One of the first steps in HIV gene expression is the recruitment of Tat protein to the transcription machinery after its binding to the RNA response element TAR. Starting from a pool of 3.2 ؋ 10 6 individual chemical entities, we were able to select a hybrid peptoid͞peptide oligomer of 9 residues (CGP64222) that was able to block the formation of the Tat͞TAR RNA complex in vitro at nanomolar concentrations. NMR studies demonstrated that the compound binds similarly to polypeptides derived from the Tat protein and induces a conformational change in TAR RNA at the Tat
A tripeptoid library was synthesized using 69 different primary amines in initially 69 individual reactions by the mix and split approach. The resulting library consisted of 328,509 (69(3)) single compounds, divided in 69 subpools each containing 4,761 entities. The 69 subpools were tested in two binding assays, one for alpha-MSH (alpha-melanotropin) and one for GRP (gastrin-releasing peptide)/bombesin. The sublibraries with the highest affinity to the MSH receptor (i.e. melanocortin type 1 or MC1 receptor) and, respectively, the GRP-preferring bombesin receptor were identified by an iterative process. Individual tripeptoids with good binding activity were resynthesized, analyzed and their dissociation constants and biological activity determined. The KD of the most potent MC1 receptor ligand was 1.58 mumol/l and that of the GRP-preferring bombesin receptor 3.40 mumol/l. Extension of this latter tripeptoid structure whose KD value increased to 280 nmol/l. A similar increase in activity was not observed with the most potent MSH tripeptoid ligand when extended by one residue, but a compound suitable for radioiodination and lacking the N-terminal amino group had a slightly higher binding activity than the tripeptoids (KD approximately 850 nmol/l). These results demonstrate that testing a peptoid library containing 328,509 single compounds led to the successful identification of new ligands for both the MC1 receptor as well as the GRP-preferring bombesin receptor.
SummaryXanthines represent a new, versatile scaffold for combinatorial chemistry. A five-step solid-phase synthesis of xanthine derivatives is described which includes alkylations, a nucleophilic displacement reaction at a heterocycle and a ring closure reaction by condensation of a nitroso function with an activated methylene group. The selected reaction sequence allows the production of a highly diverse small-molecule combinatorial compound library.
A strategy for high-throughput evaluation of combinatorial compound libraries is reported, which circumvents the necessity to test complex mixtures. The method is based on a new combination of protecting groups, solid-phase linker and tags. The bulk of the library first undergoes a binding assay with the components grafted on beads. A selection of beads carrying strong ligands is stripped from the labelled target and distributed into microvessels. The ligands are cleaved and rinsed into microeluates. Subsequently, a more detailed characterization with a functional assay in solution determines the best performers, which are identified through the peptidic tag left behind on the corresponding mother bead.
Our approach to achieve a partial synthesis of methanopterin (1) started from 6-acetyl-O 4 -isopropyl-7-methylpterin (20) which was obtained either by condensation from 6-isopropoxypyrimidine-2,4,5triamine (19) and pentane-2,3,4-trione (6) or from 6-isopropoxy-5-nitrosopyrimidine-2,4-diamine (21) and pentane-2,4-dione (¼ acetylacetone; 22) (Scheme 2). NaBH 4 reduction of 20 led to 6-(1hydroxyethyl)-O 4 -isopropyl-7-methylpterin (23) which was converted into the corresponding 6-(1chloroethyl) and 6-(1-bromoethyl) derivatives 24 and 25. A series of nucleophilic displacement reactions in the side chain and at position 4 were performed as model reactions to give 26 -29, 32 -35, and 39 -41. Hydrolysis of the substituents at C(4) led to the corresponding pterin derivatives 30, 31, 36 -38, and 42. Analogously, 25 reacted with 1-(4-aminophenyl)-1-deoxy-2,3: 4,5-di-O-isopropylidene-d-ribitol (43), prepared from N-(4-bromophenyl)benzamide (47) via 49 and 50 to give 1-{4-{{1-[2-amino-7-methyl-4-(1methylethoxy)pteridin-6-yl]ethyl}amino}phenyl}-1-deoxy-d-ribitol (44) in 62% yield (Scheme 3). Acid cleavage of the isopropylidene groups at room temperature led to 45 and on boiling to 1-{4-{[1-(2-amino-3,4-dihydro-7-methyl-4-oxopteridin-6-yl)ethyl]amino}phenyl}-1-deoxy-d-ribitol (46). The next step, however, attachment of the ribofuranosyl moiety with 55 or 56 to the terminal 1-deoxy-d-ribitol OH group could not been achieved. The second component, bis(4-nitrobenzyl) 2-{[(2-cyanoethoxy)(diisopropylamino)phosphino]oxy}pentanedioate (61), to built-up methanopterin (1) was synthesized from 2hydroxypentanedioic acid (59) and worked well in another model reaction on phosphitylation with N 6benzoyl-2',3'-O-isopropylideneadenosine and oxidation to give 62 (Scheme 6).
SummaryThe synthesis of 6,7, Ptr) (9) starting from 2,4-diamino-6-hydroxymethylpteridine (I) is described. A special protecting group strategy had to be applied to perform the structural modifications. The introductio n of the azido function was achieved by a Mitsunobu reaction converting the N 2 -N ,N -dimethylaminomethylen-6 -hydroxymethyl-5-pivaloyl-5,6,7 ,8-tetrahydropterin (7) with diphenylphosphoryl azide into the corresponding 6-azidomethyl derivative (8). The newly synthesized compounds have been characterized by elemental analyses, UV, IR and IH-NMR spectra. The photolysis of 6-AzmH 4 Ptr (9) and its inactivation effect upon aromatic amino acid mono oxygenases has been studied
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