Antiheparin properties of synthetic polycations]

The toxicity of differently structured polymeric compounds for mice (intravenously) and their antiheparin properties in in vitro and in vivo tests were studied. The antiheparin activity display polymers containing amino groups or onium atoms of nitrogen, sulphur and phosphorus in various structures. The antagonism to heparin is due to the polymers being endowed with polycationic properties.     

Antiheparin activity of polymers based on the alkaloid, lupinine]

Antiheparin activity and acute toxicity of a monomer and polymer quaternary ammonium salts obtained on the basis of the alkaloid lupinine were investigated. The monomer was shown to have no antiheparin activity even when administered in high doses. Lupinine polymetakryloyl iodoethylate is a selective heparin antagonist. This activity of polymers is relative to the extent of polymerization. A compound with a molecular weight of 20 000 turned out the most effective in the tested series of ready-made polymers. Acute toxicity of polycations is considerably less than that elicited by the monomer. The effect of heparin complete neutralization is accompanied by transitory thrombocytopenia.     

The effect of polycations on the activity of pepsin

A study has been made of the interaction of selected polycations with pepsin A (E.C. 3.4.23.1). Protamine, polybrene, spermine, spermidine and poly(L-lysine) all acted as inhibitors of the enzyme at low concentrations, but at higher concentrations of the polycations the inhibition was less pronounced. A more detailed study of the anomalous inhibition was made using protamine, polybrene and poly(L-lysine) and it was shown experimentally that, when used by themselves, each of these polycations acted as a weak proteolytic catalyst. The order of catalytic effectiveness was: protamine greater than polybrene much greater than poly(L-lysine). Thus, it is now possible to explain why the observed inhibition of pepsin decreases when the concentration of the inhibiting polycation is increased.     

Comparison of the activity of polyanions and polycations on the classical and alternative pathways of complement

Polyanions and polycations inhibit activity of the alternative and classical pathways of complement. We compared polyanions (commercial porcine heparin, chondroitin sulfate A, chondroitin sulfate B (dermatan sulfate), chondroitin sulfate C and heparatin sulfate) with polycations (salmon sperm protamine sulfate, poly-L-lysine, poly-L-arginine, polybrene and a synthetically prepared portion of platelet factor 4) for ability to inhibit alternative and classical pathway activity. The polyanions had considerably more activity on the alternative than on the classical pathway, whereas the polycations more profoundly inhibited classical than alternative pathway activity. For example, heparin, a polyanion, at 1.0 micrograms (7.7 x 10(-7) M based upon an Mr average of 13000)/10(7) cellular intermediates, inhibited alternative pathway activity and classical pathway activity by 77 and 14%, respectively, whereas protamine sulfate, a polycation, at 0.25 micrograms/10(7) cellular intermediates, inhibited these two pathways by 34 and 98%, respectively. These studies suggest that the capacity to inhibit complement activity is a common feature of highly charged substances and the polyanions preferentially inhibit the alternative pathway while polycations preferentially inhibit the classical pathway. In vivo these highly charged substances could play an important role in the tissues in regulating the activity of both pathways of complement.     

基于假病毒技术探讨黄芩苷抗HIV-1活性研究

目的:基于假病毒技术探究黄芩苷抗HIV-1活性作用.方法:利用HIV-1骨架质粒NL4-3 mCherry Luciferase、HIV-1包膜蛋白质粒pCMV-VSV-G和人胚肾细胞HEK-293T构建HIV-1假病毒药物筛选体系,并对该体系进行优化和安全性验证.测定黄芩苷对293T细胞活性、HIV-1假病毒活性、H...     

The interaction of papain with polycations

An investigation has been made of the interactions of the enzyme papain with the polycations protamine, polybrene, poly(L-lysine), spermine, spermidine and the neutral polymer polyvinylpyrrolidone (PVP). At low concentrations, each behaves as an inhibitor of the enzyme. As their concentrations increased above a certain level, the activity of the systems increased, and their inhibition of the enzyme appeared to be less pronounced. When acting by themselves in the presence of the substrate haemoglobin, each of the polycations was a weak proteolytic catalyst with a ranking of catalytic effectiveness of protamine greater than polybrene greater than poly(L-lysine) greater than polyvinyl-pyrrolidone greater than spermidine greater than spermine. This effect could explain the anomalous inhibition of papain by these polycations. The interaction of papain with dansyl protamine (DNSP) and the extent of complex formation were studied using a fluorescence polarization technique and the results showed that there was a strong interaction occurred. The strength of binding was assessed by determination of the critical electrolyte concentration (0.2 M, NaNO3). The stoichiometry of the DNSP-papain complex was found to be 63 base moles of DNSP to one mole of papain.     

Antiheparin and toxic properties of quaternary ammonium salts of the monodispersed oligomers of conidine

A distinct relationship has been demonstrated of antiheparin and toxic properties of quaternary ammonium salts of monodispersed oligomers of conidine to their molecular weight and, therefore, to the length of the macromolecular chain. Conidine monomer having the highest acute toxicity does not interact with heparin. As the number of functional links is increased up to 25, antiheparin activity of the compounds under study ascends, whereas toxic effects become less powerful. An oligomer with 25 functional links has appeared to be the best combination of the characteristics under consideration. Further increase of the chain length and, therefore, of the molecular weight is followed by a slight lowering of anticoagulant activity and increase of acute toxicity.     

Improving oral drug bioavailability with polycations?

The administration of drugs via the oral route is challenging due to the (bio)chemical aggressivity of the digestive system and to the presence of barriers that hinder cell uptake and access to the bloodstream. Indeed, the gastrointestinal tract is characterized by large variations of pH, the presence of enzymes and surfactants, and by absorption barriers such as mucus and the epithelium. Thus, many compounds such as proteins and nucleic acids do not reach the systemic circulation due to their premature degradation and/or large size. Among the different strategies that have been investigated to address these challenges, polycations have been explored to improve the oral absorption of many types of drugs. Because of their multiple positive charges and repetitive structure, polycations can protect sensitive drugs against rapid degradation and interact with the gastrointestinal mucosa. Moreover, cationic polymers promote drug transfer across intestinal barriers through various mechanisms, including the opening of the tight junctions, and change in uptake pathway. This contribution provides an overview of the most common polycations currently investigated as absorption enhancers for the oral route, and discusses the manner in which they are employed (co-administration, micro- and nanoparticles, conjugation) to improve the oral drug delivery of different classes of therapeutics.     

Inhibition of lymphocyte DNA-synthetic responses by spermine-derived polycations

The polyamines putrescine, spermidine, and spermine are oxidized by the enzyme diamine oxidase to form the corresponding aldehyde derivatives. These aldehydes have been shown to undergo a variety of spontaneous reactions, some of which result in polycationic addition compounds. We have chemically synthesized some spermine-derived polycations by reaction with the dialdehyde glutaraldehyde followed by reduction of the resulting Schiff base with sodium borohydride. Their migration on ion exchange and gel filtration columns was consistent with the formation of polycations with properties similar to those reported for the spontaneous reaction products. When added to cultures of alloantigen or mitogen stimulated lymphocytes, these polycations were potent inhibitors of the incorporation of tritiated thymidine and blast cell formation. This inhibition was reversible, non-cytotoxic, and only apparent if the polycation was added early in the culture period. The concentration of polycation necessary to achieve 50% inhibition of the lymphocyte response decreased as the cationic nature relative to spermine increased.     

基于双荧光素酶报告基因系统的人源SREBP1基因启动子的活性研究

目的 构建固醇调节元件结合蛋白 1（SREBP1）启动子双荧光素酶报告基因表达载体,为研究针对 SREBP1靶点的天然活性抑制剂筛选奠定基础。方法 利用生物信息学软件,对SREBP1基因5′端上游5 000 bp序 列进行启动子预测与分析。应用 Gibson Assembly 方法构建启动子双荧光素酶载体,并将构建成功的 pGL3- SREBP1-pro重组质粒和内参质粒pRL-SV40共转染肝癌HepG2细胞并给予天然抑制剂大黄素,检测SREBP1转录活 性的抑制效果。结果 SREBP1基因5′端上游2 000 bp区域内有启动子特征序列,存在转录因子结合位点;重组质粒 经酶切及测序鉴定证实为阳性克隆,瞬时转染肝癌HepG2细胞后经双荧光素酶报告基因系统检测,所克隆的片段序 列具有启动子活性,该活性可被大黄素所抑制。结论 成功构建了pGL3-SREBP1-pro双荧光素酶报告基因表达载 体,并证实其活性,可为进一步用于针对SREBP1靶点的天然活性抑制剂筛选奠定     

FFA1-selective agonistic activity based on docking simulation using FFA1 and GPR120 homology models

BACKGROUND AND PURPOSE

The free fatty acid FFA1 receptor and GPR120 are GPCRs whose endogenous ligands are medium- and long-chain FFAs, and they are important in regulating insulin and GLP-1 secretion respectively. Given that the ligands of FFA1 receptor and GPR120 have similar properties, selective pharmacological tools are required to study their functions further.

EXPERIMENTAL APPROACH

We used a docking simulation approach using homology models for each receptor. Biological activity was assessed by phosphorylation of ERK and elevation of intracellular calcium ([Ca²⁺]_i) in cells transfected with FFA1 receptor or GPR120. Insulin secretion from murine pancreatic beta cells (MIN6) was also measured.

KEY RESULTS

Calculated hydrogen bonding energies between a series of synthetic carboxylic acid compounds and the homology models of the FFA1 receptor and GPR120, using docking simulations, correlated well with the effects of the compounds on ERK phosphorylation in transfected cells (R²= 0.65 for FFA1 receptor and 0.76 for GPR120). NCG75, the compound with the highest predicted selectivity for FFA1 receptors from this structure-activity relationship analysis, activated ERK and increased [Ca²⁺]_i as potently as the known FFA1 receptor-selective agonist, Compound 1. Site-directed mutagenesis analysis based on the docking simulation showed that different amino acid residues were important for the recognition and activation by FFA1 receptor agonists. Moreover, NCG75 strongly induced ERK and [Ca²⁺]_i responses, and promoted insulin secretion from MIN6 cells, which express endogenous FFA1 receptors.

CONCLUSION AND IMPLICATIONS

A docking simulation approach using FFA1 receptor and GPR120 homology models could be useful in predicting FFA1 receptor-selective agonists.     

Synthesis and biological activity of potent HIV-1 protease inhibitors based on Phe-Pro dihydroxyethylene isosteres

Peptidomimetic inhibitors of HIV-1 PR are still a key resource in the fight against AIDS. Here we describe the synthesis and biological activity of HIV-1 PR inhibitors based on four novel dihydroxyethylene isosteres of the Phe-Pro and Pro-Pro dipeptides. The isosteres, containing four stereogenic centers, were synthesized in high yield and excellent stereoselectivity via the cyclization of epoxy amines derived from α-amino acids. The inhibitors were assembled by coupling the isosteres with suitable flanking groups and were screened against recombinant HIV PR showing activities in the subnanomolar to micromolar range. Two Phe-Pro-based inhibitors active at the nanomolar level were further investigated: both inhibitors combine the ability to suppress HIV-1 replication in infected MT-2 cells with low cytotoxicity against the same cells, thereby displaying a high therapeutic index. These results demonstrate the potential of the new Phe-Pro dihydroxyethylene isostere as a core unit of powerful HIV-1 PR inhibitors.     

Physical activity and physical activity adherence in the elderly based on smoking status

This study assessed the impact of current smoking status and lifetime smoking status on physical fitness and physical activity regimen adherence as part of a larger study on walking for exercise in elderly primary care patients at a Veterans Affairs Medical Center. At baseline, 218 participants self-reported smoking status which was verified by carbon monoxide expiration. Former and current smokers responded to questions about length of time quit, average daily cigarette intake, and years a smoker. Smoking measures were re-collected at 6- and 12-month follow-ups if the participants indicated a change in smoking status. Veterans completed multiple measures of physical activity (e.g., 6-min walk, 7-day Physical Activity Recall), and adherence to a physical activity goal was assessed. The Physical Component Summary (PCS) subscale of the Medical Outcomes Study Short Form-36 (MOS SF-36) was used to assess health-related quality of life. Hierarchical regression models indicated smoking status was a predictor of the baseline 6-min walk such that smokers walked significantly shorter distances than nonsmokers. In addition, smoking status was found to be a significant predictor of adherence; however, the overall model that included smoking status as a predictor did not demonstrate a significant effect on adherence. Neither smoking status nor pack years were predictors of baseline self-reported physical activity or changes in physical activity post intervention. Results are consistent with recommendations to use physical exercise as an aid to tobacco cessation, even in aging men with extensive smoking histories.