Novel cyclosal nucleotides with reduced inhibitory potency toward human butyrylcholinesterase

Two novel cycloSal-d4T monophosphates (d4TMPs) with increased steric demand have been synthesized via a new synthetic route. While 3-cyclohexyl-cycloSal d4TMP did not show a significantly reduced inhibitory potency toward human butyrylcholinesterase, the opposite was the case for the second novel pronucleotide, bis-(cycloSal-d4TMP).     

Effects of cycloSal-d4TMP derivatives in H9 cells with induced AZT resistance phenotype.

Cytotoxic and antiretroviral activity of cycloSal-d4TMP derivatives were tested in a new AZT-resistant H9 cell subline (H9rAZT250). The results showed, that cycloSal-d4TMP derivatives overcame resistance of HIV-1 to d4T in H9rAZT250 cells, which exert decreased thymidine kinase (TK) gene expression.     

d4TMP delivery from 7-substituted cycloSal-d4TMPs

Benzyl-substituted cycloSal-d4T monophosphates were prepared and evaluated for their ability to release d4TMP selectively. In contrast to previously reported derivatives, two of the new compounds release d4TMP as the sole product while two others gave the expected benzyl phosphate diesters. However, these diesters were surprisingly stable against degradation to release d4TMP.     

Benzyl-functionalized cycloSal-d4T monophosphates

Synthetic routes to benzyl-functionalized cycloSal-d4T monophosphates (7CH2X-cycloSal-d4TMP) have been developed. Their hydrolytic behavior in basic aqueous solution (pH = 7.3) was studied and their hydrolysis half-lives were determined. It turned out that two different degradation pathways are leading to different products: beside the formation of the expected d4TMP and a styrene type derivative, a phenyl-d4T-phosphodiester was obtained as well. The product distribution was specified.     

New developments of the "lock-in" modified cycloSal-d4TMPs

New developments of the "lock-in" modified cycloSal-d4TMP are reported. Novel prodrugs with variations in the linker chain were introduced. The synthesis, hydrolysis properties in different media (PBS, CEM/0- and liver extracts) and the antiviral activities against HIV are shown.     

Study of different substituted cyclic and acyclic benzylpronucleotides of d4T relative to their hydrolytic stability and antiviral activity

CycloSal-d4TMP and two different bis(benzyl) phosphate triesters of the antivirally active nucleoside analog d4T were studied with regard to their chemical hydrolysis behavior at pH 7.3, in CEM/0 cell extracts, and their anti-HIV activity. In contrast to triesters 2-4, bis-(o-AB)-d4TMP 1 was found to be chemically exquisitely stable. All compounds led to the formation of d4TMP in cell extracts and all triesters achieved the TK-bypass.     

d4TMP Delivery from 7-Substituted cycloSal-d4TMPs

Abstract

Benzyl-substituted cycloSal-d4T monophosphates were prepared and evaluated for their ability to release d4TMP selectively. In contrast to previously reported derivatives, two of the new compounds release d4TMP as the sole product while two others gave the expected benzyl phosphate diesters. However, these diesters were surprisingly stable against degradation to release d4TMP.     

Cyclo-saligenyl-2′,3′-dideoxy-2′,3′-didehydroythymidinemonophosphate (cycloSal-d4TMP) — A New Pro-Nucleotide Approach

Abstract

The synthesis of cycloSal-d4TMP 3a-g as new pro-nucleotide approach for d4TMP 2 is described. Phosphotriesters 3 release the d4TMP 2 selectively by a controlled, chemically induced tandem reaction. CycloSal-phosphotriesters 3 exhibited high biological activity against HIV-1/HIV-2 in CEM cells which was completely retained in CEM TK^? cells.     

Study of Different Substituted Cyclic and Acyclic Benzylpronucleotides of d4T Relative to Their Hydrolytic Stability and Antiviral Activity

Abstract

CycloSal-d4TMP and two different bis(benzyl) phosphate triesters of the antivirally active nucleoside analog d4T were studied with regard to their chemical hydrolysis behavior at pH 7.3, in CEM/0 cell extracts, and their anti-HIV activity. In contrast to triesters 2–4, bis-(o-AB)-d4TMP 1 was found to be chemically exquisitely stable. All compounds led to the formation of d4TMP in cell extracts and all triesters achieved the TK-bypass.     

A new cyclic phosphoramidate D4T prodrug approach cycloAmb-D4T-phosphoramidates.

A new potential phosphoramidate prodrug approach for d4T 1 is described. In hydrolyses studies the cycloAmb-d4T-phosphoramidates 2 and 3 proved to deliver d4TMP following a tandem reaction.     

BENZYL-FUNCTIONALIZED cycloSal-d4T MONOPHOSPHATES

Synthetic routes to benzyl-functionalized cycloSal-d4T monophosphates (7CH₂X-cycloSal-d4TMP) have been developed. Their hydrolytic behavior in basic aqueous solution (pH = 7.3) was studied and their hydrolysis half-lives were determined. It turned out that two different degradation pathways are leading to different products: beside the formation of the expected d4TMP and a styrene type derivative, a phenyl-d4T-phosphodiester was obtained as well. The product distribution was specified.     

A th-1 Mutant of Arabidopsis thaliana Is Defective for a Thiamin-Phosphate-Synthesizing Enzyme: Thiamin Phosphate Pyrophosphorylase

We have examined the activity of the thiamin phosphate pyrophosphorylase in Arabidopsis thaliana wild type and in a mutant (th-1) which requires exogenous thiamin for growth. Mutant and wild-type plants grown in 1 × 10⁻⁷ molar thiamin were used for the examination of the production of thiamin and thiamin monophosphate (TMP) using 4-methyl-5-hydroxyethylthiazole phosphate and 2-methyl-4-amino-5-hydroxymethylpyrimidine pyrophosphate as substrates. While the wild-type strain formed both thiamin and TMP, the th-1 mutant did not. When TMP was added to the extracts, the th-1 mutant, as well as wild type, produced thiamin. Accordingly, it was concluded that the th-1 mutant was defective in the activity of TMP pyrophosphorylase. Some of the characteristics of the enzyme from the wild-type plant were examined. The optimum temperature for the reaction is 45°C, and the K_m values for the substrates are 2.7 × 10⁻⁶ molar for 4-methyl-5-hydroxyethylthiazole phosphate and 1.8 × 10⁻⁶ molar for 2-methyl-4-amino-5-hydroxymethylpyrimidine pyrophosphate.     

Effects of cycloSal-D4TMP Derivatives in H9 Cells with Induced AZT Resistance Phenotype

Abstract

Cytotoxic and antiretroviral activity of cycloSal-d4TMP derivatives were tested in a new AZT-resistant H9 cell subline (H9^rAZT²⁵⁰). The results showed, that cycloSal-d4TMP derivatives overcame resistance of HIV-1 to d4T in H9^rAZT²⁵⁰ cells, which exert decreased thymidine kinase (TK) gene expression.     

Structure-based design of new DHFR-based antibacterial agents: 7-aryl-2,4-diaminoquinazolines

Dihydrofolate reductase (DHFR) inhibitors such as trimethoprim (TMP) have long played a significant role in the treatment of bacterial infections. Not surprisingly, after decades of use there is now bacterial resistance to TMP and therefore a need to develop novel antibacterial agents with expanded spectrum including these resistant strains. In this study, we investigated the optimization of 2,4-diamnoquinazolines for antibacterial potency and selectivity. Using structure-based drug design, several 7-aryl-2,4-diaminoquinazolines were discovered that have excellent sub-100 picomolar potency against bacterial DHFR. These compounds have good antibacterial activity especially on gram-positive pathogens including TMP-resistant strains.     

PTSD大鼠中缝背核神经元TMP酶表达变化的实验研究

目的研究创伤后应激障碍大鼠中缝背核神经元细胞TMP(三偏磷酸酶)活性分布及其表达变化。方法采用SPS刺激方法,建立PTSD样大鼠SPS模型,随机分为SPS刺激后1d、7d、14d和正常对照组,应用光、电镜酶组化技术方法,分别对各组中缝背核神经元细胞TMP活性分布及其变化进行观察和定量检测。结果光镜下TMP酶反应阳性产物为棕褐色颗粒分布于细胞质中;电镜下TMP酶反应阳性产物为高电子密度的黑色颗粒沉淀,分布于各组中缝背核神经元细胞内溶酶体上。SPS刺激后1d、7d、14d中缝背核神经元TMP活性表达比正常对照组明显增强,并于7d达到高峰。结论创伤后应激障碍大鼠中缝背核神经元细胞TMP活性增强,提示TMP参与了中缝背核神经元细胞凋亡产物的降解和处理过程。     

The influence of potassium 4-toluenethiosulfonate on membrane potential and ATPase activity of plasmatic membranes of loach embryos

The influence of novel biologically active substance potassium 4-toluenethiosulfonate in concentration of 4 x 10(-5) M on the changes of electrophysiological parameters of embryonic cells in early development of fish (Misgurnus fossilis L.) was investigated including the changes of membrane potential (TMP) and enzyme activity of plasmatic membranes of loach embryos during the period of synchronous division of blastomers in the early period of development. The evaluation of influence of these matters was studied and aperiodic changes of the level of TMP were shown. The diminishing of amplitude in every period by 7/12 mV and diminishing of growth of maximal values of vibrations of TMP by 39 mV in comparison to control was noticed. It was related to inhibition of some biosynthetic processes and results in the decline of activity of membrane enzyme (Na+, K(+)-ATPase) by 75.5 +/- 4.1% and 78.4 +/- 10.4% both at the action of high (4 x 10(-3) M) and low (4 x 10(-9) M) concentrations, accordingly, at first hours of development with subsequent renewal of its activity to the level of control only for the actions of low concentration.     

铈代替铅电镜酶组织化学显示培养脊神经节神经元TMP活性

目的探讨铈法显示脊神经节神经元初代培养细胞的TMP(三偏磷酸酶)活性。方法应用电镜酶组化技术,以铈作为捕捉剂代替常规以铅作为捕捉剂来检测观察脊神经节初代培养的神经元内溶酶体的TMP活性。结果脊神经节神经元初代培养细胞内存在TMP阳性的溶酶体,其TMP阳性反应颗粒较常规以铅作为捕捉剂来显示溶酶体不仅孵育液充分溶解呈清亮透明,电镜下反应产物颗粒也微细均匀,而且避免了反应产物的扩散及非特异性沉淀。结论以铈作为捕捉剂来检测溶酶体的TMP活性用于电镜下观察,其方法稳定、易行,是一种能很好显示溶酶体标记酶的电镜酶组化技术方法。     

A New Cyclic Phosphoramidate D4T Prodrug Approach cycloAmb-D4T-Phosphoramidates

Abstract

A new potential phosphoramidate prodrug approach for d4T 1 is described. In hydrolyses studies the cycloAmb-d4T-phosphoramidates 2 and 3 proved to deliver d4TMP following a tandem reaction.     

川芎嗪抗链霉素耳毒性作用及其对豚鼠耳蜗外毛细胞钾通道的影响

本文旨在研究川芎嗪（tetramethylpyrazine,TMP）拮抗链霉素耳毒性作用及其对豚鼠耳蜗外毛细胞K^＋通道的影响,探讨TMP拈抗链霉素耳毒性的离子通道机制。60只豚鼠随机分为6组,应用听觉脑干反应（auditory brainstem response,ABR）技术检测豚鼠ABR听阈,观测TMP的抗链霉素耳毒作用;并采用全细胞膜片钳技术观察TMP对耳蜗外毛细胞Ca^2＋敏感艮电流的影响。结果显示,TMP明显降低链霉素导致的豚鼠ABR听阈升高,提示TMP具有抗链霉素耳毒性作用;TMP能明显增大豚鼠耳蜗外毛细胞Ca^2＋敏感艮电流,并呈浓度依赖关系。结果提示,TMP通过增大艮通道电导而拮抗链霉素耳毒性作用。