ABSTRACT
The P4 region of a series of oxamyl dipeptide caspase inhibitors was optimized by the combination of anti-apoptotic activity in the Jurkat/Fas (JFas) cellular assay and membrane permeability in the PAMPA assay. Two highly potent anti-apoptotic agents with moderate membrane permeability, 29 and 36, showed strong in vivo efficacy in a murine model of alpha-Fas-induced liver injury.
Subject(s)
Caspase Inhibitors , Cysteine Proteinase Inhibitors/chemical synthesis , Liver Diseases/drug therapy , Animals , Apoptosis/drug effects , Cell Membrane Permeability/drug effects , Cysteine Proteinase Inhibitors/pharmacology , Humans , Jurkat Cells , Mice , Structure-Activity Relationship , fas ReceptorABSTRACT
(-)-6-[2-[4-(3-Fluorophenyl)-4-hydroxy-1-piperidinyl]-1-hydroxyethyl]-3,4-dihydro-2(1H)-quinolinone was identified as an orally active NR2B-subunit selective N-methyl-d-aspartate (NMDA) receptor antagonist. It has very high selectivity for NR2B subunits containing NMDA receptors versus the HERG-channel inhibition (therapeutic index=4200 vs NR2B binding IC(50)). This compound has improved pharmacokinetic properties compared to the prototype CP-101,606.
Subject(s)
N-Methylaspartate/antagonists & inhibitors , N-Methylaspartate/metabolism , Pain , Piperidines/chemistry , Quinolones/chemistry , Receptors, N-Methyl-D-Aspartate/metabolism , Animals , Cytochrome P-450 CYP2D6/metabolism , Cytochrome P-450 CYP2D6 Inhibitors , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Ether-A-Go-Go Potassium Channels/metabolism , Inhibitory Concentration 50 , Molecular Structure , Pain/drug therapy , Piperidines/pharmacology , Quinolones/pharmacology , Rats , Structure-Activity RelationshipABSTRACT
A rare beta-thalassemia mutation at the splicing junction [namely, G-->C in intervening sequence (IVS) I-1] was found in a Japanese family. The proband and his mother were heterozygous for the mutation. Analysis of mRNA extracted from the reticulocyte-rich fraction obtained from the proband's mother revealed that the mutant beta-globin gene did not produce any detectable, stable mRNA including exon 1 and exon 2, since the polymorphism in exon 1 on her mutant gene was not detected in the RT-PCR products.
Subject(s)
Globins/genetics , Point Mutation , RNA Splice Sites/genetics , beta-Thalassemia/genetics , Adult , Child, Preschool , DNA Mutational Analysis , Exons/genetics , Female , Humans , Japan , Male , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
To examine the role of the fibrinogen gamma chain in the assembly and secretion of this multichain protein, we synthesized a series of fibrinogen variants with truncated gamma chains, terminating between residues gamma379 and the C-terminus, gamma411. The variant fibrinogens were synthesized from altered gamma-chain complementary DNAs in cultured Chinese hamster ovary cells. Immunoassays of the culture media demonstrated that only those variants with gamma chain longer than 386 residues were secreted and that the concentration of fibrinogen decreased with the length of the gamma chain, from 1.4 microg/mL for normal fibrinogen to 0.39 microg/mL for gamma 387 fibrinogen. Immunoassays of cell lysates showed that all variant gamma chains were synthesized, although the levels varied significantly. For variants longer than 386 residues, levels decreased with length but remained near normal. In contrast, expression of the 4 variants with 386 residues or less was about 20-fold reduced. Quantitative reverse transcription-polymerase chain reaction demonstrated that the gamma-chain messenger RNA level was independent from chain length. Western blot analyses showed that lysates expressing variants with 387 residues or more contained species comparable to the known intermediates in fibrinogen assembly, including half-molecules. For shorter variants, these intermediates were not evident. We conclude that residues near the C-terminus of the gamma chain are essential for fibrinogen assembly, and more specifically, that gamma387 is critical. We propose that the loss of residue gamma387 destabilized the structure of gamma chain, preventing assembly of alphagamma and betagamma dimers, essential intermediates in the assembly of normal fibrinogen.
Subject(s)
Fibrinogen/biosynthesis , Fibrinogen/chemistry , Isoleucine/physiology , Amino Acid Sequence , Animals , CHO Cells , Cricetinae , Fibrinogen/genetics , Fibrinogen/metabolism , Humans , Models, Molecular , Molecular Sequence Data , Protein Structure, Tertiary , Protein Subunits , RNA, Messenger/biosynthesis , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , Sequence Deletion , Sequence Homology, Amino AcidABSTRACT
A novel preparation of (+)-methyl pederate (4), a key intermediate in syntheses of mycalamides (1), marine natural products from a New Zealand sponge of the genus Mycale, is described. The key step involves palladium-catalyzed intramolecular allylic alkylation of the carbonate 21, derived from (+)-(4R,5R,E)-5-(tert-butyldimethylsiloxy)-4-methyl-2-hexenol (13), yielding lactones 5 in 87% yield. Demethoxycarbonylation of the cyclization products 5 and further functional group transformations led to (+)-methyl pederate (4).
