(Settore Innovazione Tecnologie Informatiche) and C

(Settore Innovazione Tecnologie Informatiche) and C.I.M. a separate window Scheme 3 Reagents and conditions: (a) 3-phenylphenol, CH2Cl2, DCC, DMAP, r.t., 18 h. Isocyanates 3b-f,h,i,k-n,w were commercially available. The synthesis of isocyanates 3g,o-v,y,z (Scheme 4) was performed via Curtius rearrangement of the opportune acylazide (10), prepared by reacting the appropriate carboxylic acid (9) with diphenylphosphorylazide (DPPA) (3g,q,r,t-v,y,z) or with oxalylchloride and sodium azide (NaN3) (3o,p,s). Open in a separate window Scheme 4 Reagents and conditions: (a) toluene, TEA, DPPA, 0.5 h (3g,q,r,t-v,y,z) or CH2Cl2, DMF, (COCl)2, 0.5 h then NaN3 in H2O, 0.5 h, 0-5 C (3o,p,s); (b) reflux, 0.5 h. Carboxylic acid 9t was obtained by reaction of ethyl 7-bromoheptanoate (11t) with morpholine and subsequent hydrolysis of ester 12t (Scheme 5). Carboxylic acids 9u and 9v were synthesized by a catalytic hydrogenation of the respective intermediates 15u and 15v, obtained by a Wittig reaction of the opportune aldehyde and the phosphonium salts 14u and 14v, in presence of lithium hexamethyldisylazane (LHMDS).52 Concerning the latter compounds, 14v was commercially available while 14u was prepared by reaction of 8-bromooctanoic acid (13u) and triphenylphosphine (TPP) (Scheme 6). Open in a separate window Scheme 5 Reagents and conditions: (a) toluene, morpholine, reflux, 0.5 h; (b) H2O, reflux, 72 h. Open in a separate window Scheme 6 Reagents and conditions: (a) CH3CN, TPP, 48 h; (b) THF, LHMDS 1M in THF, R-C(O)H, 24 h; (c) AcOEt, H2, Pd/C 10%, 3 h. Results and Discussion We measured FAAH activity in rat brain membranes, using [3H]anandamide as a substrate. The IC50 values for compounds 4a-z, 6, and 8 are reported in Tables 1a and 1b, along with that of the reference compound 2. Table 1a Inhibitory Potencies (IC50) of Tested Compounds 2 and 4a-z on FAAH Activity. = 0.95), poor correlations were found between pIC50 and lipophilicity, as confirmed by the regression Equations 1 and 2. procedure. Docking was followed by energy minimization of the complexes, allowing movements of the side chain of the residues at maximum 8 ? from the inhibitor, using MMFF94s force field to a gradient of 0.1 kcal/(mol?). In the first orientation (orientation A), all the docked ligands JMV 390-1 place their biphenyl fragment in the acyl binding pocket (also occupied by the acyl chain of covalently bound MAP in the 1MT5 pdb structure) and the substituent at the nitrogen atom in the cytosolic access channel. In the second one (orientation B), the em meta /em -biphenyl fragment lies in the cytosolic access channel, whereas the carbamate nitrogen substituent occupies the acyl binding pocket. In both orientations, the carbonyl oxygen closely interacts with the NH groups of the oxyanion hole, whereas the NH of the carbamate forms a hydrogen bond with the backbone oxygen of Met191. At JMV 390-1 the end of this preliminary optimization cycle, the energy-minimized complexes were JMV 390-1 transported to the LIAISON package for subsequent SGB-LIE calculations. (e) LIE Calculation To compute the interaction terms required in the LIE model, a second energy minimization was performed with LIAISON, applying OPLS2001 force field,67 in combination with the Surface Generalized Born continuum model. The Truncated Newton (TN) technique was applied to an energy gradient of 0.01 kcal/(mol?) using a residue Rabbit Polyclonal to MNT based cut-off distance of 10 ? for the free ligands, and to 0.05 kcal/(mol?) for FAAH-inhibitor complexes, with a residue based cut-off distance of 15 ?. During energy minimization of the complexes, only the side chains of the protein residues within 20 ? from the catalytic Ser241 were allowed to move. The JMV 390-1 protein backbone was maintained with fixed geometry to preserve the structure of the binding site. The energy terms reported.