[B10,22-Asp,B25-Tyr-NH2]-去β链C端五肽胰岛素的合成和性质

本文报道了[B10,22-Asp,B25-Tyr-NH2]-去B链羧端五肽胰岛素的制备及其生物活性。结果表明,这一类似物的生物活力比去五肽胰岛素(DPI)的活力高一倍,但却比Gerald所报道的[B10-Asp,B25-Tyr-NH_2]-DPI的活力低很多,说明后者的高活性可能依赖于分子中B22-Arg的存在。     

Ring A structural modified derivatives of withaferin A and the evaluation of their cytotoxic potential

Regio-/stereoselective Michael addition to ring A of withaferin-A was performed using an optimized reaction procedure to synthesise a library of 2,3-dihydro,3-β-substituted withaferin-A derivatives. The analogues thus obtained were evaluated for in vitro cytotoxicity against various human cancer cell lines. 3-Azido analogue exhibited 35-fold increase (IC(50)=0.02-1.9 μM) in cytotoxicity against almost the entire cell lines tested when compared to the parent molecule. However, further modifications of 3-azido analogue with various alkynes under Husigen's cycloaddition conditions generated a variety of triazole derivatives with reduced cytotoxicity.     

DPP IV resistance and insulin releasing activity of a novel di-substituted analogue of glucose-dependent insulinotropic polypeptide, (Ser2-Asp13)GIP

Structure-function studies suggest that preservation of the N-terminus and secondary structure of glucose-dependent insulinotropic polypeptide (GIP) is important for biological activity. Therefore, a novel di-substituted analogue of GIP, (Ser(2)-Asp(13))GIP, containing a negatively charged Asp residue in place of an Ala in position 13, was synthesised and evaluated for in vitro biological activity. Incubation with dipeptidyl peptidase IV (DPP IV) showed the half-lives of GIP and (Ser(2)-Asp(13))GIP to be 2.3 and >4h, respectively. Insulin releasing studies in clonal pancreatic BRIN-BD11 cells demonstrated that (Ser(2)-Asp(13))GIP (10(-12)to 10(-7)mol/l) was significantly less potent (60-90%; P<0.05 to P<0.001) than native GIP. The peptide failed to display antagonistic properties as it did not significantly alter insulin secretion when incubated in the presence of GIP (10(-7)mol/l). These results demonstrate that despite increased resistance to DPP IV, substituting Ala in position 13 with a negatively charged Asp, thus producing the di-substituted analogue (Ser(2)-Asp(13))GIP, significantly reduces biological activity, most likely due to modifications within the secondary structure.     

Subtype selective NMDA receptor antagonists: evaluation of some novel alkynyl analogues

A benzylpiperidine analogue with an acetylenic linker, 5-(3-[4-(4-fluorobenzyl)-piperidin-1-yl]-prop-1-ynyl)-1,3-dihydrobenzimidazol-2-one (3), was identified as a chemical lead with excellent activity at the NR1A/2B receptor (IC50=3 nM). Efforts to optimize this activity led to focused modifications around the structural motif of 3. The synthesis and SAR studies are discussed.     

Antagonists of oxytocin featuring replacement with modified beta-mercaptopropionic acids at position 1.

Twenty analogues were synthesized of [Pmp1, D-Trp2, Arg8]oxytocin, PA, (Pmp = beta,beta-pentamethylene-beta-mercaptopropionic acid), a potent antagonist of the uterotonic effect of oxytocin in the rat (uterotonic test in vitro, pA2 = 7.77) and in the baboon. Systematic substitution of Pmp1 was made with beta-mercaptopropionic acids featuring replacement of the 4-methylene group of the cyclohexyl ring of Pmp with isosteric O, S, NH or with C=O. Since the more hydrophilic NH and C=O substitutions showed a sharply decreased antagonistic potency (rat uterotonic test in vitro), additional modifications were made to reduce their hydrophilicity. Acylation of the NH group with various acyl groups, and ketalization or thioketalization of C=O with more or less bulky substituents led to a partial restoration of potency, the N-carbamyl- and the 2-mercapto-2-adamantaneacetyl analogues being equipotent with PA. Internal cyclization by amidation of the NH-group with Gly-9, resulted in a bicyclic analogue, (cyclo 1-9)[(HN)Pmp1, Gly9]PA which was equipotent with PA. When Pen-6 was introduced into the bicyclic derivative instead of Cys-6, to reduce the flexibility of the rings, the resulting (cyclo 1-9)[(HN)Pmp1, Pen6, Gly9]PA had somewhat better potency (pA2 = 8.17) in the uterotonic test and no detectable activity in the antidiuretic assay. In the case of substitution of PA with beta,beta-(3-thiapentamethylene)-beta-mercaptopropionic acid, (S)Pmp, there was also an increase in inhibitory potency in the uterotonic test (pA2 = 8.08): the analogue had extremely weak antidiuretic activity. To establish the importance of the steric effects of the Pen-6 substitution, analogues [Pen6]PA and [(S)Pmp1, Pen6]PA were made and found to be very potent, with a pA2 of 8.72 and 8.86, respectively. The high potency of the latter analogue and its extremely weak action in the diuretic assay makes it an attractive candidate for studies on the inhibition of the biological effects of oxytocin and for the prevention of preterm labour.     

Potato micro-tuber inducing hydroxylasiodiplodins from Lasiodiplodia theobromae

Three hydroxylasiodiplodins were isolated from the mycelium extracts of Lasiodiplodia theobromae IFO 31059; and their structures were identified as (3R),(4S)-4-hydroxylasiodiplodin, (3R),(6R)-6-hydroxy-de-O-methyllasiodiplodin and (3R),(5R)-5-hydroxy-de-O-methyllasiodiplodin. All showed potato micro-tuber inducing activity.     

GABA-Mimetic Activity and Effects on Diazepam Binding of Aminosulphonic Acids Structurally Related to Piperidine-4-Sulphonic Acid

The relationship between structure, in vivo activity, and in vitro activity of some analogues of the gamma-aminobutyric acid (GABA) agonist piperidine-4-sulphonic acid (P4S) was studied. The syntheses of 1,2,3,6-tetrahydropyridine-4-sulphonic acid (DH-P4S) and (RS)-pyrrolidin-3-yl-methanesulphonamide (PMSA-amide) are described. Like P4S, its unsaturated analogue DH-P4S and the five-ring isomer (RS)-pyrrolidin-3-yl-methanesulphonic acid (PMSA) were bicuculline methochloride (BMC)-sensitive inhibitors of the firing of neurones in the cat spinal cord. Whereas isonipecotic acid was less potent than its unsaturated analogue isoguvacine as a GABA-mimetic and as an inhibitor of GABA binding, the opposite relative potencies of P4S and DH-P4S were observed, P4S being proportionally more potent than DH-P4S. In contrast with P4S and DH-P4S, PMSA, which is an analogue of the potent GABA uptake inhibitor and BMC-sensitive GABA-mimetic homo-beta-proline, was a relatively weak inhibitor of GABA uptake in vitro. PMSA-amide was more than two orders of magnitude weaker than PMSA as an inhibitor of GABA binding and did not significantly affect GABA uptake in vitro. The effects of 3-aminopropanesulphonic acid (3-APS), PMSA, P4S, and DH-P4S on the binding of [3H]diazepam in vitro at 30 degrees C, in the presence or absence of chloride ions, were studied and compared with those of the structurally related amino acids GABA, homo-beta-proline, isonipecotic acid, and isoguvacine. Under these conditions the aminosulphonic acids were weaker than the respective amino acids in enhancing [3H]diazepam binding, the difference being more pronounced in the absence of chloride.     

Modification of near active site residues in organophosphorus hydrolase reduces metal stoichiometry and alters substrate specificity

Organophosphorus hydrolase (OPH, EC 8.1.3.1) is a dimeric, bacterial enzyme that detoxifies many organophosphorus neurotoxins by hydrolyzing a variety of phosphonate bonds. The histidinyl residues at amino acid positions 254 and 257 are located near the bimetallic active site present in each monomer. It has been proposed that these residues influence catalysis by interacting with active site residues and the substrate in the binding pocket. We replaced the histidine at position 254 with arginine (H254R) and the one at position 257 with leucine (H257L) independently to form the single-site-modified enzymes. The double modification was also constructed to incorporate both changes (H254R/H257L). Although native OPH has two metals at each active site (four per dimer), all three of these altered enzymes possessed only two metals per dimer while retaining considerable enzymatic activity for the preferred phosphotriester (P-O bond) substrate, paraoxon (5-100% kcat). The three altered enzymes achieved a 2-30-fold increase in substrate specificity (kcat/Km) for demeton S (P-S bond), an analogue for the chemical warfare agent VX. In contrast, the substrate specificity for diisopropyl fluorophosphonate (P-F bond) was substantially decreased for each of these enzymes. In addition, H257L and H254R/H257L showed an 11- and 18-fold increase, respectively, in specificity for NPPMP, the analogue for the chemical warfare agent soman. These results demonstrate the ability to significantly enhance the specificity of OPH for various substrates by site-specific modifications, and it is suggested that changes in metal requirements may affect these improved catalytic characteristics by enhancing structural flexibility and improving access of larger substrates to the active site, while simultaneously decreasing the catalytic efficiency for smaller substrates.     

Antitumor agents 283. Further elaboration of desmosdumotin C analogs as potent antitumor agents: activation of spindle assembly checkpoint as possible mode of action

In our ongoing study of the desmosdumotin C (1) series, twelve new analogues, 21-32, mainly with structural modifications in ring-A, were prepared and evaluated for in vitro antiproliferative activity against several human tumor cell lines. Among them, the 4'-iodo-3,3,5-tripropyl-4-methoxy analogue (31) showed significant antiproliferative activity against multiple human tumor cell lines with ED(50) values of 1.1-2.8 μM. Elongation of the C-3 and C-5 carbon chains reduced activity relative to propyl substituted analogues; however, activity was still better than that of natural compound 1. Among analogues with various ether groups on C-4, compounds with methyl (2) and propyl (26) ethers inhibited cell growth of multiple tumor cells lines, while 28 with an isobutyl ether showed selective antiproliferative activity against lung cancer A549 cells (ED(50) 1.7 μM). The gene expression profiles showed that 3 may modulate the spindle assembly checkpoint (SAC) and chromosome separation, and thus, arrest cells at the G2/M-phase.     

Critical modifications of the ISO-1 scaffold improve its potent inhibition of macrophage migration inhibitory factor (MIF) tautomerase activity

Based on the scaffold of (S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1), an inhibitor of the proinflammatory cytokine MIF, two critical modifications and chiral resolution have significantly improved the potency of the inhibition. Compound (R)-17 is 20-fold more potent than ISO-1 and inhibits MIF tautomerase activity with an IC(50) of 550 nM.     

Antimalarial benzo[c]phenanthridines

Analogues of the antimalarial alkaloid nitidine have been prepared with high potency against both chloroquine-sensitive and -resistant strains of Plasmodium falciparum in vitro. Simple modifications, using an established synthetic route, resulted in an analogue with IC(50) below 5ng/mL against a chloroquine-sensitive strain of P. falciparum. N-Ethylethoxidine had IC(50) below 30ng/mL against both chloroquine-sensitive and chloroquine-resistant strains of P. falciparum.     

虎眼万年青属植物Ornithogalum thyrsoide 的甾体皂苷及黄酮类化学成分

运用硅胶、Sephadex LH-20、ODS柱层析及半制备高效液相色谱技术对一引种虎眼万年青属植物Ornithogalum thyrsoides的化学成分进行研究,从其乙醇提取物的乙酸乙酯部分分离得到7个化合物.通过MS、NMR等波谱技术确定其结构分别为(23S,24S,25S)-23,24-dihydroxyruscogenin- 1-O-α-L-rhamnopyranosyl(1 →2)-α-L-arabinopyranoside(1),(24S,25S)-3β,24-dihydroxyspirost-5 -en- 1β-yl O-α -L-rhamnopyranosyl-(1→2)-α-L-arabinopyranoside(2),β-胡萝卜苷(3)β-谷甾醇(4),山奈酚-4′-O-β-D-吡喃葡萄糖苷(5),千层纸素A(6)和汉黄芩素(7).其中,化合物1、5～7为首次从该属植物中分离得到.体外药理活性结果显示这7个化合物对HCT-8、Bel-7402、BGC-823、A549及A2780实体瘤细胞株无明显细胞毒活性.     

羊角拗茎的化学成分及其抗炎活性研究

探讨羊角拗茎的化学成分及其抗炎活性,采用硅胶、Sephadex LH-20等色谱分离技术分离纯化羊角拗茎乙醇提取物的乙酸乙酯萃取物中的化合物,通过质谱和核磁共振等波谱技术对其结构进行鉴定,确定出了13个化合物,包括常春藤皂苷元(1)、(-)-loliolide(2)、(3S,5R,6S,7E)-3,5,6-trihydroxy-7-megastigmen-9-one(3)、(3R,6R,7E)-3-hydroxy-4,7-megastigmadien-9-one(4)、松脂素(5)、表松脂素(6)、4,4′-dihydroxy-3,3′-dimethoxybenzophenone(7)、6-羟基柚皮素(8)、东莨菪素(9)、6-羟基-7,8-二甲氧基香豆素(10)、吲唑(11)、香草酸(12)、对羟基苯甲酸(13)。其中化合物1～4和6～13为首次从羊角拗中分离得到。采用LPS诱导小鼠单核巨噬细胞RAW 264.7体外细胞炎症模型测试结果表明,化合物5能显著抑制NO的产生,IC₅₀值为18.09±1.09μM。     

Synthesis of tritium labeled Ac-[Nle4, D-Phe7]-alpha-MSH4-11-NH2: a superpotent melanotropin with prolonged biological activity

Ac-[Nle4, D-Phe7]-alpha-MSH4-11-NH2 an octapeptide, is a melanotropin analogue (Ac-Nle-Glu-His-D-Phe-Arg-Trp-Gly-Lys-NH2), which is a superpotent agonist of frog and lizard skin melanocytes and mouse S 91 (Cloudman) melanoma cells. This melanotropin possesses ultraprolonged activity on melanocytes, both in vitro and in vivo, and the peptide is resistant to inactivation by serum enzymes. The tritium-labeled congener was prepared by direct incorporation of [3H]-labeled norleucine into the peptide. The melanotropic activity of the labeled peptide is identical to the unlabeled analogue. This labeled peptide should be useful for studies on the localization and characterization of melanotropin receptors.     

Non-thiazolidinedione antihyperglycaemic agents. Part 3: The effects of stereochemistry on the potency of alpha-methoxy-beta-phenylpropanoic acids.

Rhizopus delemar lipase catalysed ester hydrolysis of the alpha-methoxy-beta-phenylpropanoate 1 affords the (R)-(+) and (S)-(-) isomers in > 84% enantiomeric excess. Absolute stereochemistry was determined by a single crystal X-ray analysis of a related synthetic analogue. The activity of these two enantiomers on glucose transport in vitro and as anti-diabetic agents in vivo is reported and their unexpected equivalence attributed to an enzyme-mediated stereospecific isomerisation of the (R)-(+) isomer. Binding studies using recombinant human PPARgamma (peroxisomal proliferator activated receptor gamma), now established as a molecular target for this compound class, indicate a 20-fold higher binding affinity for the (S) antipode relative to the (R) antipode.     

New Nystatin-Related Antifungal Polyene Macrolides with Altered Polyol Region Generated via Biosynthetic Engineering of Streptomyces noursei

Polyene macrolide antibiotics, including nystatin and amphotericin B, possess fungicidal activity and are being used as antifungal agents to treat both superficial and invasive fungal infections. Due to their toxicity, however, their clinical applications are relatively limited, and new-generation polyene macrolides with an improved therapeutic index are highly desirable. We subjected the polyol region of the heptaene nystatin analogue S44HP to biosynthetic engineering designed to remove and introduce hydroxyl groups in the C-9-C-10 region. This modification strategy involved inactivation of the P450 monooxygenase NysL and the dehydratase domain in module 15 (DH15) of the nystatin polyketide synthase. Subsequently, these modifications were combined with replacement of the exocyclic C-16 carboxyl with the methyl group through inactivation of the P450 monooxygenase NysN. Four new polyene macrolides with up to three chemical modifications were generated, produced at relatively high yields (up to 0.51 g/liter), purified, structurally characterized, and subjected to in vitro assays for antifungal and hemolytic activities. Introduction of a C-9 hydroxyl by DH15 inactivation also blocked NysL-catalyzed C-10 hydroxylation, and these modifications caused a drastic decrease in both antifungal and hemolytic activities of the resulting analogues. In contrast, single removal of the C-10 hydroxyl group by NysL inactivation had only a marginal effect on these activities. Results from the extended antifungal assays strongly suggested that the 9-hydroxy-10-deoxy S44HP analogues became fungistatic rather than fungicidal antibiotics.     

Synthesis and biochemical characterization of a phosphorylated analogue of the response regulator CheB.

CheB is a response regulator protein in the bacterial chemotaxis two-component signal transduction pathway. Methylesterase CheB functions together with methyltransferase CheR to modulate the level of glutamate methylation in transmembrane chemoreceptors in response to environmental stimuli. The level of glutamate methylation in turn indirectly controls the direction of flagellar rotation. Like most two-component response regulators, CheB is activated in vivo by phosphorylation of a single aspartate, Asp 56, in its regulatory domain. Extensive biochemical and crystallographic studies have been completed on the inactive, unphosphorylated form of CheB. Because of the inherent lability of aspartyl phosphate bonds and the intrinsic phosphatase activity of CheB, the activated, phosphorylated form of CheB cannot be isolated for further characterization. We present a synthetic scheme to prepare an analogue of phosphorylated CheB using site-specific mutagenesis and chemical modification strategies. Initially, the two native cysteines found in CheB were substituted by serines and a cysteine was substituted for Asp 56 to yield D56C/C207S/C309S CheB. The unique cysteine in the substituted form of CheB was modified by sodium thiophosphate, Na(3)SPO(3), using two sequential disulfide bond exchange reactions. The analogue, D56C/C207S/C309S CheB-SPO(3), contained a thiophosphate group covalently bonded to the protein through a disulfide linkage at residue 56. Mass spectrometry showed that the protein was singly modified. Reverse phase chromatography showed that greater than 95% of the protein was modified under optimized conditions and that the analogue had a half-life of 28 days. In in vitro methylesterase assays in the presence of Mg(2+), the analogue exhibited activity equivalent to that of fully phosphorylated C207S/C309S CheB. Thus, D56C/C207S/C309S CheB-SPO(3) is a stable analogue that may be useful for characterization of the active form of CheB.     

Novel cyclohexene derivatives as anti-sepsis agents: synthetic studies and inhibition of NO and cytokine production

In order to develop an anti-sepsis agent, a series of cyclohexene derivatives were synthesized and evaluated for their biological activities. Through modification of the sulfonamide spacer moiety depicted by formula II, it was found that the benzylsulfone derivative 10a had potent inhibitory activity against the production of NO. Further modifications of the phenyl ring, ester moiety, and benzyl position of benzylsulfone derivatives III were carried out. Among these compounds, (R)-(+)-10a and (6R, 1S)-(+)-22a showed strong inhibitory activity not only against NO production but also against inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in vitro. Furthermore, (R)-(+)-10a and (6R, 1S)-(+)-22a protected mice from LPS-induced lethality in a dose-dependent manner.     

Synthesis and evaluation of novel pseudomycin side-chain analogues. Part 3.

To increase the therapeutic utility of C-18 side-chain bearing pseudomycin analogue 2, we prepared additional analogues and prodrugs of 2 containing further modifications at various positions within its core structure. Each of the newly synthesized derivatives (10-15) exhibited reduced tail vein toxicity relative to the parent compound. Some of the new pseudomycin derivatives (e.g., 14) also showed improved in vivo antifungal activity relative to its corresponding parent compound.