Transient receptor potential vanilloid type 1 (TRPV1) is a ligand-gated ion channel that functions as an integrator of multiple pain stimuli including heat, acid, capsaicin and a variety of putative endogenous lipid ligands. TRPV1 antagonists have been shown to decrease inflammatory pain in animal models and to produce limited hyperthermia at analgesic doses. Here, we report that ABT-102, which is a potent and selective TRPV1 antagonist, is effective in blocking nociception in rodent models of inflammatory, post-operative, osteoarthritic, and bone cancer pain. ABT-102 decreased both spontaneous pain behaviors and those evoked by thermal and mechanical stimuli in these models. Moreover, we have found that repeated administration of ABT-102 for 5-12 days increased its analgesic activity in models of post-operative, osteoarthritic, and bone cancer pain without an associated accumulation of ABT-102 concentration in plasma or brain. Similar effects were also observed with a structurally distinct TRPV1 antagonist, A-993610. Although a single dose of ABT-102 produced a self-limiting increase in core body temperature that remained in the normal range, the hyperthermic effects of ABT-102 effectively tolerated following twice-daily dosing for 2 days. Therefore, the present data demonstrate that, following repeated administration, the analgesic activity of TRPV1 receptor antagonists is enhanced, while the associated hyperthermic effects are attenuated. The analgesic efficacy of ABT-102 supports its advancement into clinical studies.

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High-throughput screening (HTS) identified benzothiazole analogue 3 as a potent fatty acid amide hydrolase (FAAH) inhibitor. Structure-activity relationship (SAR) studies indicated that the sulfonyl group, the piperidine ring and benzothiazole were the key components to their activity, with 16j being the most potent analogue in this series. Time-dependent preincubation study of compound 3 was consistent with it being a reversible inhibitor. Activity-based protein-profiling (ABPP) evaluation of 3 in rat tissues revealed that it had exceptional selectivity and no off-target activity with respect to other serine hydrolases. Molecular shape overlay of 3 with a known FAAH inhibitor indicated that these compounds might act as transition-state analogues, forming putative hydrogen bonds with catalytic residues and mimicking the charge distribution of the tetrahedral transition state. The modeling study also indicated that hydrophobic interactions of the benzothiazole ring with the enzyme contributed to its extraordinary potency. These compounds may provide useful tools for the study of FAAH and the endocannabinoid system.

We have isolated and sequenced cDNA clones encoding the human U1-70K snRNP protein, and have mapped this locus (U1AP1) to human chromosome 19. The gene produces two size classes of RNA, a major 1.7-kb RNA and a minor 3.9-kb RNA. The 1.7-kb species appears to be the functional mRNA; the role of the 3.9-kb RNA, which extends further in the 5' direction, is unclear. The actual size of the hU1-70K protein is probably 52 kd, rather than 70 kd. The protein contains three regions similar to known nucleic acid-binding proteins, and it binds RNA in an in vitro assay. Comparison of the cDNA sequences indicates that there are multiple subclasses of mRNA that arise by alternative pre-mRNA splicing of at least four alternative exon segments. This suggests that multiple forms of the hU1-70K protein may exist, possibly with different functions in vivo.

We have studied the interaction of two of the Ul small nuclear ribonucleoprotein (snRNP)-specific proteins, Ul-70K and U1-A, with Ul small nuclear RNA (snRNA). The U1-70K protein is a Ul-specific RNA-binding protein. Deletion and mutation analyses of a ,-galactosidase/Ul-70K partial fusion protein indicated that the central portion of the protein, including the RNP sequence domain, is both necessary and sufficient for specific Ul snRNA binding in vitro. The highly conserved eight-amino-acid RNP consensus sequence was found to be essential for binding. Deletion and mutation analyses of Ul snRNA showed that both the Ul-70K fusion protein and the native HeLa U1-70K protein bound directly to loop I of Ul snRNA. Binding was sequence specific, requiring 8 of the 10 bases in the loop. The U1-A snRNP protein also interacted specifically with Ul snRNA, principally with stem-loop II.The splicing of eucaryotic mRNA precursors (pre-mRNAs) involves a group of small nuclear ribonucleoprotein particles (U snRNPs) that are subassemblies of a larger ribonucleoprotein complex, the spliceosome. At least four different U snRNPs are required for splicing. Ul snRNP mediates recognition of 5' splice sites, and U2, U4/6, and U5 snRNPs mediate recognition of the 3' ends of intervening sequences (reviewed in references 10 and 27). An understanding of the mechanisms by which the various U snRNPs interact with pre-mRNAs, with each other, and with other components of the spliceosome will ultimately require an analysis of the individual snRNP components and their functions.Human Ul snRNP consists of the 164-nucleotide Ul small nuclear RNA (snRNA), a group of at least seven core (Sm) proteins common to all U snRNPs, and three specific proteins: U1-70K, U1-A, and U1-C (5). In the absence of an snRNA, the Sm core proteins associate to form a 6S particle (9). It seems likely that the core snRNP proteins serve a scaffold function, organizing snRNP architecture, and that the specific proteins mediate the functions of Ul snRNP in pre-mRNA splicing.The recent isolation of cDNAs encoding all three of the human Ul-specific snRNP proteins (24-26, 29, 37) has provided considerable structural information on these polypeptides. The U1-70K and U1-A proteins both contain sequence motifs that occur in several known nucleic acidbinding proteins, including the RNP sequence domain (1, 28; reviewed in reference 7) which is apparently involved in RNA binding (6,19). The RNP sequence domain contains within it a short, even more highly conserved segment, the RNP consensus sequence (rY'GlyPheGlyPhelleXPh¾e; reviewed in reference 7). In addition, the U1-70K protein contains two arginine-rich tracts reminiscent of those in protamines, sperm histones, and chicken galline, proteins that bind DNA * Corresponding author. t Paper 3044 from the