloro-4-deoxy-alpha-D-galactopyranoside (4), 1,4:3,6-dianhydro-beta-D-fructofuranosyl 3,6-anhydro-4-chloro-4-deoxy-alpha-D-galactopyranoside (6) and 1,6-dichloro-1,6-dideoxy-beta-D-fructofuranosyl-3,6-anhydro-4-chloro-4-deoxy-alpha-D-galactopyranoside (8) were prepared from chlorinated sucrose. The structures of these anhydrides were confirmed by their (1)H and (13)C NMR spectra, ESIMS and elemental analysis. The crystal structures of 6 and the acetate of 4 (5) are presented. The relative reactivity of the chloromethyl groups towards S(N)2 reactions in 1,6-dichloro-1,6-dideoxy-beta-d-fructofuranosyl 4,6-dichloro-4,6-dideoxy-alpha-D-galactopyranoside was found to be in order 6>6'>1'.