Bertorelli, R

Burnett, Da

Greenlee, Wj

Grilli, Mariagrazia

Heimark, L

Li, C

Lozza, G

Pramanik, B

Qiang, L

Reggiani, A.

Sasikumar, Tk

English

SASIKUMAR TK

QIANG L

BURNETT DA

GREENLEE WJ

LI C

HEIMARK L

PRAMANIK B

GRILLI, Mariagrazia

BERTORELLI R

LOZZA G

REGGIANI A.

Archivio Istituzionale della Ricerca- Università del Piemonte Orientale

Bioorganic & Medicinal Chemistry Letters 19 (2009) 3199–3203Contents lists available at ScienceDirectBioorganic & Medicinal Chemistry Lettersjournal homepage: www.elsevier .com/ locate/bmclTricyclic thienopyridine–pyrimidones/thienopyrimidine–pyrimidonesas orally efficacious mGluR1 antagonists for neuropathic painT. K. Sasikumar a,*, Li Qiang a, Duane A. Burnett a, William J. Greenlee a, Cheng Li a, Larry Heimark a,Birendra Pramanik a, Mariagrazia Grilli b, Rosalia Bertorelli b, Gianluca Lozza b, Angelo Reggiani ba Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USAb Schering-Plough Research Institute, San Raffaele Science Park, Via Olgettina, 58, 20132 Milan, Italya r t i c l e i n f o a b s t r a c tArticle history:Received 12 March 2009Revised 21 April 2009Accepted 23 April 2009Available online 3 May 2009Keywords:Tricyclic thienopyridine–pyrimidoneThienopyrimidine–pyrimidoneTriazafluorenonemGluR1 antagonistNeuropathic pain0960-894X/$ - see front matter  2009 Published bydoi:10.1016/j.bmcl.2009.04.104* Corresponding author. Tel.: +1 908 740 4373; faxE-mail addresses: sasikumartk@yahoo.com,spcorp.com (T.K. Sasikumar).Introduction of small unsaturated alkylamino groups at the 4-position of the A-ring of the tricyclic frame-work (triazafluorenone) afforded extremely potent and selective mGluR1 antagonists with desirableproperties. Compounds 11q and 11s are active in the SNL pain model with ED50s 3.3 and 6.4 mg/kgrespectively. Metabolic outcome of propargyl amino moiety was studied. 2009 Published by Elsevier Ltd.ONOOOHOOHN NNONCN OOO(-)-CPCCOEt EM-TBPCGlutamate is the major excitatory neurotransmitter in the cen-tral nervous system and mediates its actions via activation of bothionotropic and metabotropic receptor families. The metabotropicglutamate receptors (mGluRs) form a family of eight subtypes(mGlu1 to mGlu8) and are assigned to three groups based on theirstructure, coupling to effector mechanisms and pharmacology.Group I mGluRs (mGluR1 and mGluR5) are post synaptic receptorswhile Group II (mGluR2 and mGluR3) and Group III (mGluR6,mGluR7 and mGluR8) are located presynaptically. It has beenfound that the Group I mGluRs play key roles in the central sensi-tization of pain and other neurologic disorders.1–6 Over the lastdecade, there have been tremendous advances in the developmentof small molecules that selectively activate or inhibit specific mGlureceptor subtypes. Given below are some of the recent examplesfrom literature (Fig. 1).7–11Compound 1was identified from our high throughput screeningas a potent mGluR1 antagonist that was active in an in vivo modelfor pain (rat spinal nerve ligation, SNL).12 A recent paper describingthe SAR around the tricyclic frame work was published.13 Theyhave not disclosed any unsaturated alkyl substitutions on the A-ring of the tricyclic structure. Our attempt to replace the dimethylamino moiety was exceptionally fruitful.14 We found that smallunsaturated alkyls gave extremely potent mGluR1 antagonists.Elsevier Ltd.: +1 908 740 7164.thavalakulamgar.sasikumar@SAR studies involving A-ring and N-aryl modifications and somein vivo results are discussed in this Letter (Fig. 2).Compounds of this type were synthesized according to Scheme1. Commercially available ethylcyano malonate was condensedwith dimethylformamide–dimethylacetal (DMF–DMA) followedby acid catalyzed cyclization gave pyridine derivative 4 in goodyields. Upon reaction with POCl3 on 4 gave 5 which was cyclizedto a common intermediate 6. Heating a solution of 6 in DMF withBAY36-7620 JNJ16259685Figure 1. Noncompetitive antagonists of mGluR1.SXYNNNOR1NR2R3SNNNONR1 = ArylR2 = unsaturated alkylsR3 = H or MeX = C or N; Y = C or N1OMeABCFigure 2. General SAR plans.SNNONH2OOMeSNNNNONR1R2R312 13 (a-z)a-cScheme 2. Reagents and conditions: (a) DMF–DMA, DMF, 100 C; (b) 4-Cl-aniline(R1 = Cl), HOAc, toluene, 110 C; (c) R2R3NH, DMSO.Table 1mGluR1 receptor binding for compounds 1, 11a–iSNNNOR1N R211 (a-i)Compd R1 R2 h-mGluR1 IC50a (nM) r-mGluR1 Kia (nM)1 9.5 7.911a 4-MeO-Ph H 2 1.111b 4-Me-Ph H 11 2.711c 4-Cl-Ph H 60 1.111d 4-Br-Ph H 12.6 0.711e 4-F-3-MeO-Ph H 6.8 1411f 4-F-Ph H 4.4 7.711g 4-Cl-Ph Me 1.1 5.411h 4-MeO-Ph Me 6.9 4.111i 4-Br-Ph Me 66 3.2a The IC50 and Ki data are an average of at least three measurements, performedon human mGlu1/5 and rat mGlu1 receptors, respectively. The standard error was10%, and variability was less than twofold from assay to assay. h-mGluR5IC50 > 3 lM.3200 T. K. Sasikumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3199–3203DMF–DMA afforded compound 7 which was cyclized to the tricy-clic compound 8 in good yields. During this course of our studies,we found that heating a solution of 6 with desired amine in tri-ethylorthoformate in the presence of acetic acid gave the cyclizedcompound in very high yields.15 Demethylation of 8 followed bytriflation of the phenol afforded 10 in good yields. Intermediate10 was reacted with unsaturated alkylamines to afford the finalcompounds in excellent yields. We used allyl, methallyl and prop-argyl amines in this current SAR study. Similar chemistry was em-ployed in the construction of thienopyrimidine–pyrimidoneanalogs as shown in Scheme 2. The known amino ester 1216 wasconverted to the final targets in three steps with good overallyields. The final step is the displacement of methoxy group inthe pyrimidine nucleus with appropriate amine in polar solventssuch as DMSO to get compounds 13a–z.The mGluR1 inhibitory potencies of the newly synthesized tri-cyclic compounds are summarized in Table 1. Initially we turnedour attention to the allyl substitution on the A-ring of the pyridinenucleus. Human mGluR1 IC50 and rat Ki are shown throughout inthis manuscript. Allylamino compounds 11a–i are generally welltolerated as shown in Table 1. Compound 11a showed humanIC50 value of 2 nM with a rat Ki of 1.1 nM. Other substitution onthe aromatic ring such as 4-Me, 4-Br, and 4-F are very well toler-ated. However the 4-chloro substitution (compound 11c) gave onlymoderate activity on the human receptor but showed excellent po-tency in the rat assay. Similar results were obtained with the 4-Brderivative (11i). Disubstitution on the aromatic ring is well toler-ated in this scaffold. N-Methylated compounds such as 11g and11h are also tolerated. It has been found that all these tricyclicderivatives are inactive in the mGluR5 assay.Next, we studied the effect of substitution on the allyl group.Methallyl derivatives generally afforded active analogs with 2–3-fold less mGluR1 affinity as shown in Table 2. Disubstitution onthe aromatic ring (compounds 11n, 11o and 11p) generally de-creases the human inhibitory potency. Compounds 11k and 11lSNNNOOTfR1NOMNC CNEtONC CNMeON OHNOMeCNSNNNONR1R2R32 3 4 510a b cj11 (a-x)iScheme 1. Reagents and conditions: (a) DMF–DMA, MeOH, 80 C; (b) HOAc (80%), 130 C100 C; (f) 4-Cl-aniline (R1 = Cl), HOAc, toluene, 110 C; (g) 4-Cl-aniline (R1 = Cl), HOAc,exhibited subnanomolar activity in rat mGluR1 assay. Methallylsubstitution was also very effective in the pyrimidine series (Table3). Simple phenyl substitution (compound 13f)17 provided goodhuman affinity but a moderate rat Ki value. It was interesting tonote that the 4-pyridyl analog displayed poor activity at both thehuman and rat receptors. Unlike in the case of pyridine analogs,disubstitution in the pyrimidine series is very well tolerated. Forexample, compound 13l, showed human IC50 of 8.9 nM and rat Kiof 2.3 nM.Replacement of the dimethylamino group of our lead com-pound with propargylamine afforded an extremely potent com-pound 11q with human IC50 0f 0.9 nM and a rat Ki of 0.6 nM asSNNNOOMeR1SNONH2OOMeCleCN6 7dSNONOOMeNeSNNNOOHR1fh89g; (c) POCl3, Et3N, 100 C; (d) HSCH2CO2Me, NaOMe, DMF, 80 C; (e) DMF–DMA, DMF,CH(OEt)3, 160 C; (h) BBr3, CH2Cl2; (i) NPhTf2, CH2Cl2; (j) R2R3NH, DMSO.Table 2mGluR1 receptor binding for compounds 11j–pSNNNOR1NH11 j-pCompd R1 h-mGluR1IC50a (nM)r-mGluR1Kia (nM)11j 4-Cl-Ph 10.6 1.211k 4-Me-Ph 16.4 0.211l 4-MeO-Ph 10.4 0.411m 4-Br-Ph 43 1.511n 3-F-4-MeO-Ph 103 1311o 3-(2,3-Dihydrobenzo[b] [1,4]Dioxin-6-yl) 196 1311p 3-(Benzo[d][1,3]dioxol-5-yl) 241 14a The IC50 and Ki data are an average of at least three measurements, performedon human mGlu1/5 and rat mGlu1 receptors, respectively. The standard error was10%, and variability was less than twofold from assay to assay. h-mGluR5IC50 > 3 lM.Table 3mGluR1 receptor binding for compounds 13a–mSNNNNOR1NH13 (a-m)Compd R1 h-mGluR1IC50a (nM)r-mGluR1Kia (nM)13a 4-Cl-Ph 7.7 2.213b 4-Me-Ph 12.4 2.913c 4-MeO-Ph 9.6 7.513d 4-Br-Ph 14.5 3.813e 3-F-4-MeO-Ph 72 6813f Ph 4.6 2513g 4-Py 955 100013h 4-F-Ph 10.7 3813i 3-Cl-Ph 53 6213j 3-(Benzo[d]thiazol-5-yl) 16 4313k 3-(Benzo[d]thiazol-6-yl) 17 4913l 3-(Benzo[b]thiophen-5-yl) 8.9 2.313m 3-(2,3-Dihydrobenzo[b] [1,4]dioxin-6-yl) 33 64a The IC50 and Ki data are an average of at least three measurements, performedon human mGlu1/5 and rat mGlu1 receptors, respectively. The standard error was10%, and variability was less than twofold from assay to assay. h-mGluR5IC50 > 3 lM.Table 4mGluR1 receptor binding for compounds 11q–xSNNNOR1N R211 (q-x)Compd R1 R2 h-mGluR1 IC50a (nM) r-mGluR1 Kia (nM)11q 4-Cl-Ph H 0.9 0.611r 4-Me-Ph H 1.5 1.611s 4-MeO-Ph H 2.1 3.511t 4-Br-Ph H 1.8 0.711u 3-F-4-MeO-Ph H 12.5 2811v 4-Cl-Ph Me 3.8 1811w 4-MeO-Ph Me 10.8 2711x 4-Br-Ph Me 1.9 8a The IC50 and Ki data are an average of at least three measurements, performedon human mGlu1/5 and rat mGlu1 receptors, respectively. The standard error was10%, and variability was less than twofold from assay to assay. h-mGluR5IC50 > 3 lM.Table 5mGluR1 receptor binding for compounds 13n–zSNNNNOR1NH13 (n-z)Compd R1 h-mGluR1 IC50a(nM)r-mGluR1 Kia(nM)13n 4-Cl-Ph 3.0 4.413o 4-Me-Ph 3.9 8.313p 4-MeO-Ph 13 2513q 4-Br-Ph 6.8 2913r 3-F-4-MeO-Ph 212 7713s 4-F-Ph 8.2 3913t 2-F-4-MeO-Ph 85 3213u 3-(2,3-Dihydrobenzo[b][1,4]dioxin-6-yl)106 16013v 3-(Benzo[d][1,3]dioxol-5-yl) 137 99713w 3-(Benzo[d]thiazol-5-yl) 45 6513x 3-(Benzo[d]thiazol-6-yl) 209 10713y 3-(Benzofuran-5-yl) 21 6313z 3-(Benzo[b]thiophen-5-yl) 219 69a The IC50 and Ki data are an average of at least three measurements, performedon human mGlu1/5 and rat mGlu1 receptors, respectively. The standard error was10%, and variability was less than 2-fold from assay to assay. h-mGluR5 IC50 > 3 lM.T. K. Sasikumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3199–3203 3201shown in Table 4. 4-Me, 4-MeO, and 4-Br substitution on the N-arylgroup afforded similarly highly potent compounds. N-Methylationlowered affinity 4–5-fold in all cases except 4-BrPh derivative(11x).Similar results were obtained in the A-ring pyrimidine series asevidenced from the Table 5. Generally, mono substitution at thepara substitution of the N-aryl ring is generally well tolerated.Compounds 13n, 13o, 13q and 13s attained single digit nanomolarat the human mGluR1 assay. However disubstituted compoundssuch as 13x and 13z exhibited a large decrease in potency as evi-denced from Table 5.Having achieved excellent potency against mGluR1 receptors,we shifted our attention toward measuring the pharmacokineticsof these types of molecules. Data for representative compounds11q and 11s are shown in Table 6.Compounds 11q and 11s showed moderate rat AUC with highbrain to plasma ratio as demonstrated in Table 6. These com-pounds were inactive in the hERG assay. It has been found thatcompounds 11q and 11s were active in the in vivo pain model(rat spinal nerve ligation, SNL)12 with ED50 values of 3.3 and6.4 mg/kg, respectively, when dosed orally. The metabolic pathwayin rat for high clearance compound 11q was further investigatedusing microsome incubation. The metabolic details are shown inFigure 3. The metabolism of acetylenic compounds commonly usedin the formulation of pharmaceuticals have been investigated pre-viously using 13C NMR and mass spectrometry.19,20 Compound 11qTable 6PK Profile and in vivo activity of selected compoundsSNNNONHRParameters R = Cl (11q) R = OMe (11s)Human mGluR1 IC50 (nM) 0.9 2.1Rat mGluR1 Ki (nM) 0.6 3.5Human mGluR5 IC50 (nM) >3000 >3000Rat SNL ED5012 3.3 mg/kg po 6.4 mg/kg poCaco-2 permeability 430 nm/s 690 nm/sEfflux substrate No NoRat PK, (10 mg/kg), AUC (ng.h/mL)18 427 682Brain conc. @ 6 h (ng/g) 67 257Brain/Plasma 1.8 2.5Clearance (mL/min/kg) 30 22Bioavailability (%) 16 34SNNNONHCl SNNNONHClOHSNNNONHClOH(11q) C18H11ClN4OS (RT = 14.1 Min)Mol. Wt.: 366.82(14) C18H11ClN4O2S (RT = 13.0 Min)Mol. Wt.: 382.82HOH(17) C18H13ClN4O3S (RT = 3.9 Min)Mol. Wt.: 400.84SNNNONHClOHOSNNNONH2Cl(15) C15H9ClN4OS (RT = 11.3 Min)Mol. Wt.: 328.78OH[O]H2O+(16)(18)Figure 3. Metabolism of propargyl group (Mass analysis of rat bile samples, 0–24 h).SNNNONHR1 R219; R1 = Me, R2 = H; h-IC50 = 314 nM20; R1 = Me, R2 = Me; h-IC50 = 171 nM21; R1 = Et, R2 = Et; h-IC50 = 358 nMαFigure 4. Effect of a-alkylation on human mGluR1 potency.3202 T. K. Sasikumar et al. / Bioorg. Med. Chem. Lett. 19 (2009) 3199–3203was incubated with rat liver microsome for 0–24 h and the metab-olites were identified by LC–MS methodology. The oxidation of thecarbon adjacent to the amine lead to the intermediate 14 followedby degradation to the final molecules 15 and 16. LC–MS peak at13 min with an observed mass of 383 is attributed to the interme-diate 14. LC–MS analysis showed a peak at 3.9 min. with a mass of401 could be water addition product (17) to intermediate 14 asshown in Figure 3. The parent amino compound 15 displayed inLC–MS at 11.3 min. This retention time of the amine 15 was inde-pendently confirmed via analysis of pure synthetic sample. Com-pound 15 was found to be the major metabolite in the rat bile.The side product of this whole sequence, compound 16, is a knownglutathione scavenger described in the literature.19In order to avoid the metabolic issues, we introduced methylgroups adjacent to the amino group. Unfortunately monomethyla-tion and dimethylation of our lead compounds afforded 200–300-fold less active compounds (19–21) as shown in Figure 4. Identicalresults were obtained for compounds with different substitutionon the right hand side aromatic ring.In summary, we have achieved a large number of single digitnanomolar mGluR1 antagonists in the tricyclic series. This excel-lent potency sometimes extrapolates to good in vivo efficacy asseen in compounds 11q and 11s. These types of compounds showmoderate PK and high brain plasma ratio. It has also been foundthat these compounds are inactive in the hERG assay. Further datawill be published elsewhere.AcknowledgmentsWe thank Drs. Deen Tulshian, Julius Matasi and Peter Korakasfor helpful discussions. We also thank Lisa Broske’s group for ani-mal dosing and Sam Wainhaus’ group for PK sample analysis.References and notes1. Higgins, G. A.; Miczek, K. A. Psychopharmacology 2005, 179, 1.2. Kew, J. N. C.; Kemp, J. A. Psychopharmacology 2005, 179, 4.3. Steckler, T.; Lavreysen, H.; Oliveira, A. M.; Aerts, N.; Van Craenendonck, H.;Prickaerts, J.; Megens, A.; Lesage, A. S. J. Psychopharmacology 2005, 179, 198.4. Niswender, C. M.; Jones, C. K.; Conn, P. J. Curr. Top. Med. Chem. 2005, 5, 847.5. Kunishima, N.; Shimada, Y.; Tsuji, Y.; Sato, T.; Yamamoto, M.; Kumasaka, T.;Nakanishi, S.; Jingami, H.; Morikawa, K. Nature 2000, 407, 971.6. Constantino, G.; Macchiarulo, A.; Pellicciari, R. Bioorg. Med. Chem 2001, 9, 847.7. Layton, M. E. Curr. Top. Med. Chem 2005, 5, 859.8. Ott, D.; Floersheim, P.; Inderbitzin, W.; Stoehr, N.; Francotte, E.; Lecis, G.;Richert, P.; Rihs, G.; Flor, P. J.; Kuhn, R.; Gasparini, F. J. Med. 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See:Zheng, G. Z.; Bhatia, P.; Daanen, J.; Kolasa, T.; Patel, M.; Latshaw, S.;Kouhen, O. F. E.; Chang, R.; Uchic, M. E.; Miller, L.; Nakane, M.; Lehto, S.G.; Honore, M. P.; Moreland, R. B.; Brioni, J. D.; Stewart, A. O. J. Med.Chem. 2005, 48, 7374.14. Our efforts in the modification of tricyclic structure is being published in thefollowing patents, WO 2006/002051 and US 2006/167029. Also see: Wu, W-L.;Burnett, D. A.; Domalski, D.; Greenlee, W. J.; Li, C.; Bertorelli, R.; Fredduzzi, S.;Lozza, G.; Veltri, A.; Reggiani, A. J. Med. Chem 2007, 50, 5550. Biology materialsand methods were described in this Letter.15. Typical one step synthesis of tricyclic compound is described as follows: Compound6 (2 g, 0.0084 mol) was suspended in 15 mL triethyl orthoformate and treatedwith acetic acid (2 mL) and 4-chloroaniline (2 g, 0.0156 mol). The contentswere heated in a sealed tube at 160 C for 16 h. 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Tricyclic thienopyridine–pyrimidones/thienopyrimidine–pyrimidones as orally efficacious mGluR1 antagonists for neuropathic pain

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Tricyclic thienopyridine\u2013pyrimidones/thienopyrimidine\u2013pyrimidones as orally efficacious mGluR1 antagonists for neuropathic pain

Tricyclic thienopyridine\u2013pyrimidones/thienopyrimidine\u2013pyrimidones as orally efficacious mGluR1 antagonists for neuropathic pain

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