Characterization of [3H]-(2S,2′R,3′R)-2-(2′,3′-dicarboxy- cyclopropyl)glycine ([3H]-DCG IV) binding to metabotropic mGlu2 receptor-transfected cell membranes

1. The binding of the new selective group II metabotropic glutamate receptor radioligand, [³H]-(2S,2′R,3′R)-2-(2′,3′-dicarboxycyclopropyl)glycine ([³H]-DCG IV), was characterized in rat mGlu₂ receptor-transfected CHO cell membranes.
2. [³H]-DCG IV binding was pH-dependent, but was not sensitive to temperature. Saturation analysis showed the presence of a single binding site, with a K_d value of 160 nM and a B_max value of 10 pmol mg⁻¹ protein. Binding was not sensitive to Na⁺-dependent glutamate uptake blockers or Cl⁻-dependent glutamate binding inhibitors. Furthermore, up to concentrations of 1 mM, the glutamate ionotropic receptor agonists, N-methyl-D-aspartic acid (NMDA), (S)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainate, did not affect [³H]-DCG IV binding.
3. Of the compounds observed to inhibit [³H]-DCG IV binding, the most potent were the recently described selective group II agonist, (+)-2-aminobicyclo-[3.1.0]hexane-2,6-dicarboxylate (LY 354740; K_i value 16 nM) and antagonist, 2-amino-2-(2-carboxycyclopropan-1-yl)-3-(dibenzopyran-4-yl) propanoic acid (LY 341495; K_i value 19 nM). As expected, for a G-protein-coupled receptor, guanosine-5′-O-(3-thiotriphosphate) (GTPγS) inhibited [³H]-DCG IV binding in a concentration-dependent manner, with an IC₅₀ value of 12 nM.
4. A highly significant correlation was observed between the potencies of compounds able to inhibit [³H]-DCG IV binding and potencies obtained for agonist activity in a GTPγ³⁵S binding functional assay. In addition, these studies identified a number of compounds with previously unknown activity at mGlu₂ receptors, including L(+)-2-amino-3-phosphonopropionic acid (L-AP3), L(+)-2-amino-5-phosphonopentanoic acid (L-AP5), 3-((RS)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (R-CPP), N-acetyl-L-aspartyl-L-glutamic acid (NAAG) and (RS)-α-methylserine-O-phosphate (MSOP).

     

Synthesis of novel 2-(3′-aryl-sydnon-4′-ylidene)-5′-substituted-[1,3,4]-thiadiazolylamines and [1,3,4]-thiadiazol-2′-yl-3-oxo-[1,2,4]-triazoles as antimicrobial agents

The title compounds (3g–n) and (6g–n) were synthesized using 3-arylsydnones as synthons, and the structures were confirmed by IR, ¹H NMR, FAB mass and CHN analysis. These compounds were evaluated for their antibacterial and the antifungal activities in terms of minimum inhibitory concentrations (MICs) against the bacterial strains E. coli, B. cereus, and the fungal strains A. niger, C. albicans. Some of the compounds have shown significant activities.     

Tolerance to μ-opioid agonists in human neuroblastoma SH-SY5Y cells as determined by changes in guanosine-5′-O-(3-[35S]-thio)triphosphate binding

1. The agonist action of morphine on membranes prepared from human neuroblastoma SH-SY5Y cells was measured by an increase in the binding of the GTP analogue [³⁵S]-GTPγS. Morphine increased the binding of [³⁵S]-GTPγS to SH-SY5Y cell membranes by 30 fmol mg⁻¹ protein with an EC₅₀ value of 76±10 nM.
2. Incubation of SH-SY5Y cells with 10 μM morphine for 48 h caused a tolerance to morphine manifested by a 2.5 fold shift to the right in the EC₅₀ value with a 31±6% decrease in the maximum stimulation of [³⁵ S]-GTPγS binding. The response caused by the partial agonist pentazocine was reduced to a greater extent.
3. Chronic treatment of the cells with the more efficacious μ-ligand [D-Ala², MePhe⁴, Gly-ol⁵]enkephalin (DAMGO, 10 μM) for 48 h afforded a greater effect than treatment with morphine. The maximal agonist effect of morphine was reduced to 58.9±6% of that seen in control cells while the maximal effect of DAMGO was reduced to 62.8±4%. There was a complete loss of agonist activity for pentazocine.
4. The development of tolerance was complete within 24 h and was blocked by naloxone and by the nonselective protein kinase inhibitor H7, but not by the putative β-adrenoceptor kinase (β-ARK) inhibitor suramin.
5. The observed tolerance effect was accompanied by a down-regulation of μ-opioid receptors determined by a decrease in the maximal binding capacity for the opioid antagonist [³H]-diprenorphine of 66±4%, but with no change in binding affinity. Binding of the agonist [³H]-DAMGO was similarly reduced.
6. The modulation of [³⁵S]-GTPγS binding in SH-SY5Y cell membranes by opioids provides a simple method for the study of opioid tolerance at a site early in the signal transduction cascade.

     

Potential antitumor α-methylene-ψ-butyrolactone-bearing nucleic acid bases. 2. Synthesis of 5′-methyl-5′-[2-(5-substituted uracil-1-yl)ethyl]-2′-oxo-3′-methylenetetrahydrofurans

Ten, heretofore unreported, 5′-methyl-5′-[2-(5-substituted uracil-1-yl)ethyl)]-2′-oxo-3′-methylenetetrahydrofurans (H, F, Cl, Br, I, CH₃, CH₃, CH₂CH₃, CH=CH₂, SePh) (7a-j) were synthesized and evaluated against four cell lines (K-562, FM-3A, P-388 and U-937). For the preparation of α-methylene-γ-butyrolactone-linked to 5-substituted uracils (7a-j), the convenient Reformasky type reaction was employed which involves the treatment of ethyl α-(bromomethyl)acrylate and zinc with the respective 1-(5-substituted uracil-1-yl)-3-butanone (6a-j). The 5-substituted uracil ketones (6a-j) were directly obtained by the respective Michael type reaction of vinyl methyl ketone with the K₂CO₃ (or NaH)-treated 5-substituted uracils (5a-j) in the presence of acetic acid in the DMF solvent. The α-methylene-γ-butyrolactone compounds showing the most significant antitumor activity are7e, 7f, 7h and7j (inhibitory concentration (IC₅₀) ranging from 0.69 to 2.9 μg/ml), while7b, 7g and7i have shown moderate to significant activity. The compounds7a, 7c and7d were found to be inactive. The synthetic intermediate compounds6a-j were also screened and found marginal to moderate activity where compounds6b and6g showed significant activity (IC₅₀:0.4∼2.8 μg/ml).     

(2R,3S)-2-[(R)-1-[3,5-二(三氟甲基)苯基]乙氧基]-3-(4-氟苯基)吗啉盐酸盐的合成

对甲氧基苯甲醛(3)和2-氨基乙醇进行还原胺化反应得2-(4-甲氧基苄胺基)乙醇(4),4和乙醛酸经成环反应得2-羟基-4-对甲氧基苄基吗啉-3-酮(5),5和三氟乙酐反应得6后与(R)-1-[3,5-二(三氟甲基)苯基]乙醇(7)缩合,再经结晶诱导不对称转化、格氏反应、氢化脱保护及成盐反应制得阿瑞吡坦关键中间体(2R,3S)-2-[(R)-1-[3,5-二(三氟甲基)苯基]乙氧基]-3-(4-氟苯基)吗啉盐酸盐,总收率约18%(以3计)。     

cis-trans-Isomerization of [E]-5-(2-bromovinyl)-2,2′-anhydrouridine in vivo in rats

1. [E]-5-(2-bromovinyl)-2,2′-anhydrouridine ([E]BVANUR) has considerable antiviral activity against herpes simplex virus type 1 (HSV-1).

2. [E]BVANUR is not a substrate of pyrimidine nucleoside phosphorylases, but it is an inhibitor of uridine phosphorylase (K_i=450 nM).

3. [E]BVANUR (trans-isomer, parent compound) undergoes isomerization to [Z]BVANUR (cis-isomer), the only metabolite in rat, which was identified by?h.p.l.c., mass spectra and n.m.r. spectroscopy.

4. Absorption of the drug from the gastrointestinal tract after oral administration is minimal. Absorption of [E]BVANUR from the abdominal cavity after i.p. administration was slow.     

Nonenzymatic and Enzymatic Hydrolysis of 5-Fluoro-2′-deoxyuridine (FUdR) Esters

The chemical and enzymatic reactivity of 5-fluoro-2-deoxyuridine prodrugs esterified at the 3 and 5 positions with several acyl groups has been investigated. The enzymatic reactivity was affected by the acyl structure, the site of esterification, and the number of esters in the prodrug molecule.     

N~6-2′-O-双丁酰-3′,5′-环化腺苷酸的生产

随着cAMP作用机理研究的深入,人们发现cAMP进入机体后很容易被磷酸二酯酶水解而失去作用的弊病。此外Posternak等人认为,由于cAMP分子具有一定的极性,影响进入细胞的能力。所以近几年来对cAMP衍生物的研究及试制开始引起人们的注意。迄今已合成的cAMP衍生物中大致不外乎以下几类:(1)N~6—NH_2部位的改变;(2)2'—OH的取代;(3)磷酰基的改变;(4)C—8的取代。而最早合成及使用最多的衍生物是N~6—2’—O—双丁酰环化腺苷酸(DBC),于1967年经Falbriard合成。我们在按Falbriard方法     

Synthesis of N-[3′-(acetylthio)alkanoyl] and N-[3′-mercaptoalkanoyl]-4-α(s)-(phenylmethyl) pyroglutamic acids and prolines as potent ACE inhibitors

Abstract  

Angiotensin converting enzyme (ACE) inhibitors have emerged as a revolution in antihypertensive therapy. Introduction of captopril, the first rationally designed ACE inhibitor, has encouraged researchers all over the world to design and synthesize target molecules controlling hypertension based on these lines. It has been observed that replacing proline part of captopril with 4-substituted prolines or 5-oxo-prolines led to significant enhancement in ACE inhibitory activity, and this observation prompted us to design and synthesize N-acyl 4-substituted pyroglutamates and prolinates with the objective of developing therapeutically better ACE inhibitors. Herein we describe an easy approach for N-acylation of 4-α(S)-(phenylmethyl) pyroglutamates with the aim of synthesizing N-[3′-(acetylthio)alkanoyl] and N-[3′-mercaptoalkanoyl]-4-α-(s)-(phenylmethyl) pyroglutamic acids and prolines as ACE inhibitors.     

2′,3′-O-(2,4,6- trinitrophenyl) adenosine 5′-triphosphate (TNP-ATP)–a nanomolar affinity antagonist at rat mesenteric artery P2X receptor ion channels

1. P2X receptor activation by α,β-meATP evoked inward currents in acutely dissociated rat mesenteric artery smooth muscle cells and contractions of whole artery rings.
2. The selective P2X₁ and P2X₃ receptor antagonist TNP-ATP inhibited P2X receptor mediated inward currents in response to 3 μM α,β-meATP (an ∼EC₉₀ concentration) with an IC₅₀ of ∼2 nM. This provides further evidence that the P2X receptor underlying membrane depolarisation associated with P2X receptor activation can be accounted for by the expression of P2X₁ receptors.
3. TNP-ATP inhibited α,β-meATP induced contractions with an IC₅₀ of ∼30 μM and had non-specific effects on smooth muscle contraction.
4. The reduced potency of TNP-ATP in whole tissue experiments probably reflects the breakdown of TNP-ATP by nucleotidases. Thus, TNP-ATP is of limited use in whole tissue experiments as a P2X receptor antagonist.

     

Synthesis of bioactive spiro-2-[3′-(2′-phenyl)-3H-indolyl]-1-aryl-3-phenylaziridines and SAR studies on their antimicrobial behavior

The present communication deals with synthesis of a new series of spiro-2-[3′-(2′-phenyl)-3H-indolyl]-1-aryl-3-phenylaziridines. Infrared (IR), ¹H nuclear magnetic resonance (NMR), mass spectral, and elemental analysis data corroborated the structure of all synthesized compounds. Synthesized compounds were screened for antimicrobial activity against a representative panel of Gram-positive and Gram-negative bacteria. All newly synthesized compounds showed remarkable antimicrobial behavior. Structure–activity relationship (SAR) investigations were applied to investigate the correlation between the molecular refractive index (M_R) parameter of the compounds and their biological activity profile. All compounds were subjected to acute toxicity studies to determine 50% lethal dose (LD₅₀) values.     

In vitro and in vivo evaluation of radiolabeled methyl N-[5-(3′-halobenzoyl)-1H-benzimidazol-2-yl]carbamate for cancer radiotherapy

The role of theranostics in cancer management is growing so is the selection of vectors used to deliver these modalities to cancer cells. We describe biological evaluation of a novel theranostic agent targeted to microtubules. Methyl N-[5-(3′-[¹³¹I]iodobenzoyl)-1H-benzimidazol-2-yl]carbamate ( 1 ) and methyl N-[5-(3′-[¹²⁵I]iodobenzoyl)-1H-benzimidazol-2-yl]carbamate ( 2 ) were synthesized from a common precursor 3′-stannylated derivative ( 4 ). Antiproliferative effects and radiotoxicity of ¹³¹I-labeled β-particle emitting 1 were examined in vitro in human neuroblastoma and glioblastoma cells lines. The therapeutic potential of 1 was also examined in a subcutaneous mouse model of human glioblastoma U-87 MG. Compound 1 at the extracellular radioactive concentration of 0.35 MBq/mL, easily achievable in vivo, kills >90% of neuroblastoma cells and >60% glioblastoma cells as measured in a clonogenic assay. D₁₀ doses established for 1 indicate that as few as 3,000 decays are sufficient to kill 90% of BE(2)-C cells. Even U-87 MG cells, the least sensitive of the tested cell lines, require <20,000 decays of intracellular ¹³¹I to reduce number of clonogenic cells by 90%. Biodistribution studies of 2 delivered either intratumorally or intraperitoneally show a similar tissue distribution for both routes of the drug administration. The whole body clearance half-lives were on average 6 hr. Intratumor administration of 1 produces significant tumor growth delay. After a single dose of 8.4 ± 0.3 MBq of compound 1 , the tumor doubling times were 3.2 ± 0.1 and 7.9 ± 0.6 days in control and treated mice, respectively. Methyl N-[5-(3′-radiohalobenzoyl)-1H-benzimidazol-2-yl]carbamates have properties compatible with a theranostic approach to cancer management.     

3′-(4-(Benzyloxy)phenyl)-1′-phenyl-5-(heteroaryl/aryl)-3,4-dihydro-1′H,2H-[3,4′-bipyrazole]-2-carboxamides as EGFR kinase inhibitors: Synthesis,anticancer evaluation,and molecular docking studies

Pyrazoline-linked carboxamide derivatives were designed, synthesized, and evaluated for potential epidermal growth factor receptor (EGFR) kinase inhibition, anticancer activity, and apoptotic and cardiomyopathy toxicity. Compounds 6m and 6n inhibit EGFR kinase at a concentration of 6.5 ± 2.91 and 3.65 ± 0.54 µM, respectively. Some of these compounds showed effects on proliferation, which were also then evaluated against four different human cancer cell lines, that is, MCF-7 (breast cancer), A549 (non-small-cell lung tumor), HCT-116 (colon cancer), and SiHa cells (cancerous tissues of the cervix uteri). The results showed that certain synthetic compounds showed significant inhibitor activity; compounds 6m and 6n were more cytotoxic than doxorubicin against A549 cancer cells, with IC₅₀ values of 10.3 ± 1.07 and 4.6 ± 0.57 µM, respectively. Additionally, compounds 6m and 6n induced apoptosis in A549 cancer cells, as evidenced by 4′,6-diamidino-2-phenylindole (DAPI) staining and phase-contrast microscopy. Potency to induce apoptosis by compound 6n was further confirmed by fluorescence-activated cell sorting using Annexin V-FITC and propidium iodide labeling. Compound 6n showed normal cardiomyocytes with no marked sign of pyknotic nuclei in cardiomyopathy and also normal histological appearance of the renal cortex when compared with that of control. Results of molecular docking studies suggested that compounds 6m and 6n can bind to the hinge region of the adenosine triphosphate-binding site of EGFR kinase, like the standard drug erlotinib. Therefore, the present study suggests that compounds 6m and 6n have potent in vitro antitumor activities against the human non-small-cell lung tumor cell line A549, which can be further explored in other cancer cell lines and in animal studies.     

Synthesis of a radiotracer for studying nicotinic acetylcholine receptors: (+/−)-exo-2-(2-[18F]fluoro-5-pyridyl)-7-azabicyclo[2.2.1]heptane

The radiochemical synthesis of (+/−)-exo-2-(2-[¹⁸F]fluoro-5-pyridyl)-7-azabicyclo[2.2.1]heptane ([¹⁸F] (UNDERLINE)1(/UNDERLINE) ) was accomplished by Kryptofix® 222 assisted nucleophilic no-carrier-added [¹⁸F]fluorination of (+/−)-exo-2-(2-bromo-5-pyridyl)-7-azabicyclo[2.2.1] heptane ( (UNDERLINE)3a(/UNDERLINE) ). The average radiochemical yield of the final product was 10% and the average specific activity was greater than >2000 mCi/μmol, calculated at end-of-synthesis. The stable fluorine ligand ([¹⁹F] (UNDERLINE)1(/UNDERLINE) ) was prepared by Kryptofix® 222 assisted nucleophilic fluorination of (+/−)-exo-2-(2-bromo-5-pyridyl) - 7 -methoxycarbonyl - 7 - azabicyclo[2.2.1]heptane ( (UNDERLINE)3b(/UNDERLINE) ) followed by acid deprotection.     

5′-RS-1,5-二氧代-(5′-乙基-5′-羟基-2′H,5′H,6′H-6-氧代吡喃)-[3′,4′,f]-Δ6(8)-四氢中氮茚的合成工艺改进

目的改进喜树碱关键中间体5′-RS-1,5-二氧代-(5′-乙基-5′-羟基-2′H,5′H,6′H-6-氧代吡喃) -[3′,4′,f]-Δ6(8)-四氢中氮茚(1)的合成工艺.方法以6-氰基-1,1-亚乙二氧基-7-(1′-乙氧羰基)丙基-5-氧代-Δ6(8)-四氢中氮茚(2)为原料,经氢化和亚硝化、脱氮、混合金属催化氧化、环合及三氟乙酸脱保护反应得到目标产物.结果与结论新工艺简化了操作、缩短了反应时间,总产率达到了72.4%.     

Synthesis and antitumor screening of new series of pyrimido-[4,5-b]quinolines and [1,2,4]triazolo[2′,3′:3,4]pyrimido[6,5-b]quinolines

New series of pyrimido[4,5-b]quinolines and [1,2,4]triazolo[2′,3′:3,4]pyrimido[6,5-b]quinolines have been synthesized. Compounds 4a, 4e, 4f, 4h, 5b, 5d, 6a, 6d, 6e, 8c, 8d, 10c–e, 10h, 11a, 11b, and 12a were tested for in vitro antitumor activity against human breast carcinoma (MCF-7) cell line, where compound 8d was found to be the most active member with IC₅₀ value of 3.62 μM. The DNA-binding affinity for the same compounds showed that compounds 8d and 10d exhibited the highest affinity to DNA. The detailed synthesis, spectroscopic, and biological data are reported.