The use of a hypoxic cell radiosensitizer AK-2123 gave no improvement in thermoradiotherapy combined with hydralazine

An electron-affinic compound, AK-2123, and the anti-hypertensive agent, hydralazine, were combined with radiation and hyperthermia for treatment of murine SCC-VII tumours. Hydralazine markedly decreased tumour perfusion while AK-2123 had no influence on it. Hydralazine enhanced the tumouricidal effects of hyperthermia alone and in combination with radiation. AK-2123 provided a radiosensitization which was significant only in tumours irradiated without supplementary hyperthermia. The greatest tumour response was achieved when thermoradiotherapy was combined with hydralazine alone; the additional use of AK-2123 with this treatment combination did not further increase the effect. It is concluded that hydralazine plus heat virtually eliminated a hypoxia-related radioresistance in tumours, thus removing the requirement for AK-2123 administration.     

De novo design, synthesis, and biological activities of high-affinity and selective non-peptide agonists of the delta-opioid receptor

On the basis of the structure-activity relationships of delta-opioid-selective peptide ligands and on a model of the proposed bioactive conformation for a potent and selective, conformationally constrained delta-opioid peptide ligand [(2S, 3R)-TMT1]DPDPE, a series of small organic peptide mimetic compounds targeted for the delta-opioid receptor have been designed, synthesized, and evaluated in radiolabeled ligand binding assays and in vitro bioassays. The new non-peptide ligands use piperazine as a template to present the most important pharmacophore groups, including phenol and phenyl groups and a hydrophobic moiety. This hydrophobic group was designed to mimic the hydrophobic character of the D-Pen residues in DPDPE, which has been found to be extremely important for increasing the binding affinity and selectivity of these non-peptide ligands for the delta-opioid receptor over the mu-opioid receptor. Compound 6f (SL-3111) showed 8 nM binding affinity and over 2000-fold selectivity for the delta-opioid receptor over the mu-opioid receptor. Both enantiomers of SL-3111 were separated, and the (-)-isomer was shown to be the compound with the highest affinity for the delta-opioid receptor found in our study (IC50 = 4.1 nM), with a selectivity very similar to that observed for the racemic compound. The phenol hydroxyl group of SL-3111 turned out to be essential to maintain high affinity for the delta-opioid receptor, which also was observed in the case of the delta-opioid-selective peptide ligand DPDPE. Binding studies of SL-3111 and [p-ClPhe4]DPDPE on the cloned wild-type and mutated human delta-opioid receptors suggested that the new non-peptide ligand has a binding profile similar to that of DPDPE but different from that of (+)-4-[((alphaR)-alpha(2S,5R)-4-allyl-2, 5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC-80), another delta-opioid-selective non-peptide ligand.     

Design, synthesis, and biological activities of potent and selective somatostatin analogues incorporating novel peptoid residues

We report the synthesis, bioactivity, and structure-activity relationship studies of compounds related to the Merck cyclic hexapeptide c[Pro6-Phe7-d-Trp8-Lys9-Thr10-Phe11], L-363,301 (the numbering in the sequence refers to the position of the residues in native somatostatin). The Pro residue in this compound is replaced with arylalkyl peptoid residues. We present a novel approach utilizing beta-methyl chiral substitutions to constrain the peptoid side-chain conformation. Our studies led to molecules which show potent binding and increased selectivity to the hsst2 receptor (weaker binding to the hsst3 and hsst5 receptors compared to L-363, 301). In vivo, these peptoid analogues selectively inhibit the release of growth hormone but have no effect on the inhibition of insulin. The biological assays which include binding to five recombinant human somatostatin receptors carried out in two independent laboratories and in vivo inhibition of growth hormone and insulin provide insight into the relationship between structure and biological activity of somatostatin analogues. Our results have important implications for the study of other peptide hormones and neurotransmitters.     

Implantation of a synthetic cornea: design, development and biological response

Our goal was to evaluate 3 different designs of synthetic corneas in vivo. All devices had a transparent hydrogel center molded to a porous peripheral skirt. Over 30 devices were implanted into rabbits and followed for up to 6 months. The devices were preseeded with rabbit stromal fibroblasts, which enhanced the rate of fibroplasia. The anterior surface of the hydrogel was modified using argon rf plasma treatments. Clinical examinations were performed, and histological analyses were conducted on tissue throughout the time course. Our optimal model ranged from 4.5 to 6 mm and had an extended porous skirt increasing the surface area for fibroplasia and ultimate anchorage of the device. Fibroplasia occurred in this model, and collagen was detected by 28 days. The anterior chamber was normal with no detectable leakage of aqueous humor. Glycosaminoglycans were detected and followed the time course outlined previously when porous material itself was inserted into the stroma. We present the first demonstration that rabbit limbal epithelial cells can migrate onto the synthetic cornea in vivo.     

The Gabriel-Colman rearrangement in biological systems: design, synthesis and biological evaluation of phthalimide and saccharin derivatives as potential mechanism-based inhibitors of human leukocyte elastase, cathepsin G and proteinase 3

Metal ion-promoted binding of Saccharomyces cerevisiae alcohol dehydrogenase to Cibacron Blue 3GA was used for its isolation by affinity precipitation with Eudragit-bound dye. The yeast cells were broken and the cell debris was separated by flocculation with poly(ethylene imine). The supernatant containing the enzyme activity was mixed with Eudragit--Cibacron Blue in the presence of zinc ions. The precipitation of the affinity complex was induced by the addition of 50 mM CaCl2 and a subsequent increase in temperature to 40 degrees C. The enzyme was desorbed by treating the precipitate with iminodiacetic acid solution. The procedure resulted in about 66% recovery of enzyme activity with more than 20-fold purification. Recycling of Eudragit--dye for enzyme purification was also shown to be possible.     

Thymidine-based gelators: design, synthesis, and evaluation

Through the simple modification of nucleoside, a new class of organogelator was synthesized. Five kinds of thymidine-based organogelators gelated in various organic solvents. Their SEM microscopic images showed three different types. The driving force of gelation is mainly hydrogen bonding interaction.     

Polyunsaturated fatty acids, Part 1: Occurrence, biological activities and applications

Polyunsaturated fatty acids form a unique class of food constituents that show a wide range of functions in biological systems. Investigations over the past two decades have uncovered their roles and those of their eicosanoid metabolites, and have highlighted their homeostatic functions in mammals. A growing number of common human medical conditions are thought to be traceable to dysfunctions in the eicosanoid system, which could in turn be due to imbalances in the intake and/or metabolism of polyunsaturated fatty acids. This, together with medical advances, has spurred the introduction of biomedical products, nutritionals, fortified foods and health supplements.     

Tri-N-Boc-tetraazamacrocycle-nucleoside conjugates: synthesis and anti-HIV activities

As far as linear N-Boc-polyamines conjugates elicited remarkable anti-HIV activity, the synthesis and anti-HIV properties of cyclic N-Boc-polyamines conjugates such as tetraazamacrocycle-nucleoside were studied. These new conjugates include an ester linkage between the two moieties. They were synthesized using Benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate coupling reagent, in the case of N-alkyl polyazamacrocycle derivatives, or through direct condensation of the acyl chloride derivative with nucleoside in the case of N-acyl polyazamacrocycle compounds. None of the new conjugates presented anti-HIV activity greater than that of the corresponding parent nucleosides.     

Search for alpha 1-adrenoceptor subtypes selective antagonists: design, synthesis and biological activity of cystazosin, an alpha 1D-adrenoceptor antagonist

Two novel quinazolines (2 and 3) related to both prazosin and its open analogue 1 were synthesized, and their biological profile at alpha 1-adrenoceptor subtypes was assessed by functional assays in rat isolated tissues, namely prostatic vas deferens (alpha 1A), spleen (alpha 1B) and aorta (alpha 1D). Furthermore, the binding profile of 3 was assessed at native alpha 2 and D2 receptors, and cloned human 5-HT1A receptors, in comparison to prazosin, (+)-cyclazosin, 1 and BMY 7383. It turned out that the cystamine-bearing quinazoline 3 (cystazosin) has a reversed affinity profile relative to (+)-cyclazosin owing to a higher affinity for alpha 1D-adrenoceptors and a significantly lower affinity for the alpha 1A and alpha 1B subtypes. Furthermore, in comparison to BMY 7378, cystazosin (3) displays a much better specificity profile since it has lower affinity for D2 and 5-HT1A receptors.     

Excitatory amino acid synthesis in hypoxic brain slices: does alanine act as a substrate for glutamate production in hypoxia?

Excitatory amino acids are an important cause of cell death in the hypoxic and ischaemic brain. Neuronal glutamate stores are depleted rapidly in hypoxia, but alanine production rises under such conditions and has been suggested to be a potential precursor of glutamate. To test this hypothesis, we have investigated amino acid metabolism using 13C NMR with superfused guinea pig cortical slices subjected to varying degrees of hypoxia. During severe hypoxia, brain slices metabolising 5 mM [2-(13)C]pyruvate exported [2-(13)C]alanine into the superfusion fluid. The metabolic fate of alanine during normoxia and hypoxia was tested by superfusion of brain slices with 10 mM glucose and 2 mM [2-(13)C,15N]alanine. Metabolism of exogenous alanine leads to the release of aspartate into the superfusion fluid. The pattern of labelling of aspartate indicated that it was synthesised via the glial-specific enzyme pyruvate carboxylase. 13C-labelled glutamate was produced with both normoxia and hypoxia, but concentrations were 30-fold lower than for labelled aspartate. Thus, although substantial amounts of glutamate are not synthesised from alanine in hypoxia, there is significant production of aspartate, which also may have deleterious effects as an excitatory amino acid.     

Ethics, epidemiology and the thrifty gene: biological determinism as a health hazard

This paper briefly describes the rise of the thrifty genotype hypothesis as an explanation for the late twentieth century epidemic of diabetes, particularly in post-colonial indigenous societies. It looks at some of the ethical consequences of the biological deterministic paradigm, particularly the popular confusion of "genes" with "race" and how this paradigm served to exclude consideration of social determinants of disease in epidemiological thinking. Some alternative hypotheses to the thrifty gene theory are explored, together with the consequences of acceptance of these other theories in terms of public health action. Finally, there is a need for epidemiology to be continually conscious, critical and transparent with respect to the general disease (and wellness) theory under which it operates if it is to be truly a science rather than a collection of methodologies.     

Aromatase inhibitors: synthesis, biological activity, and binding mode of azole-type compounds

The enantiomers of the potent nonsteroidal inhibitor of aromatase fadrozole hydrochloride 3 have been separated and their absolute configuration determined by X-ray crystallography. On the basis of a molecular modeling comparison of the active enantiomer 4 and one of the most potent steroidal inhibitors reported to date, (19R)-10-thiiranylestr-4-ene-3,17-dione, 7, a model describing the relative binding modes of the azole-type and steroidal inhibitors of aromatase at the active site of the enzyme is proposed. It is suggested that the cyanophenyl moiety present in the most active azole inhibitors partially mimics the steroid backbone of the natural substrate for aromatase, androst-4-ene-3,17-dione, 1. The synthesis and biological testing of novel analogues of 3 used to define the accessible and nonaccessible volumes to ligands in the model of the active site of aromatase are reported.     

Protein structure-based design, synthesis, and biological evaluation of 5-thia-2,6-diamino-4(3H)-oxopyrimidines: potent inhibitors of glycinamide ribonucleotide transformylase with potent cell growth inhibition

The design, synthesis, biochemical, and biological evaluation of a novel series of 5-thia-2,6-diamino-4(3H)-oxopyrimidine inhibitors of glycinamide ribonucleotide transformylase (GART) are described. The compounds were designed using the X-ray crystal structure of human GART. The monocyclic 5-thiapyrimidinones were synthesized by coupling an alkyl thiol with 5-bromo-2, 6-diamino-4(3H)-pyrimidinone, 20. The bicyclic compounds were prepared in both racemic and diastereomerically pure forms using two distinct synthetic routes. The compounds were found to have human GART KiS ranging from 30 microM to 2 nM. The compounds inhibited the growth of both L1210 and CCRF-CEM cells in culture with potencies down to the low nanomolar range and were found to be selective for the de novo purine biosynthesis pathway. The most potent inhibitors had 2,5-disubstituted thiophene rings attached to the glutamate moiety. Placement of a methyl substituent at the 4-position of the thiophene ring to give compounds 10, 18, and 19 resulted in inhibitors with significantly decreased mFBP affinity.     

Selective Pneumocystis carinii dihydrofolate reductase inhibitors: design, synthesis, and biological evaluation of new 2,4-diamino-5-substituted-furo[2,3-d]pyrimidines

Nonclassical antifolates, 2,4-diamino-5-substituted-furo[2, 3-d]pyrimidines 3-12 with bridge region variations of C8-S9, C8-N9, and C8-O9 and 1-naphthyl, 2-naphthyl, 2-phenoxyphenyl, 4-phenoxyphenyl, and 2-biphenyl side chains were synthesized as phenyl ring appended analogues of previously reported 2, 4-diamino-5-(anilinomethyl)furo[2,3-d]pyrimidines. The phenyl ring appended analogues were designed to specifically interact with Phe69 of dihydrofolate reductase (DHFR) from Pneumocystis carinii (pc) to afford selective inhibitors of pcDHFR. Additional substituted phenyl side chains which include 2,5-dichloro, 3,4-dichloro, 3,4,5-trichloro, 3-methoxy, and 2,5-dimethoxy analogues 13-17 were also synthesized. The compounds were prepared by nucleophilic displacement of 2,4-diamino-5-(chloromethyl)furo[2,3-d]pyrimidine(2) with the appropriate thiol, amine, or naphthol. Compound 2 was obtained from 2,4-diamino-6-hydroxypyrimidine and 1, 3-dichloroacetone. The compounds were evaluated as inhibitors against DHFR from P. carinii, Toxoplasma gondii, and rat liver. Two analogues, 2,4-diamino-5-[(2'-naphthylthio)methyl]furo[2, 3-d]pyrimidine (5) and 2,4-diamino-5-[(2'-phenylanilino)methyl]furo[2,3-d]pyrimidine (11) showed significant selectivity and potency for pcDHFR compared to trimethoprim. The X-ray crystal structure of 5 with pcDHFR was also carried out, which corroborated the design rationale and indicated a hydrophobic interaction of the naphthalene ring of 5 and Phe69 of pcDHFR which is responsible, in part, for the more than 18-fold selectivity of 5 for pcDHFR as compared with rat liver DHFR.     

The design, synthesis, and initial evaluation of benzophenone-containing peptides as potential photoaffinity labels of oligosaccharyltransferase

The benzophenone photophore was incorporated into protected tripeptides and tetrapeptides as photoactivatable probes to study the multimeric enzyme oligosaccharyltransferase (OST). These peptides contain the -Asn-X-Thr- sequon which is required for OST-catalyzed N-glycosylation. Two tripeptides, Bz-Asn-Bpa-Thr-NH2 (3b) and Bz-Asn-Lys[N epsilon-(4-Bz)Bz]-Thr-NH2 (4b), were found to be good OST substrates. They were competitive inhibitors versus standard peptide substrate [14C]Bz-Asn-Leu-Thr-NH2 and their Ki values were determined to be 41 +/- 6 microM and 21 +/- 6 microM, respectively, using synthetic (GlcNAc)2-PP-dolichol.     

Design, synthesis, and biological activities of four angiotensin II receptor ligands with gamma-turn mimetics replacing amino acid residues 3-5

Disulfide cyclization is a powerful method for reducing the conformational space of a peptide. This in turn may enable the study of its bioactive conformation. Several analogues of angiotensin II (Ang II) containing a disulfide bridge between amino acids 3 and 5 have been reported. Among these the cyclic octapeptides c[Hcy3,5]-Ang II, c[Cys3,5]-Ang II, and c[Pen 3,5]-Ang II showed significant activity at Ang II receptors. We have performed conformational analysis studies using theoretical calculations and 1H-NMR spectroscopy on tripeptide model compounds of these cyclic octapeptides which show that the cyclic moieties of c[Cys3,5]-Ang II and c[Pen3,5]-Ang II preferentially assume an inverse gamma-turn conformation. On the basis of these results, we substituted amino acid residues 3-5 in Ang II with two different gamma-turn mimetics giving four diastereomeric Ang II analogues. Interestingly, two of these are equipotent to Ang II in binding to AT1 receptors. In the contractile test using rabbit aorta rings, one of the analogues is an agonist with full contractile activity approximately equipotent to c[Pen3,5]-Ang II but 300-fold less potent than Ang II. This low potency may suggest that Ang II does not adopt a gamma-turn in the 3-5 region when interacting with the receptor.