Am. third NOS isoform, the inducible NOS (iNOS), is usually calcium-independent, not usually expressed under physiological conditions, and is induced by endotoxin and/or cytokines, such as lipopolysaccharide (LPS), interleukin-1 (IL-1), tumor necrosis factor (TNF-) and interferon- (IFN). Once induced, iNOS produces high and sustained levels of NO. The overexpression of iNOS, and the producing excessive production of NO which results in cellular cytotoxicity and tissue damage, has been implicated in the pathogenesis of a number of inflammatory diseases, such as rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, multiple sclerosis and asthma [3-8]. Therefore, iNOS inhibitors may find power for the treatment of these diseases. Because of the importance of the constitutive forms in normal physiology, high selectivity for iNOS is usually advantageous to avoid blocking the basic homeostatic functions of the eNOS and nNOS isoforms. The three NOS isoforms differ in their location and function, but are comparable in that they are only active in the dimeric form [9-1]. Preventing the dimerization of inactive NOS monomers into active homodimers has emerged as a novel pharmacological strategy to develop isoform-selective NOS inhibitors. Highly potent and selective imidazopyri-midine-based iNOS dimerization inhibitors, exemplified by compounds 1 and 2 (Fig. ?11), were discovered recently. These compounds significantly decreased levels of NO production [10, 11]. Based on the crystal structure of 2 bound to murine iNOS monomeric oxygenase domain name (iNOS 114) [12-14], the imidazole group binds to the heme, while the benzodioxolane group fits closely between residues in the iNOS monomer active site and the pyrimidine ring, resulting in a U-shaped conformation of the molecule in its active site. This prevents Glu377 of helix 7A from occupying the position that leads to dimer formation. Based on this binding mode, new inhibitors using alternate linkers such as hydroxyethylamine, hydroxypiperidine, hydroxypyrimidine, etc, to connect the benzodioxolane and imidazole moieties have been reported [12-14]. As part of our research program on new chemical classes of iNOS inhibitors, we designed and synthesized a series of imidazopyrimidine derivatives with the general formula I (Fig. ?11) as isosteric analogs of 1 1 and 2. In the structure of these compounds, the central piperazine and pyrrolidine heterocycle themes in 1 [10, 11] and 2 [11] were replaced with cycloalkenyl, cycloalkyl and phenyl rings. Some of these new agents were potent iNOS dimerization inhibitors in cell-based iNOS assays. Open in a separate windows Fig. (1) In compounds 1 and 2, the piperazine and pyrrolidine heterocycles are connected to the pyrimidine ring analogs 5 and 8 by treatment with DBU in refluxing benzene. The synthesis of the target compound 9 was also straightforward. The reaction of chloropyrimidine 19 with 2-ethoxycarbonylphenylzinc bromide in the presence of Pd(PPh3)4 under Negishi coupling condition afforded the coupled product 26 in 84% yield. The ester 26 was then converted to the target compound 9 in a similar manner as for the synthesis of 3 and 6 from 24a,b. Next, we made various modifications on the molecule 9 at the tether linking the middle phenyl ring to the benzodioxolane group to further investigate the SAR of this new chemical series. The compounds 10-16 were prepared according to Scheme 2. 2-Iodophenylacetic acid (28) was condensed with piperonylamine using TBTU as coupling reagent to provide the amide 29, which was then coupled with the organotin derivative 23 using Pd(CH3CN)2Cl2 as catalyst under microwave conditions to yield 10. Similar to the preparation of ester 26, Negishi coupling of 19 with 2-cyanophenylzinc bromide furnished 30 in excellent yield. The cyano derivative 30 was then converted to the primary amine 31 by hydrogenation. Compound 31 was then converted to the amide 11 using the above-mentioned TBTU coupling method, and converted to the urea analog 12 by condensation with 3,4-(methylenedioxy) phenyl isocyanate. Stille coupling of bromide 33 and 35 with 23 using the same reaction condition as for 24a,b yielded amide 13, and sulfonamide 14, respectively. Compounds 15 and 16 were prepared according to the same reaction conditions described above for the synthesis of 9. The imidazopyrimidines 3-16 were evaluated for their abilities to inhibit cytokine-mediated induction of iNOS activity in DLD-1 cells (Tables ?11 and ?22). The initial strategy consisted of replacing the central pyrrolidine group of 2 with a cyclopentene moiety. This modification led to a 10-fold decrease in the iNOS potency. Hydrogenation of the double bond functionality of 3 provided compound 4, which displayed similar iNOS.Chem. of NO. The overexpression of iNOS, and the resulting excessive production of NO which results in cellular cytotoxicity and tissue damage, has been implicated in the pathogenesis of a number of inflammatory diseases, such as rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, multiple sclerosis and asthma [3-8]. Therefore, iNOS inhibitors may find utility for the treatment of these diseases. Because of LDN-214117 the importance of the constitutive forms in normal physiology, high selectivity for iNOS is advantageous to avoid blocking the basic homeostatic functions of the eNOS and nNOS isoforms. The three NOS isoforms differ in their location and function, but are similar in that they are only active in the dimeric form [9-1]. Preventing the dimerization of inactive NOS monomers into active homodimers has emerged as a novel pharmacological strategy to develop isoform-selective NOS inhibitors. Highly potent and selective imidazopyri-midine-based iNOS dimerization inhibitors, exemplified by compounds 1 and 2 (Fig. ?11), were discovered recently. These compounds significantly decreased levels of NO production [10, 11]. Based on the crystal structure of 2 bound to murine iNOS monomeric oxygenase domain (iNOS 114) [12-14], the imidazole group binds to the heme, while the benzodioxolane group fits closely between residues in the iNOS monomer active site and the pyrimidine ring, resulting in a U-shaped conformation of the molecule in its active site. This prevents Glu377 of helix 7A from occupying the position that leads to dimer formation. Based on this binding mode, new inhibitors using alternative linkers such as hydroxyethylamine, hydroxypiperidine, hydroxypyrimidine, etc, to connect the benzodioxolane and imidazole moieties have been reported [12-14]. As part of our research program on new chemical classes of iNOS inhibitors, we designed and synthesized a series of imidazopyrimidine derivatives with the general formula I (Fig. ?11) as isosteric analogs of 1 1 and 2. In the structure of these compounds, the central piperazine and pyrrolidine heterocycle templates in 1 [10, 11] and 2 [11] were replaced with cycloalkenyl, cycloalkyl and phenyl rings. Some of these new agents were potent iNOS dimerization inhibitors in cell-based iNOS assays. Open in a separate window Fig. (1) In compounds 1 and 2, the piperazine and pyrrolidine heterocycles are connected to the pyrimidine ring analogs 5 and 8 by treatment with DBU in refluxing benzene. The synthesis of the target compound 9 was also straightforward. The reaction of chloropyrimidine 19 with 2-ethoxycarbonylphenylzinc bromide in the presence of Pd(PPh3)4 under Negishi coupling condition afforded the coupled product 26 in 84% yield. The ester 26 was then converted to the target compound 9 in a similar manner as for the synthesis of 3 and 6 from 24a,b. Next, we made various modifications on the molecule 9 at the tether linking the middle phenyl ring to the benzodioxolane group to further investigate the SAR of this new chemical series. The compounds 10-16 were prepared according to Scheme 2. 2-Iodophenylacetic acid (28) was condensed with piperonylamine using TBTU as coupling reagent to provide the amide 29, which was then coupled with the organotin derivative 23 using Pd(CH3CN)2Cl2 as catalyst under microwave conditions to yield 10. Similar to the preparation of ester 26, Negishi coupling of 19 with 2-cyanophenylzinc bromide furnished 30 in excellent yield. The cyano derivative 30 was then converted to the primary amine 31 by hydrogenation. Compound 31 was then converted to the amide 11 using the above-mentioned TBTU coupling method, and converted to the urea analog 12 by condensation with 3,4-(methylenedioxy) phenyl isocyanate. Stille coupling of bromide 33 and 35 with 23 using the same reaction condition as for 24a,b yielded amide 13, and sulfonamide 14, respectively. Compounds 15 and 16 were prepared according to the same reaction conditions described above for the synthesis of 9. The imidazopyrimidines 3-16 were evaluated for their abilities to inhibit cytokine-mediated induction of iNOS activity in DLD-1 cells.2002:1880C1882. interleukin-1 (IL-1), tumor necrosis factor IgG2b Isotype Control antibody (PE) (TNF-) and interferon- (IFN). Once induced, iNOS produces high and sustained levels of NO. The overexpression of iNOS, and the resulting excessive production of NO which results in cellular cytotoxicity and tissue damage, has been implicated in the pathogenesis of a number of inflammatory diseases, such as rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, multiple sclerosis and asthma [3-8]. Therefore, iNOS inhibitors may find utility for the treatment of these diseases. Because of the importance of the constitutive forms in normal physiology, high selectivity for iNOS is advantageous to avoid blocking the basic homeostatic functions of the eNOS and nNOS isoforms. The three NOS isoforms differ LDN-214117 in their location and function, but are similar in that they are only active in the dimeric form [9-1]. Preventing the dimerization of inactive NOS monomers into active homodimers has emerged as a novel pharmacological strategy to develop isoform-selective NOS inhibitors. Highly potent and selective imidazopyri-midine-based iNOS dimerization inhibitors, exemplified by compounds 1 and 2 (Fig. ?11), were discovered recently. LDN-214117 These compounds significantly decreased levels of NO production [10, 11]. Based on the crystal structure of 2 bound to murine iNOS monomeric oxygenase domain (iNOS 114) [12-14], the imidazole group binds to the heme, while the benzodioxolane group fits closely between residues in the iNOS monomer active site and the pyrimidine ring, resulting in a U-shaped conformation of the molecule in its active site. This prevents Glu377 of helix 7A from occupying the position that leads to dimer formation. Based on this binding mode, fresh inhibitors using alternate linkers such as hydroxyethylamine, hydroxypiperidine, hydroxypyrimidine, etc, to connect the benzodioxolane and imidazole moieties have been reported [12-14]. As part of our research system on fresh chemical classes of iNOS inhibitors, we designed and synthesized a series of imidazopyrimidine derivatives with the general method I (Fig. ?11) while isosteric analogs of 1 1 and 2. In the structure of these compounds, the central piperazine and pyrrolidine heterocycle themes in 1 [10, 11] and 2 [11] were replaced with cycloalkenyl, cycloalkyl and phenyl rings. Some of these fresh agents were potent iNOS dimerization inhibitors in cell-based iNOS assays. Open in a separate windowpane Fig. (1) In compounds 1 and 2, the piperazine and pyrrolidine heterocycles are connected to the pyrimidine ring analogs 5 and 8 by treatment with DBU in refluxing benzene. The synthesis of the LDN-214117 target compound 9 was also straightforward. The reaction of chloropyrimidine 19 with 2-ethoxycarbonylphenylzinc bromide in the presence of Pd(PPh3)4 under Negishi coupling condition afforded the coupled product 26 in 84% yield. The ester 26 was then converted to the prospective compound 9 in a similar manner as for the synthesis of 3 and 6 from 24a,b. Next, we made various modifications within the molecule 9 in the tether linking the middle phenyl ring to the benzodioxolane group to further investigate the SAR of this fresh chemical series. The compounds 10-16 were prepared according to Plan 2. 2-Iodophenylacetic acid (28) was condensed with piperonylamine using TBTU as coupling reagent to provide the amide 29, which was then coupled with the organotin derivative 23 using Pd(CH3CN)2Cl2 as catalyst under microwave conditions to yield 10. Similar to the preparation of ester 26, Negishi coupling of 19 with 2-cyanophenylzinc bromide furnished 30 in superb yield. The cyano derivative 30 was then converted to the primary amine 31 by hydrogenation. Compound 31 was then converted to the amide 11 using the above-mentioned TBTU coupling method, and converted to the urea analog 12 by condensation with 3,4-(methylenedioxy) phenyl isocyanate. Stille coupling of bromide 33 and 35 with 23 using the same reaction condition as for 24a,b yielded amide 13, and sulfonamide 14, respectively. Compounds 15 and 16 were prepared according to the same reaction conditions explained above for the synthesis of 9. The imidazopyrimidines 3-16 were evaluated for his or her capabilities to inhibit cytokine-mediated induction of iNOS activity in DLD-1 cells (Furniture ?11 and ?22). The initial strategy consisted of replacing the central pyrrolidine group of 2 with.