HETEROCYCLES

Short Paper | Regular issue | Vol. 91, No. 4, 2015, pp. 835-848
Received, 1st January, 2015, Accepted, 16th February, 2015, Published online, 24th February, 2015.
DOI: 10.3987/COM-15-13166

■ Convenient Synthesis of Novel Phenylpyrimido[1,2-c]thienopyrimidinones as IL-6/STAT3 Inhibitors

Jae Hoo Park, So Young Hong, Jungah Kim, Hyuck Joo Lee, Hyun Ho Lee, Ka Young Kim, Seung Woong Lee, Hyun-Mee Oh, Mun-Chul Rho, Beom-Gyu Lee, and Yang-Heon Song*

Department of Chemistry, Mokwon University, Daejeon, Doan-dong 800, 302-729, Korea

Abstract

New phenylpyrimido[1,2-c]thienopyrimidinones 4A and 4B were easily prepared in good yields by the one-pot reaction of formamidine derivatives 2 of 4-aminothienopyrimidine 1 with phenylacetyl chlorides. The application of this convenient and reliable method could be used for the synthesis of a variety of pyrimido[1,2-c]thienopyrimidinone derivatives of biological importance. Some of the compounds synthesized showed strong IL-6/STAT3 inhibition.

Amongst pro-inflammatory cytokines, interleukin-6 (IL-6) plays a key role in the induction of initiation and extension of the inflammatory process and immune response.1 And, an over-production of IL-6 is responsible for several diseases such as rheumatoid arthritis, psoriasis, inflammatory bowel disease, osteoarthritis, multiple myeloma and also for human atherosclerotic plaque.2 IL-6 binds to its receptor and leads to the activation of the Janus kinase (Jak)/Signal Transducer and Activator of Transcription-3 (STAT3).3 STAT3 is also frequently over-expressed or persistently activated in most tumors and cancer, and activated STAT3 was found to suppress tumor-immune surveillance.4 Therefore, the inhibition of STAT3 activation pathway stimulated by IL-6 represents a useful therapeutic strategy for discovery of new anti-inflammatory and anticancer drugs and is currently under intense investigation.5
In the other hand, thienopyrimidines, pyrido[1,2-a]pyrimidinones and pyrimido[1,6-a]pyrimidinones have been reported to have a wide range of biological properties, and their structural motif is present in the antitumor (I),6 antiplatelet agent (II),7 anxiolytic agent (III)8 and antiviral agent (IV).9 As part of a programme to discover novel IL-6/STAT3 inhibitors containing thienopyrimidines and to synthesize thienopyrimidine derivatives,10 we focused our attention on the new heterocyclic scaffold, phenylpyrimido[1,2-c]thienopyrimidinones 4A and 4B which are incorporated thiophene moiety into pyrimidopyrimidinones in attempt to improve the IL-6/STAT3 inhibitory activity. Herein, we describe a convenient and reliable method for the preparation of these compounds having biological activity.

The first synthetic route to target 4A (route A) is described in Scheme 1. The multi-step synthetic approach was investigated by using the conventional method previously described in the literature for the synthesis11 (Gould-Jacob type reaction) and the functionalization12 (Suzuki-Miyaura reaction) of pyridopyrimidinone analogues. However, the synthetic reaction for the precursor of 4Aa (R1, R2 = X = H), pyrimido[1,2-c]thienopyrimidinone skeleton, gave a mixture of inseparable products in 25% yield under high temperature (>260 C) and longer reaction time. Modified methods using the condensation of 4-aminothienopyrimidine 1 with 3-dimethylamino-2-arylpropenoates,13 and using microwave assisted synthesis14 were not also effective. Deniaud reported recently the synthetic method of pyrimido[1,2-a]pyrimidine-2,6-diones by using formamidines and acetyl chloride.15 In this literature, however, only methoxy or methoxycarbonyl group was introduced on the 7-position of pyrimido[1,2-a]pyrimidine-2,6-diones that were formed by cyclization reaction, and the scope and generality of the reaction to other heterocycles was not examined.
With the purpose of devising a milder and more convenient method of synthesizing 4A and 4B, another synthetic route B (this work) was examined by the cyclization reaction of formamidine derivative, (E)-N,N-dimethyl-N-(thieno[2,3-d]pyrimidin-4-yl)formimidamide (2Aa, R1, R2 = H), of 1Aa with 2-phenylacetyl chloride (3a) via 2Aa’. The key reactant 2Aa was easily prepared in quantitative yield by the reaction of 1Aa with dimethylformamide dimethyl acetal (DMF-DMA), and used without isolation for the next reaction with 2-phenylacetyl chloride. We indeed observed that the reaction proceeded smoothly in THF at room temperature affording the desired compound 4Aa as the only product in 48% yield (Table 1, entry 1). As shown in Table 1, reaction optimization for 4Aa was investigated at the various conditions such as different catalyst, reaction temperature and solvent. Use of ethanol or DMF as solvent resulted in inferior yield of product (Table 1, entries 3, 4, 10, 11). When the same reaction was carried out in dichloromethane in the presence of 0.1 equiv Et3N as a catalyst at room temperature, 4Aa was obtained in best yield within 2 h (Table 1, entry 6). The solid product was easily isolated with filtration, washing and drying. Notably, it was found that the stoichiometric usage of Et3N did not increased the yield as compared to its catalytic amount (Table 1, entry 7), and longer reaction time also did not affect the yield of 4Aa (Table 1, entry 8).

Next, as depicted in Table 2, the reaction of 2Aa with 3b-h was further evaluated under optimized reaction condition. Desired products 4Ab-g were obtained in good yields without side product. The results also indicated that 3b-f having electron-withdrawing group on phenyl ring gave products in slightly better yields (Table 2, entries 2-6) as compared with 3g having electron-donating group (Table 2, entry 7). No desired product 4Ah, however, was formed when the reaction was carried out with 3h (Table 2, entry 8). This could be due to the steric hindrance of two methyl groups at ortho position on phenyl of 3h. The reaction of 2Ba with 3a-g was also successfully applied to the synthesis of 4Ba-g as products. Notably, the reaction with 2Aa gave the corresponding products in little higher yield than that of 2Ba. The cyclohexyl- and cyclopentylthiophene derivatives, 2Ab and 2Ac, also performed well, affording the respective 4Ai-o and 4Ap-t in good yield (Table 2, entries 17-28). All compounds were characterized from their spectroscopic data.

To explore the diversity of this methodology, various heterocyclic amines were used in cyclization reaction. Under the same reaction condition, phenylpyrido[1,2-a]pyrimidinones, phenylpyrimido[1,2-c]- quinazolinones and phenylpyrazino[1,2-a]pyrimidinones 5 – 11 were also obtained in good yield (Table 3).

The synthesized compounds 4A and 4B were tested for their inhibitory activities on STAT3-dependent luciferase activity induced by IL-6, according to the reported method.16 The inhibitory activities of compounds were investigated as compared to genistein was used as a positive control which inhibited the STAT3-dependent luciferase activity with an IC50 value of 15 μM in this assay system.17 Among these compounds, some of strong IL-6/STAT3 inhibitors are listed in Table 4.

Phenylpyrimido[1,2-c]thieno[2,3-e]pyrimidinone analogues (4Bc, 4Bd, 4Bg) have slightly potent inhibitory activities than phenylpyrimido[1,2-c]thieno[3,2-e]pyrimidinone analogues (4Ac, 4Ag). The presence of cyclohexane or cyclopentane fused thiophene ring resulted in the improved activity of compounds (4Ai-4At). The most potent compound 4Ak showed inhibitory activity (IC50 value of 0.52 μM) with 30-fold when compared to genistein. None of the compounds tested had any cytotoxicity in Hep3B cells with MTT assay (data not shown).
In summary, we have showed the convenient synthesis of novel phenylpyrimido[1,2-c]thienopyrimidinone derivatives, and investigated preliminary biological activity for them. This simple and reliable one-pot synthetic method may allow the preparation of a larger library of these compounds of biological importance. Further investigations on their biological evaluation on IL-6/STAT3 inhibition and the application of methodology to other heterocyclic scaffold are currently underway.

EXPERIMENTAL
Melting points were determined in capillary tubes on Büchi apparatus and are uncorrected. Reactions were checked and monitored on thin-layer chromatography of Merck Kieselgel 60F254. The 1H NMR spectra were recorded on Unity Inova 400NB FT NMR spectrometer (400 MHz) with Me4Si as internal standard and chemical shifts are given in ppm (δ). Mass spectra were recorded on a HP 59580 B spectrometer. Elemental analyses were performed on a Carlo Erba 1106 elemental analyzer.
General procedure for the preparation of (E)-N,N-dimethyl-N-(thieno[2,3-d]pyrimidin-4-yl)formimidamide (2A) and (2B).
A mixture of thieno[2,3-d]pyrimidin-4-amine 1A or 1B (30 mmol) and dimethylformamide dimethyl acetal (DMF-DMA) (36 mmol) was heated to 110 C for 8 h. The solution was evaporated to dryness, and the crude product was used without isolation for the next reaction.
(E)-N,N-Dimethyl-N-(thieno[2,3-d]pyrimidin-4-yl)formimidamide (2Aa).
Yield: 95%; mp 96-97 C (recrystallized from n-hexane); 1HNMR (400 MHz, DMSO-d6) δ 8.89 (s, 1H), 8.55 (s, 1H), 7.64 (d, 1H, J = 5.8 Hz), 7.48 (d, 1H, J = 5.8 Hz), 3.20 (s, 6H); MS (ESI): m/z 206.63 (M+); Anal. Calcd for C9H10N4S: C, 52.41; H, 4.89; N, 27.16. Found: C, 52.23; H, 4.77; N, 27.02.
(E)-N,N-Dimethyl-N’-(5,6,7,8-tetrahydrobenzo[4,5]thieno[2,3-d]pyrimidin-4-yl)formimidamide (2Ab).
Yield: 98%; mp 136-137 C (recrystallized from n-hexane); 1HNMR (400 MHz, CDCl3) δ 8.56 (s, 1H), 8.35 (s, 1H), 3.03 (s, 6H), 2.99 (s, 2H), 2.68 (s, 2H), 1.72 (s, 4H); MS (ESI): m/z 261.1 (M++1); Anal. Calcd for C13H16N4S: C, 59.97; H, 6.19; N, 21.52. Found: C, 59.89; H, 6.10; N, 21.40.
(E)-N’-(6,7-Dihydro-5H-cyclopenta[4,5]thieno[2,3-d]pyrimidin-4-yl)-N,N-dimethylformimidamide (2Ac).
Yield: 97%; mp 145-146 C (recrystallized from n-hexane); 1HNMR (400 MHz, CDCl3) δ 8.76 (s, 1H), 8.52 (s, 1H), 3.18-3.11 (m, 8H), 2.99 (s, 2H), 2.46 (s, 2H); MS (ESI): m/z 246.8 (M+); Anal. Calcd for C12H14N4S: C, 58.51; H, 5.73; N, 22.74. Found: C, 58.64; H, 5.61; N, 22.86.
(E)-N,N-Dimethyl-N-(thieno[3,2-d]pyrimidin-4-yl)formimidamide (2Ba).
Yield: 95%; mp 107-108 C (recrystallized from CHCl3 and petroleum ether); 1HNMR (400 MHz, DMSO-d6) δ 8.91 (s, 1H), 8.78 (s, 1H), 7.82 (d, 1H, J = 5.8 Hz), 7.48 (d, 1H, J = 5.8 Hz), 3.22 (s, 6H); MS (ESI): m/z 206.57 (M+); Anal. Calcd for C9H10N4S: C, 52.41; H, 4.89; N, 27.16. Found: C, 52.52; H, 4.78; N, 27.09.
General procedure for the preparation of phenylpyrimido[1,2-c]thieno[3,2-e]pyrimidinone (4Aa-t) and phenylpyrimido[1,2-c]thieno[2,3-e]pyrimidinone (4Ba-g).
A suspension of 2A or 2B (4 mmol) and 3 (4.8 mmol) in the presence of a catalytic amount of triethylamine in CH2Cl2 (10 mL) was stirred at rt for 2 h. The solid product was filtered, washed with EtOAc and dried. The crude product was recrystallized from EtOH to give 4A or 4B.
3-Phenyl-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Aa).
Yield: 82%; mp 231-232 C; 1HNMR (400 MHz, DMSO-d6) δ 9.69 (s, 1H), 8.59 (s, 1H), 8.04 (d, 1H, J = 5.8 Hz), 7.90 (d, 1H, J = 5.8 Hz), 7.80 (d, 2H, J = 7.8 Hz), 7.47 (t, 2H, J = 7.8 Hz), 7.38 (t, 1H, J = 7.8 Hz); MS (ESI): m/z 279.82 (M+); Anal. Calcd for C15H9N3OS: C, 64.50; H, 3.25; N, 15.04 Found: C, 64.31; H, 3.14; N, 15.20.
3-(4-Bromophenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Ab).
Yield: 85%; mp 253-254 C; 1HNMR (400 MHz, DMSO-d6) δ 9.58 (s, 1H), 8.61 (s, 1H), 8.02 (d, 1H, J = 5.8 Hz), 7.88 (d, 1H, J = 5.8 Hz), 7.76 (d, 2H, J = 8.0 Hz), 7.73 (d, 2H, J = 8.0 Hz); MS (ESI): m/z 358.4 (M+); Anal. Calcd for C15H8BrN3OS: C, 50.29; H, 2.25; N, 11.73. Found: C, 50.12; H, 2.19; N, 11.61.
3-(2-Chlorophenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Ac).
Yield: 84%; mp 243-244 C; 1HNMR (400 MHz, DMSO-d6) δ 9.55 (s, 1H), 8.36 (s, 1H), 8.04 (d, 1H, J = 5.8 Hz), 7.89 (d, 1H, J = 5.8 Hz), 7.46 (d, 1H, J = 8.0 Hz), 7.40-7.30 (m, 3H); MS (ESI): m/z 313.5 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.30; H, 2.50; N, 11.41.
3-(3-Chlorophenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Ad).
Yield: 89%; mp 259-260 C; 1HNMR (400 MHz, DMSO-d6) δ 9.59 (s, 1H), 8.65 (s, 1H), 8.04 (d, 1H, J = 5.8 Hz), 7.90 (m, 2H), 7.76 (d, 1H, J = 8.0 Hz), 7.49-7.43 (m, 2H); MS (ESI): m/z 313.8 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.50; H, 2.50; N, 11.44.
3-(4-Chlorophenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Ae).
Yield: 92%; mp 255-256 C; 1HNMR (400 MHz, DMSO-d6) δ 9.58 (s, 1H), 8.61 (s, 1H), 8.02 (d, 1H, J = 5.8 Hz), 7.88 (d, 1H, J = 5.8 Hz), 7.83 (d, 2H, J = 7.8 Hz), 7.50 (d, 2H, J = 7.8 Hz); MS (ESI): m/z 313.9 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.55; H, 2.48; N, 11.24.
3-(4-Nitrophenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Af).
Yield: 90%; mp 344-345 C; 1HNMR (400 MHz, DMSO-d6) δ 9.63 (s, 1H), 8.77 (s, 1H), 8.28 (d, 2H, J = 8.0 Hz), 8.13 (d, 2H, J = 8.0 Hz), 8.06 (d, 1H, J = 5.8 Hz), 7.91 (d, 1H, J = 5.8 Hz); MS (ESI): m/z 324.8 (M+); Anal. Calcd for C15H8N4O3S: C, 55.55; H, 2.49; N, 17.28. Found: C, 55.44; H, 2.40; N, 17.38.
3-(4-Methoxyphenyl)-4H-pyrimido[1,2-c]thieno[3,2-e]pyrimidin-4-one (4Ag).
Yield: 72%; mp 208-209 C; 1HNMR (400 MHz, DMSO-d6) δ 9.56 (s, 1H), 8.52 (s, 1H), 8.00 (d, 1H, J = 5.8 Hz), 7.85 (d, 1H, J = 5.8 Hz), 7.73 (d, 2H, J = 8.0 Hz), 7.00 (d, 2H, J = 8.0 Hz), 3.77 (s, 3H); MS (ESI): m/z 309.1 (M+); Anal. Calcd for C16H11N3O2S: C, 62.12; H, 3.58; N, 13.58. Found: C, 62.01; H, 3.55; N, 13.38.
3-Phenyl-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Ai).
Yield: 77%; mp 214-215 C; 1HNMR (400 MHz, CDCl3) δ 9.69 (s, 1H), 8.50 (s, 1H), 7.78 (d, 2H, J = 7.8 Hz), 7.50 (t, 2H, J = 7.8 Hz), 7.41 (d, 1H, J = 8.0 Hz), 3.29-3.27 (m, 2H), 2.95-2.92 (m, 2H), 1.98-1.94 (m, 4H); MS (ESI): m/z 334.2 (M+); Anal. Calcd for C19H15N3OS: C, 68.45; H, 4.53; N, 12.60. Found: C, 68.34; H, 4.42; N, 12.72.

3-(4-Bromophenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Aj).
Yield: 82%; mp 211-212 C; 1HNMR (400 MHz, CDCl3) δ 9.60 (s, 1H), 8.41 (s, 1H), 7.60 (d, 2H, J = 8.0 Hz), 7.53 (d, 2H, J = 8.0 Hz), 3.20-3.19 (m, 2H), 2.88-2.87 (m, 2H), 1.89-1.87 (m, 4H); MS (ESI): m/z 413.9 (M+); Anal. Calcd for C19H14BrN3OS: C, 55.35; H, 3.42; N, 10.19. Found: C, 55.22; H, 3.52; N, 10.08.
3-(2-Chlorophenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Ak).
Yield: 80%; mp 234-235 C; 1HNMR (400 MHz, CDCl3) δ 9.54 (s, 1H), 8.24 (s, 1H), 7.43 (d, 1H, J = 7.6 Hz), 7.36 (d, 1H, J = 7.6 Hz), 7.27-7.25 (m, 2H), 3.19-3.17 (m, 2H), 2.85-2.83 (m, 2H), 1.85-1.84 (m, 4H); MS (ESI): m/z 367.3 (M+); Anal. Calcd for C19H14ClN3OS: C, 62.04; H, 3.84; N, 11.42. Found: C, 61.95; H, 3.77; N, 11.30.
3-(3-Chlorophenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Al).
Yield: 88%; mp 238-239 C; 1HNMR (400 MHz, CDCl3) δ 9.60 (s, 1H), 8.42 (s, 1H), 7.72 (s, 1H), 7.60 (d, 1H, J = 7.9 Hz), 7.35-7.28 (m, 2H), 3.21-3.19 (m, 2H), 2.88-2.87 (m, 2H), 1.88-1.87 (m, 4H); MS (ESI): m/z 367.1(M+); Anal. Calcd for C19H14ClN3OS: C, 62.04; H, 3.84; N, 11.42. Found: C, 62.11; H, 3.79; N, 11.33.
3-(4-Chlorophenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Am).
Yield: 90%; mp 230-231 C; 1HNMR (400 MHz, CDCl3) δ 9.67 (s, 1H), 8.48 (s, 1H), 7.74 (d, 2H, J = 8.0 Hz), 7.45 (d, 2H, J = 8.0 Hz), 3.28-3.26 (m, 2H), 2.94-2.93 (m, 2H), 1.98-1.95 (m, 4H); MS (ESI): m/z 368.2 (M+); Anal. Calcd for C19H14ClN3OS: C, 62.04; H, 3.84; N, 11.42. Found: C, 62.16; H, 3.77; N, 11.36.
3-(4-Nitrophenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4An).
Yield: 91%; mp 300-301 C; 1HNMR (400 MHz, CDCl3) δ 9.63 (s, 1H), 8.52 (s, 1H), 7.94 (d, 2H, J = 8.0 Hz), 7.27 (d, 2H, J = 8.0 Hz), 3.22 (s, 2H), 2.89 (s, 2H), 1.89 (s, 4H); MS (ESI): m/z 378.1 (M+); Anal. Calcd for C19H14N4O3S: C, 60.31; H, 3.73; N, 14.81. Found: C, 60.20; H, 3.66; N, 14.70.
3-(4-Methoxyphenyl)-9,10,11,12-tetrahydro-4H-benzo[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4-one (4Ao).
Yield: 75%; mp 210-211 C; 1HNMR (400 MHz, CDCl3) δ 9.60 (s, 1H), 8.38 (s, 1H), 7.66 (d, 2H, J = 8.0 Hz), 6.94 (d, 2H, J = 8.0 Hz), 3.79 (s, 3H), 3.20 (s, 2H), 2.87 (s, 2H), 1.88 (s, 4H); MS (ESI): m/z 363.8 (M+); Anal. Calcd for C20H17N3O2S: C, 66.10; H, 4.71; N, 11.56. Found: C, 66.01; H, 4.63; N, 11.44.
3-Phenyl-10,11-dihydrocyclopenta[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4(9H)-one (4Ap).
Yield: 78%; mp 205-206 C; 1HNMR (400 MHz, CDCl3) δ 9.50 (s, 1H), 8.36 (s, 1H), 7.62 (d, 2H, J = 8.0 Hz), 7.26 (t, 2H, J = 8.0 Hz), 7.11 (d, 1H, J = 8.0 Hz), 3.14 (s, 2H), 2.97 (s, 2H), 2.43 (s, 2H); MS (ESI): m/z 320.1 (M+); Anal. Calcd for C18H13N3OS: C, 67.69; H, 4.10; N, 13.16. Found: C, 67.59; H, 4.01; N, 13.31.
3-(4-Bromophenyl-10,11-dihydrocyclopenta[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4(9H)-one (4Aq).
Yield: 80%; mp 185-186 C; 1HNMR (400 MHz, CDCl3) δ 9.54 (s, 1H), 8.21 (s, 1H), 7.43 (d, 2H, J = 8.0 Hz), 7.16 (d, 2H, J = 8.0 Hz), 3.56 (s, 2H), 2.98 (s, 2H), 2.53 (s, 2H); MS (ESI): m/z 397.8 (M+); Anal. Calcd for C18H12BrN3OS: C, 54.28; H, 3.04; N, 10.55. Found: C, 54.10; H, 3.13; N, 10.41.
3-(2-Chlorophenyl-10,11-dihydrocyclopenta[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4(9H)-one (4Ar).
Yield: 77%; mp 254-255 C; 1HNMR (400 MHz, CDCl3) δ 9.45 (s, 1H), 8.19 (s, 1H), 7.35 (d, 1H, J = 8.0 Hz), 7.33 (d, 1H, J = 7.9 Hz), 7.28 (m, 2H), 3.13 (s, 2H), 2.96 (s, 2H), 2.43 (s, 2H); MS (ESI): m/z 354.1 (M+); Anal. Calcd for C18H12ClN3OS: C, 61.10 H, 3.42; N, 11.88. Found: C, 61.00; H, 3.35; N, 11.75.
3-(4-Chlorophenyl-10,11-dihydrocyclopenta[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4(9H)-one (4As).
Yield: 88%; mp 255-256 C; 1HNMR (400 MHz, CDCl3) δ 9.54 (s, 1H), 8.38 (s, 1H), 7.61 (d, 2H, J = 8.0 Hz), 7.33 (d, 2H, J = 8.0 Hz), 3.18 (s, 2H), 3.01 (s, 2H), 2.47 (s, 2H); MS (ESI): m/z 353.9 (M+); Anal. Calcd for C18H12ClN3OS: C, 61.10 H, 3.42; N, 11.88. Found: C, 61.22; H, 3.49; N, 11.79.
3-(4-Methoxyphenyl-10,11-dihydrocyclopenta[4,5]thieno[3,2-e]pyrimido[1,2-c]pyrimidin-4(9H)-one (4At).
Yield: 72%; mp 206-207 C; 1HNMR (400 MHz, CDCl3) δ 9.53 (s, 1H), 8.36 (s, 1H), 7.60 (d, 2H, J = 8.0 Hz), 6.89 (d, 2H, J = 8.0 Hz), 3.75 (s, 3H), 3.17 (s, 2H), 3.00 (s, 2H), 2.46 (s, 2H); MS (ESI): m/z 350.2 (M+); Anal. Calcd for C19H15N3O2S: C, 65.31 H, 4.33; N, 12.03. Found: C, 65.25 H, 4.26; N, 12.12.
8-Phenyl-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Ba).
Yield: 77%; mp 257-258 C; 1HNMR (400 MHz, DMSO-d6) δ 9.62 (s, 1H), 8.56 (s, 1H), 8.42 (d, 1H, J = 5.8 Hz), 7.81 (d, 2H, J = 8.0 Hz), 7.73 (d, 1H, J = 5.8 Hz), 7.48 (t, 2H, J = 8.0 Hz), 7.40 (t, 1H, J = 8.0 Hz) ; MS (ESI): m/z 279.95 (M+); Anal. Calcd for C15H9N3OS: C, 64.50; H, 3.25; N, 15.04 Found: C, 64.66; H, 3.34; N, 15.17.
8-(4-Bromophenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Bb).
Yield: 75%; mp 237-238 C; 1HNMR (400 MHz, DMSO-d6) δ 9.57 (s, 1H), 8.56 (s, 1H), 8.39 (d, 1H, J = 5.8 Hz), 7.74 (d, 2H, J = 8.0 Hz), 7.69 (d, 1H, J = 5.8 Hz), 7.63 (d, 2H, J = 8.0 Hz); MS (ESI): m/z 358.8 (M+); Anal. Calcd for C15H8BrN3OS: C, 50.29; H, 2.25; N, 11.73. Found: C, 50.19; H, 2.15; N, 11.79.
8-(2-Chlorophenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Bc).
Yield: 78%; mp 203-204 C; 1HNMR (400 MHz, DMSO-d6) δ 9.54 (s, 1H), 8.40 (d, 1H, J = 5.8 Hz), 8.31 (s, 1H), 7.69 (d, 1H, J = 5.8 Hz), 7.56-7.54 (m, 1H), 7.46-7.40 (m, 3H); MS (ESI): m/z 313.3 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.50; H, 2.49; N, 11.21.
8-(3-Chlorophenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Bd).
Yield: 79%; mp 335-336 C; 1HNMR (400 MHz, DMSO-d6) δ 9.58 (s, 1H), 8.60 (s, 1H), 8.40 (d, 1H, J = 5.8 Hz), 7.87 (s, 1H), 7.75 (d, 1H, J = 7.8 Hz), 7.70 (d, 1H, J = 5.8 Hz), 7.49-7.40 (m, 2H); MS (ESI): m/z 313.8 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.34; H, 2.56; N, 11.40.
8-(4-Chlorophenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Be).
Yield: 83%; mp 229-230 C; 1HNMR (400 MHz, DMSO-d6) δ 9.53 (s, 1H), 8.52 (s, 1H), 8.35 (d, 1H, J = 5.8 Hz), 7.77 (d, 2H, J = 8.0 Hz), 7.65 (d, 1H, J = 5.8 Hz), 7.45 (d, 2H, J = 8.0 Hz); MS (ESI): m/z 313.9 (M+); Anal. Calcd for C15H8ClN3OS: C, 57.42; H, 2.57; N, 11.30. Found: C, 57.49; H, 2.51; N, 11.41.
8-(4-Nitrrophenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Bf).
Yield: 88%; mp 331-332 C; 1HNMR (400 MHz, DMSO-d6) δ 9.62 (s, 1H), 8.73 (s, 1H), 8.44 (d, 1H, J = 5.8 Hz), 8.29 (d, 2H, J = 7.8 Hz), 8.11 (d, 2H, J = 7.8 Hz), 7.73 (d, 1H, J = 5.8 Hz); MS (ESI): m/z 324.1 (M+); Anal. Calcd for C15H8N4O3S: C, 55.55; H, 2.49; N, 17.28. Found: C, 55.48; H, 2.41; N, 17.40.
8-(4-Methoxyphenyl)-7H-pyrimido[1,2-c]thieno[2,3-e]pyrimidin-7-one (4Bg).
Yield: 70%; mp 228-229 C; 1HNMR (400 MHz, DMSO-d6) δ 9.55 (s, 1H), 8.47 (s, 1H), 8.34 (d, 1H, J = 5.8 Hz), 7.72 (d, 2H, J = 8.0 Hz), 7.66 (d, 1H, J = 5.8 Hz), 6.99 (d, 2H, J = 8.0 Hz), 3.76 (s, 3H); MS (ESI): m/z 309.7 (M+); Anal. Calcd for C16H11N3O2S: C, 62.12; H, 3.58; N, 13.58. Found: C, 62.21; H, 3.48; N, 13.47.
General procedure for the preparation of phenylpyrido[1,2-a]pyrimidinones, phenylpyrimido[1,2-c]quinazolinones and phenylpyrazino[1,2-a]pyrimidinones 5 – 11.
A suspension of formamidine derivative of various heterocyclic amines (4 mmol) and 3 (4.8 mmol) in the presence of a catalytic amount of triethylamine in CH2Cl2 (10 mL) was stirred at rt for 2 h. The solid product was filtered, washed with EtOAc and dried. The crude product was recrystallized from EtOH to give 5 – 11.
3-Phenyl-4H-pyrido[1,2-a]pyrimidin-4-one (5).
Yield: 78%; mp 166-167 C; 1HNMR (400 MHz, DMSO-d6) δ 9.14 (d, 1H, J = 7.6 Hz), 8.63 (s, 1H), 8.02 (t, 1H, J = 7.5 Hz), 7.86 (d, 2H, J = 7.9 Hz), 7.78 (d, 1H, J = 7.6 Hz), 7.49-7.48 (m, 3H), 7.39 (t, 1H, J = 8.0 Hz); MS (ESI): m/z 223.03 (M+).
3-(2-Chlorophenyl)-4H-pyrido[1,2-a]pyrimidin-4-one (6).
Yield: 85%; mp 160-161 C; 1HNMR (400 MHz, DMSO-d6) δ 9.04 (d, 1H, J = 7.6 Hz), 8.32 (s, 1H), 7.99 (t, 1H, J = 7.5 Hz), 7.75 (d, 1H, J = 7.5 Hz), 7.53 (d, 1H, J = 7.9 Hz), 7.47 (t, 1H, J = 8.0 Hz), 7.42 (d, 1H, J = 8.0 Hz), 7.40 (t, 2H, J = 8.0 Hz); MS (ESI): m/z 256.67 (M+).
7-Bromo-3-(4-chlorophenyl)-4H-pyrido[1,2-a]pyrimidin-4-one (7).
Yield: 82%; mp 203-204 C; 1HNMR (400 MHz, DMSO-d6) δ 9.01 (s, 1H), 8.62 (s, 1H), 8.08 (d, 1H, J = 7.3 Hz), 7.85 (d, 2H, J = 7.9 Hz), 7.67 (d, 1H, J = 7.3 Hz), 7.48 (d, 2H, J = 7.9 Hz); MS (ESI): m/z 335.58 (M+).
3-Phenyl-4H-pyrimido[1,2-c]quinazolin-4-one (8).
Yield: 92%; mp 237-238 C; 1HNMR (400 MHz, DMSO-d6) δ 9.49 (s, 1H), 8.72 (d, 1H, J = 7.6 Hz), 8.60 (s, 1H), 7.97 (t, 2H, J = 8.0 Hz), 7.81-7.80 (m, 3H), 7.47 (t, 2H, J = 8.0 Hz), 7.39 (t, 1H, J = 7.9 Hz); MS (ESI): m/z 274.1 (M+).
3-(4-Methoxyphenyl)-4H-pyrimido[1,2-c]quinazolin-4-one (9).
Yield: 85%; mp 202-203 C; 1HNMR (400 MHz, DMSO-d6) δ 9.41 (s, 1H), 8.70 (d, 1H, J = 7.8 Hz), 8.55 (s, 1H), 7.94 (t, 2H, J = 8.0 Hz), 7.77-7.63 (m, 3H), 7.01 (d, 2H, J = 8.0 Hz), 3.76 (s, 3H); MS (ESI): m/z 303.31 (M+).
3-(4-Bromophenyl)-4H-pyrazino[1,2-a]pyrimidin-4-one (10).
Yield: 76%; mp 254-255 C; 1HNMR (400 MHz, DMSO-d6) δ 9.22 (s, 1H), 8.90 (s, 1H), 8.83 (d, 1H, J = 7.6 Hz), 8.33 (d, 1H, J = 7.6 Hz), 8.30 (d, 2H, J = 8.0 Hz), 8.18 (d, 2H, J = 8.0 Hz); MS (ESI): m/z 302.13 (M+).
3-(4-Nitrophenyl)-4H-pyrazino[1,2-a]pyrimidin-4-one (11).
Yield: 81%; mp 207-208 C; 1HNMR (400 MHz, DMSO-d6) δ 9.15 (s, 1H), 8.76 (d, 1H, J = 7.3 Hz), 8.75 (s, 1H), 8.25 (d, 1H, J = 7.4 Hz), 7.82 (d, 2H, J = 8.0 Hz), 7.65 (d, 2H, J = 8.0 Hz); MS (ESI): m/z 268.23 (M+).

ACKNOWLEDGEMENTS
This work was supported by the Korea Research Foundation (project number 2010-0021038).

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