Preparation and Antiviral Activity of Some New C₃- and C_S-Symmetrical Tri-Substituted Triazine Derivatives Having Benzylamine Substituents

Abstract

We report the preparation of new C₃- and C_S-symmetrical molecules constructed on a triazine (TAZ) template. Anti-herpes simplex virus type 1 (anti-HSV-1) and cytotoxic activities against Vero cells of synthesized TAZ derivatives were evaluated. The results suggested that the presence of an electron-donating group(s) on the benzene ring in benzylamine groups on the TAZ template is an important structural factor for expressing a high level of anti-HSV-1 activity and low cytotoxicity for these C₃ types of TAZ derivatives. Among the tested TAZ derivatives, compounds 4f and 7h showed the highest anti HSV-1 activities (EC₅₀=0.98 and 1.23 µM, respectively) and low cytotoxic activities to Vero cells (50% cytotoxic concentration (CC₅₀)=292.2 and >200 µM, respectively).

In the search for new types of antiviral active compounds, extensive efforts have been made to find promising new antiviral candidates.¹⁾ It is well known that many types of receptors or functionalized proteins on membranes in a native state often have a high order of symmetrical interfaces.²⁾ C₃- or C₂-geometric symmetrical molecules have frequently been found in various synthetic biologically active compounds.³⁾ Aiming to develop new synthetic bioactive compounds, we targeted such geometrical symmetric molecules constructed on a symmetrical template.

In our previous studies on antiviral compounds in the tri-substituted 1,3,5-triazine (TAZ) series, we found that many of the new C₃- and C_S-symmetrical TAZ derivatives synthesized previously showed a wide range of bioactivities against herpes simplex virus type 1 (HSV-1) or cytotoxicic activities to Vero cells.⁴⁾ Regarding tri-substituted TAZ derivatives, we have reported that some symmetrical TAZ derivatives having alkoxy and/or alkylamino substituents show a high level of anti-HSV-1 activity⁵⁾ (Fig. 1). Notably, we found that some TAZ derivatives that have benzylamine substituents on a TAZ template such as compound 4h show a high level of anti-HSV-1 activity (EC₅₀=5.4 µM) and low cytotoxicity (50% cytotoxic concentration (CC₅₀)=291.9 µM). In terms of a structure–activity relationship (SAR) study and finding new types of antiviral active leads, molecular modification by the introduction of benzylamine functionalities into a TAZ template seems to be interesting.

Fig. 1. Symmetrical Target Molecules Constructed on Tri-Substituted TAZ Derivatives

Here, we describe the synthesis of new types of symmetrical TAZ derivatives that have various benzylamine substituents on symmetrical TAZ templates and the results of their biological evaluations.

Results

Synthesis of New Target Derivatives of Symmetrical Tri-Substituted TAZ Derivatives

Our strategies for the synthesis of target TAZ derivatives by using 2,4,6-trichlorotriazine (TCT AZ: 1) as a starting material are summarized in Chart 1.

Chart 1. Synthesis of C₃- and C_S-Symmetrical TAZ Derivatives

The pathway for the synthesis of C₃- and C_S-type symmetrical TAZ derivatives (4 and 7) together with intermediates of TAZ derivatives is shown in Chart 1. All of the substitution reactions of the starting TCT AZ (1) or monoalkoxy-dichloro-TAZ (5)⁵⁾ with various benzylamines (2a–k) were performed in a one-pot reaction and the desired new TAZ derivatives (4 and 7) were obtained. All of the target C₃-type TAZ derivatives (4) were obtained in moderate to good yields (42–85%) as shown in Table 1, and C_S-type symmetrical TAZ derivatives (7) were also obtained in moderate yields (Table 2). For the synthesis of these new targeted C₃- and C_S-symmetrical tri-substituted TAZ derivatives, we used the same procedure as that described for the preparation of symmetrical TAZ derivatives (see Experimental).

Table 1. Synthesis of C₃-Symmetrical Benzylamine-Type TAZ Derivatives (4)

Entry	ArCH2NH2 (2)	Reaction time of MW (min)a)	Products (yield %)
1	2a	60	4a (85)
2	2b	60	4b (53)
3	2b	40b)	3b (46)c), 4b (5)c)
4	2c	60	4c (75)
5	2d	60	4d (42)
6	2e	30	4e (66)
7	2f	50	4f (80)
8	2g	20	4g (57)
9	2i	20	4i (63)

a) In the all reactions except for Entry 3, stirring of the reaction mixtures (ratio of 1 : 2=1 : 10) in dioxane were conducted at 0°C for 30 min, at room temperature for 1 h, and then irradiation of microwave (100 W, 100°C) for a proper shown time. b) Stirring the reaction mixture (ratio of 1 : 2b=1 : 6) in THF was conducted at room temperature for 10 min, and then irradiation of microwave (90 W, 70°C) for 20 min. c) Yields after recrystallization.

Table 2. Synthesis of C_S-Symmetrical Benzylamine-Type TAZ Derivatives (7)

Entry	ArCH2NH2 (2)	Ratio of 1 : 2 (: DIPEA)a)	Reaction conditionsa)	Product (yield %)
1	2a	1 : 1 : 1 (DIPEA)	0°C 2 h	6a (97)
2	2a	1 : 5	MW 60 min	7a (79)
3	2b	1 : 5	MW 45 min	7b (82)b)
4	2e	1 : 5	MW 60 min	7e (75)b)
5	2f	1 : 6	MW 30 min	7f (75)b)
6	2g	1 : 5	MW 30 min	7g (72)
7	2h	1 : 3	MW 30 min	7h (34)b)
8	2i	1 : 5	MW 30 min	7i (89)
9	2j	1 : 5	MW 30 min	7j (84)b)
10	2k	1 : 5	MW 45 min	7k (78)

a) In the all reactions except for Entry 1, the reaction mixtures were subjected to microwave irradiation (100 W, 100°C) in dioxane. In the reaction of Entry 1, the reaction mixture with N,N-diisopropylethylamine (DIPEA) was used in THF. b) Yields after recrystallization.

All of the structures of the synthesized C₃- and C_S-symmetrical TAZ derivatives (4 and 7) and a few intermediates (3, 6) were confirmed by spectroscopic and analytical data. The geometries of the obtained symmetrical TAZ derivatives described above were confirmed from ¹³C-NMR spectroscopic data⁶⁾ (see Experimental for details).

Biological Activity and Discussion

In many synthetic receptor-type molecules mimicking the supramolecular interaction of biomolecules, it is expected that the ability to form multiple hydrogen bonds plays an important role in guest molecule recognition. Considering the hydrogen bonded interaction in a supramolecular host-guest system, it is also expected that a convergent orientation of incorporated functional groups is an effective array to produce multiple hydrogen bonding interactions between host and guest molecules.^7,8) From this point of view, title C₃- and C_S-geometrical molecules that have donors or acceptors for hydrogen bonds on a symmetrical TAZ template are thought to offer new biologically active molecules.

From biological evaluations of anti-HSV-1 activities (EC₅₀) by plaque reduction assays⁹⁾ and cytotoxic activities (CC₅₀) against Vero cells of synthesized TAZ derivatives (Table 3), most of the C₃-symmetrical benzylamine-type TAZ derivatives (4) presented hydrogen bond donor protons of a benzylamine (–NH–) functionality. Many of the C₃-symmetrical benzylamine-type TAZ derivatives (4) showed a significantly high level of anti-HSV-1 activity, and compound 4f showed a high level of anti-HSV-1 activity (EC₅₀=0.98 µM) and a low level of cytotoxic activity (CC₅₀=292.2 µM) as did compound 4h⁵⁾ (EC₅₀=5.4 µM and CC₅₀=291.9 µM) reported previously. All of the compounds that have electron-withdrawing substituents (4c and 4d) including non-substituted compound 4a showed considerably high levels of anti-HSV-1 activity (EC₅₀=7.4–13.2 µM) and cytotoxic activity (CC₅₀=5.5–94.9 µM). Regarding the two biological activities (anti-HSV-1 activity and cytotoxic activity), the opposite order in electron-withdrawing (attracting) effects between the two activities was observed (4a<4c<4d for anti-HSV-1 activity and 4a>4c>4d for cytotoxic activity). Lower levels of cytotoxic activity (CC₅₀=13.0–94.9 µM) were observed for compounds 4c and 4d than for non-substituted compound 4a.

Table 3. Anti-HSV-1 Activity (EC₅₀) and Cytotoxicity (CC₅₀) against Vero Cells of Synthesized New C₃- and C_S-Symmetrical Benzylmine-Type TAZ Derivatives (4, 7) and Intermediates (3, 6)

a) Data were taken from ref. 5). b) Data were taken from ref. 15).

Considerably high levels of anti-HSV-1 activity (EC₅₀=0.98–38.7 µM) were observed for C₃-type compounds (4b, 4f, 4h⁵⁾ and 4i) that have electron-donating substituents such as methyl or methoxy groups on the benzene rings except for compounds 4g and 4e (see Table 3), and these compounds showed cytotoxic activities over 200 µM. The obtained biological results indicated that the nature of substituents on the benzene rings prefers electron-donating substituents for expressing a high level of anti-HSV-1 activity and low level of cytotoxicity for these C₃ types of TAZ derivatives. We also consider that benzylamine groups in C₃-type molecules may act as hydrogen-bond donors to receptor binding sites.

The exceptional anti-HSV-1 activity for compound 4g may be due to intramolecular hydrogen bond formation between the basic benzylamine moiety and the ortho-methoxy group in the molecule 4g that weakens such intermolecular hydrogen-bonding interaction of the TAZ molecule 4g to receptor binding sites (see Fig. 2). This exceptional result for compound 4g also indicates the importance of intermolecular hydrogen bond formation ability of C₃-type TAZ molecules in the supramolecular interaction with its binding site for expressing a high level of anti-HSV-1 activity.

Fig. 2. Intramolecular Hydrogen Bond Formation in Compound 4g

Despite the considerably high level of cyctotoxic activity (CC₅₀=22.6 µM) of compound 4e, the reason for exceptionally low level of anti-HSV-1 activity (EC₅₀>100 µM) for this compound is still unclear at present. The positional effect of methoxy groups on benzene rings in these TAZ derivatives may be required for expression of anti-HSV-1 activity.

Among the C_S-symmetrical benzylamine-type TAZ derivatives (7a, 7b, 7e–k) as shown in Table 3, a C_S-symmetrical benzylamine-type TAZ derivative (7h) that has two benzylamine groups with electron-donating substituents and an isopropoxy group showed a considerably high level of anti-HSV-1 activity (EC₅₀=1.23 µM); however, many structurally related C_S-symmetrical derivatives (7a, 7b, 7e, 7f, 7g and 7k) showed low levels of anti-HSV-1 activity (EC₅₀=>100 µM) compared to that of compound 7h. Compounds 7i and 7j showed moderate levels of anti-HSV-1 activity (EC₅₀=72.1 and 12.6 µM, respectively). Interestingly, many compounds in this series (7a, 7b, 7g, 7i, 7j and 7k) showed moderate levels of cytotoxic activity (CC₅₀=14.6–192.4 µM, see Table 3). Intermediates 3 and 6 listed in Table 3 showed almost no anti-HSV-1 or cytotoxic activity (see Table 3).

Regarding calculated log P values¹⁰⁾ for the compounds (see Supplementary Materials), there were few distinct correlations between log P values and anti-HSV-1 activity (EC₅₀) values shown in Table 3.

The results showing high levels of anti-HSV-1 activity for C₃- or C_S-symmetrical compounds (4f and 7h) strongly suggest that at least the presence of two hydrogen bond donor benzylamine protons in the target TAZ derivatives is required as an important structural factor for the expression of a high level of anti-HSV-1 activity. A similar assumption is also substantiated by our previous observation of the disappearance of anti-HSV-1 activity by N-acetylated C₃-symmetrical amine-type molecules.¹¹⁾

In the comparison with the anti-HSV-1 activity of acyclovir, it was found that the mode of action of acyclovir is different from the modes of action of TAZ derivatives described in this paper; however, it is noteworthy that some new symmetrical TAZ derivatives such as 4f and 7h showed similar levels of anti-HSV-1 activity with a large safety index value.

The results of calorimetric experiments¹²⁾ showed sugar recognition properties of highly active compounds with good SI values (CC₅₀/EC₅₀) such as compound 4f (SI=298). From the point of the view of substituents as hydrogen-bond donors or acceptors, ITC experiments suggested that the reaction of compound 4f with sugars showed a thermodynamic signature different from that of the entropically driven reaction of tri-isopropoxy-substituted TAZ derivative. Thus, we consider that such an enthalpic C₃-type TAZ derivative having benzylamine functionalities may act as a hydrogen-bond donor to proposed receptor binding sites. On the basis of the obtained structural information on biological activities in this series, we are further investigating chemical modifications to new TAZ framework molecules in order to find new promising leads.

Experimental

Melting points were determined using a micro melting point apparatus (Yanaco MP-S3) without correction. IR spectra were measured by a JASCO FTIR-4100 IR spectrophotometer. MicromATR Vision [an apparatus of attenuated total reflectance (ATR)] was used for a neat sample operation. Low- and high-resolution mass spectra (LR-MS and HR-MS) were obtained by a JEOL JMS HX-110 double-focusing model equipped with an FAB ion source interfaced with a JEOL JMA-DA 7000 data system. ^lH- and ¹³C-NMR spectra were obtained by ECG600R. Chemical shifts were expressed in δ ppm downfield from an internal tetramethylsilane (TMS) signal for ¹H-NMR and the carbon signal of the corresponding solvent [CDCl₃ (77.00 ppm), dimethyl sulfoxide (DMSO)-d₆ (39.50 ppm), THF-d₈ (67.20 ppm)] for ¹³C-NMR. The signal assignments were confirmed by two-dimensional (2D)-NMR analyses: ¹H–¹H 2D correlation spectroscopy (COSY), ¹H–¹³C heteronuclear multiple-quantum coherence (HMQC), and ¹H–¹³C heteronuclear multiple-bond connectivity (HMBC). Microanalyses were performed with a Yanaco MT-6 CHN corder. Routine monitoring of reactions was carried out using precoated Kieselgel 60F₂₅₄ plates (E. Merck). Microwave irradiation experiments were carried out in a CEM Discover Focused Microwave System. For the separation of crude products in small experimental scales (below 2 mmol), we applied Centrifugal chromatography separation of the reaction products on silica gel (Kanto 60N or Able-Biott) with a UV detector. Commercially available starting materials were used without further purification, and dry solvents were used in all reactions.

General Procedure for the Preparation of C₃-Symmetrical Benzylamine-Type TAZ Derivatives (4) (Table 1): Example: Synthesis of N²,N⁴,N⁶-Tribenzyl-1,3,5-triazine-2,4,6-triamine (4a) (Entry 1)

To a solution of TCT AZ (1, 277 mg, 1.50 mmol) in dioxane (2.5 mL), a solution of benzylamine (2a, 1.61 g, 15.0 mmol) in dioxane (1.5 mL) was added dropwise at 0°C, and the mixture was continuously stirred for 30 min at room temperature and then for another 1 h at room temperature. Then the mixture was subjected to microwave irradiation (MW) at 100°C (100 W) for 60 min with stirring. After addition of water (30 mL), the mixture was extracted with EtOAc (2×25 mL). The combined organic layer was dried over MgSO₄. After evaporation of the solvent, the residual solid was purified by centrifugal chromatography (n-hexane–EtOAc=50 : 50) to give an analytically pure 4a (507 mg, 1.28 mmol, 85%) as a pale yellow solid.

4a: mp 103.0–104.5°C (from 2-PrOH). IR cm⁻¹: 3405, 3249 (NH), 1536, 1500 (C=N), 1163 (C–N). ¹H-NMR (CDCl₃) δ: 4.53 (6H, br s, CH₂), 5.39 (3H, br s, NH), 7.18–7.32 (15H, m, H, C2′, 3′, 4′, 5′, 6′), ¹³C-NMR (CDCl₃) δ: 44.56 (ArCH₂), 127.02 (C4′), 127.51 (C2′, 6′), 128.44 (C3′, 5′), 139.45 (C1′), 166.27 (C2, 4, 6). Positive-ion FAB-MS m/z: 397 (M+H⁺). HR-FAB-MS m/z: 397.2147 (Calcd for C₂₄H₂₅N₆: 397.2147). Anal. Calcd for C₂₄H₂₄N₆: C, 72.70; H, 6.10; N, 21.20. Found: C, 72.70; H, 6.19; N, 21.05.

N²,N⁴,N⁶-Tris(4-methylbenzyl)-1,3,5-triazine-2,4,6-triamine (4b) (Entry 2)

Compound 4b was prepared from the reaction of 1 with 4-methylbenzylamine (2b) under the conditions shown in Table 1. Purification by centrifugal chromatography (CH₂Cl₂–EtOH=97 : 3) afforded 4b (53%) as a white solid. An analytical sample of 4b was obtained as white crystals by concentration of the filtrate.

4b: mp 116–117°C (from 2-PrOH). IR cm⁻¹: 3440 (NH), 1575, 1504 (C=N), 1346, 1245 (C–N), 808 (1,4-disubstituted benzene). ¹H-NMR (CDCl₃) δ: 1.83 (1.6H, br s, NH), 2.31 (9H, s, CH₃), 4.50 (6H, br s, Hα), 5.17 (0.6H, br s, NH), 5.27 (0.8H, br s, NH), 7.08 (6H, d, J=7.3 Hz, H3′, 5′), 7.16 (6H, d, J=7.3 Hz, H2′, 6′). ¹³C-NMR (CDCl₃) δ: 21.06 (CH₃), 44.40 (Cα), 127.54 (C2′, 6′), 129.13 (C3′, 5′), 136.40 (C1′), 136.62 (C4′), 166.28 (C2, 4, 6). Positive-ion FAB-MS m/z: 439 (M+H)⁺. HR-FAB-MS m/z: 439.2603 (Calcd for C₂₇H₃₁N₆: 439.2610). Anal. Calcd for C₂₇H₃₀N₆: C, 73.94; H, 6.89; N, 19.16. Found: C, 73.91; H, 6.98; N, 19.07.

6-Chloro-N²,N⁴-bis(4-methylbenzyl)-1,3,5-triazine-2,4-diamine (3b) (Entry 3)

Compound 3b was obtained from the reaction of 1 and 2b in THF (ratio of 1 : 2b=1 : 6) under the conditions shown in Table 1 in 46% yield after recrystallization from 2-PrOH as a white solid together with 4b (5%) obtained by concentration of the filtrate.

3b: mp 235–237°C (from 2-PrOH). IR cm⁻¹: 3248 (NH), 1541, 1511 (C=N), 1343, 1091 (C–N and/or C–O), 796, 582 (C–Cl). ¹H-NMR (THF-d₈) δ: 2.27, 2.28* (6H, s, CH₃), 4.44–4.49 (4H, m, CH₂), 4.71 (0.4H, br s, NH), 7.01–7.09 (4H, m, H3′, 5′), 7.11–7.19 (4H, m, H2′, 6′), 7.29 (0.8H, br s, NH), 7.40 (0.8H, br s, NH). ¹³C-NMR (THF-d₈, 67.20) δ: 20.93 (CH₃), 44.58, 44.75, 44.80* (CH₂), 127.85, 128.14*, 128.27 (C2′, C6′) 129.47 (C3′, 5′), 136.84*, 136.90, 136.98, 137.02 (C4′), 137.13, 137.16* (C1′), 166.93, 167.08*, 167.24 (C2, 4), 169.58*, 170.10* (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 354 (M+H)⁺. HR-FAB-MS m/z: 354.1482 (Calcd for C₁₉H₂₁ClN₅: 354.1485). Anal. Calcd for C₁₉H₂₀ClN₅: C, 64.49; H, 5.70; N, 19.79. Found: C, 64.47; H, 5.76; N, 19.72.

N²,N⁴,N⁶-Tris(4-chlorobenzyl)-1,3,5-triazine-2,4,6-triamine (4c) [112484-31-8] (Entry 4)

Compound 4c was prepared from the reaction of 1 with 4-chlorobenzylamine (2c) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=97 : 3) gave 4c (75%) as a white solid. An analytical sample of 4c was obtained as colorless crystals by recrystallization from propionitrile (EtCN).

4c: mp 156–158°C (from EtCN). IR cm⁻¹: 3396, 3246 (NH), 1526 (C=N), 1330, 1161 (C–N). ¹H-NMR (DMSO-d₆) δ: 4.31 (3.3H, br s, Hα of tautomer B), 4.40 (2.7H, d, J=5.5 Hz, Hα of tautomer A), 7.05 (0.6H, br s, NH of A), 7.15 (3.3H, br s, H2′, 6′ of B), 7.19 (1.2H, br s, NH of A and B), 7.23 (3.3H, br s, H3′, 5′ of B), 7.28 (2.7H, d, J=7.6 Hz, H2′, 6′ of A), ca. 7.32 (1.2H, br s, NH of B), 7.34 (2.7H, d, J=7.6 Hz, H3′, 5′ of A). (The integral values of the peaks at δ_H 7.15–7.34 were calculated from the spectral data after addition of D₂O.) ¹³C-NMR (DMSO-d₆) δ: 42.37 (Cα of A), 42.66 (Cα of B), 127.88 (C3′, C5′ of B), 127.98 (C3′, C5′ of A), 128.80 (C2′, 6′ of A), 128.96 (C2′, 6′ of B), 130.82 (C4′ of A and B), 139.86 (C1′ of A and B), 165.72 (C2, 4, 6 of A), 165.89 (C2, 4, 6 of B). Positive-ion FAB-MS m/z: 499 (M+H⁺). HR-FAB-MS m/z: 499.0975 (Calcd for C₂₄H₂₂Cl₃N₆: 499.0972). Anal. Calcd for C₂₄H₂₁Cl₃N₆: C, 57.67; H, 4.23; N, 16.81. Found: C, 57.63; H, 3.97; N, 16.81.

N²,N⁴,N⁶-Tris[4-(trifluoromethyl)benzyl]-1,3,5-triazine-2,4,6-triamine (4d) (Entry 5)

Compound 4d was prepared from the reaction of 1 with 4-(trifluoromethyl)benzylamine (2d) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (n-hexane–EtOAc=30 : 70) gave 4d (42%) as a white solid. An analytical sample of 4d was obtained as a white solid by recrystallization from 2-PrOH.

4d: mp 128–130°C (from 2-PrOH). IR cm⁻¹: 3462, (NH), 1530, 1502 (C=N), 1323, 1105, 1066 (C–N and/ or C–O). ¹H-NMR (DMSO-d₆) δ: 4.39, 4.42*, 4.51, 4.52 (6H, br s, Cα), 7.19*, 7.42 (3H, br s, NH), 7.31, 7.48* (6H, br s, C2′, 6′), 7.49, 7.66* (6H, d, J=8.2 Hz, C3′, 5′). ¹³C-NMR (DMSO-d₆) δ: 42.74, 42.97* (Cα), 124.27* (q, J=273.1 Hz, CF₃) 124.39 (q, J=271.7 Hz, CF₃), 124.74*, 124.96 (C3′, 5′), 127.02*, 127.11 (q, J=31.8 Hz, C4′), 127.55, 127.67* (C2′, C6′), 145.66, 145.77* (C1′), 165.80, 166.00* (C2, 4, 6). (The observed signals of the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 601 (M+H)⁺. HR-FAB-MS m/z: 601.1763 (Calcd for C₂₇H₂₂F₉N₆: 601.1762). Anal. Calcd for C₂₇H₂₁F₉N₆·0.7H₂O: C, 52.89; H, 3.68; N, 13.71. Found: C, 53.06; H, 3.38; N, 13.69.

N²,N⁴,N⁶-Tris(4-methoxybenzyl)-1,3,5-triazine-2,4,6-triamine (4e) (Entry 6)

Compound 4e was prepared from the reaction of 1 with 4-methoxybenzylamine (2e) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=97 : 3) gave 4e (66%) as a white solid. An analytical sample of 4e was obtained as a white solid by recrystallization from EtOH.

4e: mp 135–136°C (from EtOH). IR cm⁻¹: 3439, 3367 (NH), 1556, 1499 (C=N), 1345, 1246, 1171, 1030 (C–N or C–O), 810 (1,4-disubstituted benzene). ¹H-NMR (DMSO-d₆) δ: 3.70 (9H, s, OCH₃), 4.33 (6H, br s, Hα), 6.77 (3H, br s, H3′, 5′ of B), 6.81–6.87^† (1H, NH), 6.83 (3H, d, J=6.9 Hz, H3′, 5′ of A), 7.01 (1.6H, br s, NH), 7.13 (3H, br s, H2′, 6′ of B), 7.18 (3H, d, J=6.9 Hz, H2′, 6′ of A), 7.16–7.23^† (0.4H, NH). (The peaks with the mark^† attached are assumed to be overlapped with the peaks of NH.) ¹³C-NMR (DMSO-d₆) δ: 42.40, 42.67* (Cα) 54.93 (OCH₃), 113.37 (C3′, 5′), 128.21 (C2′, 6′ of A), 128.51* (C2′, 6′ of B), 132.80*, 132.90 (C1′), 157.86 (C4′), 165.74*, 165.86 (C2, 4, 6). (Signals for only the predominant tautomer B are asterisked.) Positive-ion FAB-MS m/z: 487 (M+H)⁺. HR-FAB-MS m/z: 487.2460 (Calcd for C₂₇H₃₁N₆O₃: 487.2458). Anal. Calcd for C₂₇H₃₀N₆O₃: C, 66.65; H, 6.21; N, 17.27. Found: C, 66.47; H, 6.15; N, 17.19.

N²,N⁴,N⁶-Tris(3,4-dimethoxybenzyl)-1,3,5-triazine-2,4,6-triamine (4f) (Entry 7)

Compound 4f was prepared from the reaction of 1 with 3,4-dimethoxybenzylamine (2f) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=95 : 5) gave 4f (80%) as a white solid. An analytical sample of 4f was obtained as a colorless solid by recrystallization from 2-PrOH. Compound 4f was converted into hydrochloride (4f·HCl) as colorless crystals by a routine procedure.

4f: mp 47°C (from 2-PrOH). IR cm⁻¹: 3378 (NH), 1501 (C=N), 1257, 1230, 1134, 1023 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: 1.80 (1.8H, br s, NH), 3.85 (9H, s, OCH₃ on C3′), 3.87 (9H, s, OCH₃ on C4′), 4.53 (6H, br s, ArCH₂), 5.19 (0.5H, br s, NH), 5.33 (0.7H, br s, NH), 6.80 (3H, d, J=7.6 Hz, H5′), 6.85 (3H, d, J=7.6 Hz, H6′), 6.86 (3H, s, H2′). ¹³C-NMR (CDCl₃) δ: 44.54 (ArCH₂), 55.82, 55.90 (OCH₃), 110.90 (C2′), 111.09 (C5′), 119.77 (C6′), 131.86 (C1′), 148.20 (C4′), 149.01 (C3′), 166.17 (C2, 4, 6). ¹H-NMR (DMSO-d₆) δ: 3.65, 3.70* (18H, br s, OCH₃ on C3′, 4′), ca. 3.7 (1H, br s, NH), 4.34, 4.35* (6H, br s, ArCH₂), 6.72–6.97 (9H, m, ArH), [7.02 (1.6H), 7.17 (0.4H)] (br s, NH). ¹³C-NMR (DMSO-d₆) δ: 42.87, 43.13* (ArCH₂), 55.35, 55.49 (OCH₃ on C3′, 4′), 111.50 (C2′, 5′), 119.11, 119.49* (C6′), 133.29 (C1′), 147.48, 148.48 (C3′, 4′), 165.81 (C2, 4, 6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 577 (M+H)⁺. HR-FAB-MS m/z: 577.2784 (Calcd for C₃₀H₃₇N₆O₆: 577.2775). Anal. Calcd for C₃₀H₃₆N₆O₆·i-PrOH: C, 62.25; H, 6.97; N, 13.26. Found: C, 62.35; H, 6.89; N, 13.22.

4f·HCl: mp 128–131°C (from 2-PrOH–H₂O). IR cm⁻¹: 3254 (br, NH⁺), 1625 (NH₂⁺), 1567, 1515 (C=N), 1263, 1234, 1137, 1023 (C–N and/or C–O). ¹H-NMR (DMSO-d₆) δ: 3.69*, 3.73*, 3.75 (18H, br s, OCH₃ on C3′, 4′), ca. 3.7 (1H, br s, NH), 4.46*, 4.48 (6H, br s, ArCH₂), 6.75–7.15 (9H, m, ArH), 8.82*, 8.88, 9.03 (3H, br s, NH). ¹³C-NMR (DMSO-d₆) δ: 43.23*, 43.45, 43.76 (ArCH₂), 55.39, 55.48*, 55.52*, 55.54 (OCH₃), 111.60*, 111.71*, 111.83 (C2′, 5′), 119.81 (C6′), 130.10*, 130.48, 131.03 (C1′), 147.96, 148.03, 148.13*, 148.64* (C3′, 4′), 154.54*, 162.64 (C2, 4, 6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 577 (M+H)⁺. HR-FAB-MS m/z: 577.2766 (Calcd for C₃₀H₃₇N₆O₆: 577.2775). Anal. Calcd for C₃₀H₃₆N₆O₆·HCl·1.6H₂O : C, 56.13; H, 6.31; N, 13.09. Found: C, 56.13; H, 6.26; N, 13.05.

N²,N⁴,N⁶-Tris(2,4-dimethoxybenzyl)-1,3,5-triazine-2,4,6-triamine (4g) (Entry 8)

Compound 4g was prepared from the reaction of 1 with 2,4-dimethoxybenzylamine (2g) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=96 : 4) gave 4g (57%) as a white solid. An analytical sample of 4g was obtained as colorless crystals by recrystallization from EtOH.

4g: mp 137–138°C (from EtOH). IR cm⁻¹: 3444, 3421 (NH), 1556, 1499 (C=N), 1206, 1157, 1115, 1031 (C–N or C–O), 924, 810 (1,2,4-trisubstituted benzene). ¹H-NMR (CDCl₃) δ: 3.78 (9H, s, OCH₃ on C4′ or C2′), 3.80 (9H, s, OCH₃ on C2′ or C4′) 4.42, 4.52* (6H, br s, Hα), 5.05 (1H, br s, NH), 5.30 (2H, br s, NH), 6.36 (3H, br s, H5′), 6.43 (3H, br s, H3′), 7.16 (3H, d, J=8.3 Hz, H6′). ¹³C-NMR (CDCl₃) δ: 39.72 (Cα), 55.23 (OCH₃ on C2′ or C4′), 55.31 (OCH₃ on C4′or C2′), 98.32 (C3′), 103.65 (C5′), 120.16 (C1′), 130.08 (C6′), 158.45 (C4′ or C2′), 160.07 (C2′or C4′), 166.15 (C2, 4, 6). Positive-ion FAB-MS m/z: 577 (M+H)⁺. HR-FAB-MS m/z: 577.2789 (Calcd for C₃₀H₃₇N₆O₆: 577.2775). Anal. Calcd for C₃₀H₃₆N₆O₆: C, 62.49; H, 6.29; N, 14.57. Found: C, 62.57; H, 6.29; N, 14.58.

N²,N⁴,N⁶-Tris(benzo[d][1,3]dioxol-5-ylmethyl)-1,3,5-triazine-2,4,6-triamine (4h)⁵⁾

Compound 4h was prepared from the reaction of 1 with piperonylamine (2h) (see ref. 5).

N²,N⁴,N⁶-Tris(3,4,5-trimethoxybenzyl)-1,3,5-triazine-2,4,6-triamine (4i) (Entry 10)

Compound 4i was prepared from the reaction of 1 with 3,4,5-trimethoxybenzylamine (2i) under the conditions shown in Table 1. Separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=96 : 4) gave 4i (63%) as a white solid. An analytical sample of 4i was obtained as colorless crystals by recrystallization from 2-PrOH.

4i: mp 64–65°C (from 2-PrOH). IR cm⁻¹: 3363 (NH), 1590, 1560, 1498 (C=N), 1328, 1230, 1119 (C–N or C–O), 1003 (C–O), 811 (1,2,3,5-tetrasubstituted benzene). ¹H-NMR (CDCl₃) δ: 3.79 (18H, s, OCH₃ on C3′, 5′), 3.82 (9H, s, OCH₃ on C4′), 4.50 (6H, br s, Hα), 5.44, 5.50 (3H, br s, NH), 6.52 (6H, s, H2′, 6′). ¹³C-NMR (CDCl₃) δ: 44.93 (Cα), 55.99 (OCH₃ on C3′, 5′), 60.72 (OCH₃ on C4′), 104.44 (C2′, 6′), 134.88 (C1′), 137.03 (C4′), 153.25 (C3′, 5′), 166.13 (C2, 4, 6). Positive-ion FAB-MS m/z: 667 (M+H)⁺. HR-FAB-MS m/z: 667.3086 (Calcd for C₃₃H₄₃N₆O₉: 667.3092). Anal. Calcd for C₃₃H₄₂N₆O₉·0.5 2-PrOH·0.5H₂O: C, 58.71; H, 6.71; N, 11.91. Found: C, 58.57; H, 6.74; N, 11.78.

Preparation of Intermediate Di-benzylamino-Substituted TAZ Derivatives (3)

6-Chloro-N²,N⁴-bis(3,4-dimethoxylbenzyl)-1,3,5-triazine-2,4-diamine (3f)

This compound was prepared in a manner similar to that for the preparation of 3h described by Vidal-Mosquera et al.¹³⁾ To a solution of 1 (922 mg, 5.0 mmol) in THF (100 mL), 2f (3.34 g, 20.0 mmol) was added dropwise at room temperature, and then the mixture was refluxed for 1.2 h. Then the white solids were filtered, and water (200 mL) was added. After the mixture had been stirred at 60°C for 10 min, the solid was filtered and washed with EtOH (50 mL) to give analytically pure compound 3f (1.51 g, 4.29 mmol, 86%) as a white solid.

3f: mp 193–194. IR cm⁻¹: 3250 (NH), 1555, 1514 (C=N), 1240, 1137, 1021 (C–N and/or C–O), 799, 736 (C–Cl). ¹H-NMR (DMSO-d₆) δ: 3.67* (3.3H, s, OCH₃ on C3′), 3.71* (3.3H, s, OCH₃ on C4′), 3.72 (3.6H, br s, OCH₃), 3.73 (1.8H, s, OCH₃), 4.33–4.40 [4H, m, Cα: including 4.38* (d, J=6.2 Hz)], 6.72–6.96 [6H, m, ArH: including 6.73* (1.1H, dd, J=8.3, 1.4 Hz, H6′), 6.79* (ca. 1.1H, d, J=8.3 Hz, H5′), 6.89* (ca. 1.1H, d, J=1.4 Hz, H2′)], [8.05 (0.3H), 8.19 (0.3H), 8.23 (0.3H), 8.31* (1.1H)] (t, J=6.2 Hz, NH). ¹³C-NMR (DMSO-d₆) δ: 43.19, 43.28, 43.41* (Cα), [55.29* (on C3′), 55.35, 55.42* (on 5′), 55.51] (OCH₃), 111.38*, 111.51 (C2′), 111.67* (C5′), 119.22, 119.29, 119.36* (C6′), 131.43, 131.61* (C1′), 147.73*, 147.78 (C4′), 148.55,*, 148.59 (C3′), 165.23, 165.36*, 165.61 (C2, 4), 167.77*, 168.30 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 446 (M+H)⁺. HR-FAB-MS m/z: 446.1586 (Calcd for C₂₁H₂₅ClN₅O₄: 446.1595). Anal. Calcd for C₂₁H₂₄ClN₅O₄: C, 56.57; H, 5.43; N, 15.71. Found: C, 56.35; H, 5.42; N, 15.68.

6-Chloro-N²,N⁴-bis(benzo[d][1,3]dioxol-5-ylmethyl)-1,3,5-triazine-2,4-diamine (3h)¹³⁾

In a manner similar to that for the preparation of 3f, the reaction of 1 and 2f only by stirring for 30 min at room temperature, product 3h was obtained in 96% yield as a white solid.

3h: mp 225–226. IR cm⁻¹: 3244 (NH), 1558 (C=N), 1280, 1038 (C–N and/or C–O), 796 (C–Cl). ¹H-NMR (DMSO-d₆) δ: 4.31–4.35 [4H, m, Cα: including 4.33* (d, J=6.2 Hz)], 5.96*, 5.97, 5.98 (4H, s, H2′), 6.68-6.70 [1.1H, m, ArH: including 6.69* (br d, J=8.3 Hz, H6′)], 6.72–6.78 [3H, m, ArH: including 6.77* (d, J=8.3 Hz, H7′)], 6.83–6.88 [1.9H, m, ArH: including 6.85* (br s, H4′)], [8.08 (0.5H), 8.21 (1.1H), 8.24 (1.1H), 8.33 (3.3H)] (t, J=6.2 Hz, NH). ¹³C-NMR (DMSO-d₆) δ: 43.14, 43.23, 43.36* (Cα), 100.76 (C2′), 107.82, 107.86* (C7′), 107.90, 108.00* (C4′), 120.26, 120.37, 120.46* (C6′), 132.92, 133.07, 133.13* (C5′), 146.03 (C3a′), 146.15 (C7a′), 165.20, 165.30*, 165.56 (C2, 4), 167.81*, 168.33 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 414 (M+H)⁺. HR-FAB-MS m/z: 414.0974 (Calcd for C₁₉H₁₇ClN₅O₄: 414.0969). Anal. Calcd for C₁₉H₁₆ClN₅O₄·0.2H₂O: C, 54.67 H, 3.96; N, 16.78. Found: C, 54.63; H, 3.90; N, 16.84.

Preparation of N-Benzyl-4-chloro-6-isopropoxy-1,3,5-triazin-2-amine (6a) (Table 2, Entry 1)

Compound 6a was obtained from the reaction of 5b with 2a under the conditions shown in Table 2. Thus, to a solution of intermediate 5b (2.08 g, 10.0 mmol) in THF (150 mL), benzylamine 2a (1.07 g, 8.00 mmol) and diisopropylethylamine (DIPEA, 1.29 g, 10.0 mmol) were added and the reaction mixture was stirred for 2 h at 0°C. After removal of the precipitated solids (DIPEA·HCl salt), evaporation of the solvent gave crude compound 6a (2.71 g, 9.74 mmol, 97%) as a white powder. An analytically pure sample of 6a was obtained as a white powder by recrystallization from acetonitrile (MeCN).

6a: mp 127–128°C (from MeCN). IR cm⁻¹: 3262, 3110 (NH), 1638, 1527 (C=N), 1294, 1231, 1105 (C–N and/or C–O), 803 (C–Cl). ¹H-NMR (CDCl₃) δ: [1.31 (2H), 1.34* (4H)] (d, J=6.2 Hz, H2″, 3″), [4.65 (0.7H), 4.67* (1.3H)] (d, J=6.2 Hz, Hα), 5.20–5.30 (1H, m, H1″), [6.33 (0.3H), 6.78* (0.7H)] (br s, NH), 7.26–7.37 (5H, m, H2′–6′). ¹³C-NMR (CDCl₃) δ: 21.63*, 21.67 (C2″, 3″), 44.99*, 45.07 (Cα), 71.66, 72.02* (C1″), 127.33, 127.62 (C2′, 6′) 127.66*, 127.72 (C4′), 128.70*, 128.74 (C3′, C5′), 137.45 (C1′), 167.07 (C2), 169.81, 170.33, 170.40, 171.47 (C4, 6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 279 (M+H)⁺. HR-FAB-MS m/z: 279.0981 (Calcd for C₁₃H₁₆ClN₄O: 279.1013). Anal. Calcd for C₁₃H₁₅ClN₄O: C, 56.02; H, 5.42; N, 20.10. Found: C, 55.96; H, 5.43; N, 19.93.

General Procedure for the Preparation of C_S-Symmetrical Benzylamine-Type TAZ Derivatives (7) (Table 2): Example 1 (No Base by MW): Synthesis of 6-Isopropoxy-N²,N⁴-bis(4-methylbenzyl)-1,3,5-triazine-2,4-diamine (7b) (Entry 3)

To a solution of intermediate 5b¹⁴⁾ (416 mg, 2.0 mmol) in dioxane (4 mL) was added 2b (1.21 g, 10.0 mmol) at room temperature. Then the mixture was subjected to MW at 100°C (100 W) for 45 min with stirring. After addition of water (50 mL), the mixture was extracted with EtOAc (2×50 mL). The combined organic layer was washed with saturated aqueous NH₄Cl solution and dried over MgSO₄. After evaporation of the solvent, the residue was recrystallized from 2-PrOH to obtain analytically pure compound 7b as a colorless solid.

7b: mp 142–143°C (from 2-PrOH). IR cm⁻¹: 3347, 3239 (NH), 1609, 1579 (C=N), 1158, 1104 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.27 (br s), 1.31* (d, J=6.2 Hz)] (6H, H2″, 3″), 2.32 (6H, br s, CH₃), 4.51, 4.55* (4H, br s, Hα), [5.13 (0.3H), 5.23 (0.7H)] (br s, H1″), [5.38, 5.45 (0.6H), 5.61 (1.4H)] (br s, NH), 7.11 (4H, d, J=6.9 Hz) (H3′, 5′), 7.17 (4H, d, J=6.9 Hz) (H2′, 6′). ¹³C-NMR (CDCl₃) δ: 21.06 (CH₃), 21.94 (C2″, 3″), 44.40, 44.50* (Cα), 69.02*, 69.43 (C1″), 127.38, 127.53* (C2′, 6′), 129.18 (C3′, 5′), 135.83*, 136.10 (C1′), 136.82 (C4′), 167.07, 167.35* (C2, 4), 169.93*, 170.30 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 378 (M+H)⁺. HR-FAB-MS m/z: 378.2284 (Calcd for C₂₂H₂₈N₅O₄: 378.2294). Anal. Calcd for C₂₂H₂₈N₅O₄: C, 70.00; H, 7.21; N, 18.55. Found: C, 70.04; H, 7.22; N, 18.56.

N²,N⁴-Dibenzyl-6-isopropoxy-1,3,5-triazine-2,4-diamine (7a) (Entry 2)

Compound 7a was prepared from the reaction of 5b with 2a under the conditions shown in Table 2 using the above method. After the workup, separation of the products by centrifugal chromatography (CH₂Cl₂–EtOH=98 : 2) gave 7a (79%) as a white solid. An analytically pure sample of 7a was obtained as colorless crystals by recrystallization from diisopropylether (2-Pr₂O).

7a: mp 117–119°C (from 2-Pr₂O). IR cm⁻¹: 3336, 3246 (NH), 1615, 1508 (C=N), 1160, 1104 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.24 (2H, br s), 1.28* (4H, d, J=5.5 Hz)] (H2″, 3″), 4.55, 4.59* (4H, br s, Hα), [5.12 (0.3H), 5.21* (0.7H)] (br s, H1″), [5.53 (0.5H), 5.73 (0.2H), 5.84 (0.3H), 5.97* (1H)] (br s, NH), 7.20–7.35 (10H, m, H2′–6′). ¹³C-NMR (CDCl₃) δ: 21.89 (C2″, 3″), 44.53, 44.67* (Cα), 69.01, 69.48* (C1″), 127.11, 127.31, 127.49 (C2′, 6′, 4′), 128.46 (C3′, C5′), 138.96*, 139.21 (C1′), 167.15, 167.36* (C2, 4), 169.87, 170.21* (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 350 (M+H)⁺. HR-FAB-MS m/z: 350.1989 (Calcd for C₂₀H₂₄N₅O: 350.1981). Anal. Calcd for C₂₀H₂₃N₅O: C, 68.74; H, 6.63; N, 20.04. Found: C, 68.73; H, 6.74; N, 20.00.

6-Isopropoxy-N²,N⁴-bis(4-methoxybenzyl)-1,3,5-triazine-2,4-diamine (7e) (Entry 4)

Colorless needles. mp 132–134°C (from EtOH). IR cm⁻¹: 3392, 3261, 3101 (NH), 1583, 1530 (C=N), 1238, 1170, 1108, 1030 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.26 (2H, br s), 1.32* (4H, d, J=5.5 Hz)] (H2″, 3″), 3.78 (6H, br s, OCH₃ on C4′), 4.48, 4.53* (4H, br s, Hα), [5.13 (0.3H), 5.24* (0.7H)] (br s, H1″), [5.42 (0.3H), 5.55* (1.0H), 5.73 (0.7H)] (br s, NH), 6.83 (4H, d, J=7.6 Hz, H3′, 5′), 7.20 (4H, d, J=7.6 Hz, H2′, 6′). ¹³C-NMR (CDCl₃) δ: 21.93 (C2″, 3″), 44.07, 44.15* (Cα), 55.22 (OCH₃), 68.97*, 69.38 (C1″), 113.87 (C3′, 5′), 128.96, 128.79* (C2′, 6′), 130.93*, 131.27 (C1′), 158.79 (C4′), 166.97, 167.26* (C2, 4), 169.89*, 170.30 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 410 (M+H)⁺. HR-FAB-MS m/z: 410.2180 (Calcd for C₂₂H₂₈N₅O₃: 410.2192). Anal. Calcd for C₂₂H₂₇N₅O₃: C, 64.53; H, 6.65; N, 17.10. Found: C, 64.51; H, 6.67; N, 17.04.

N²,N⁴-Bis(3,4-dimethoxybenzyl)-6-isopropoxy-1,3,5-triazine-2,4-diamine (7f) (Entry 5)

Colorless crystals. mp 146–147°C (from MeCN). IR cm⁻¹: 3397, 3248 (NH), 1553, 1516 (C=N), 1262, 1162, 1136, 1026 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.28 (2H, br s), 1.33 (4H, d, J=4.8 Hz)] (H2″, 3″), 3.80–3.90 (12H, m, OCH₃ on C3′,4′), 4.56 (4H, br s, Hα), [5.15 (0.3H), 5.26 (0.7H)] (br s, H1″), [5.42 (1.4H), 5.59 (0.6H)] (br s, NH), 6.76–6.90 (6H, m, H2′, 5′, 6′). ¹³C-NMR (CDCl₃) δ: 21.94 (C2″, 3″), 44.62 (Cα), 55.82, 55.90* (OCH₃), 69.11*, 69.59 (C1″), 110.78 (C2′), 111.12 (C5′), 119.64, 119.74* (C6′), 131.34*, 131.57 (C1′), 148.28 (C4′), 149.04 (C3′), 167.02, 167.33* (C2, 4), 170.28*, 170.31 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 470 (M+H)⁺. HR-FAB-MS m/z: 470.2397 (Calcd for C₂₄H₃₂N₅O₅: 470.2403). Anal. Calcd for C₂₄H₃₁N₅O₅: C, 61.39; H, 6.65; N, 14.92. Found: C, 61.25; H, 6.79; N, 14.91.

N²,N⁴-Bis(2,4-dimethoxybenzyl)-6-isopropoxy-1,3,5-triazine-2,4-diamine (7g) (Entry 6)

Colorless crystals. mp 134–136°C (from 2-PrOH). IR cm⁻¹: 3417, 3248 (NH), 1612, 1592, 1526 (C=N), 1228, 1206, (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.22–1.29 (2H), 1.29–1.40 (4H)] (br s, H2″, 3″), [3.79 (8H), 3.82 (4H)] (s, OCH₃), [4.55, 4.51 (br s), 4.56* (d, J=5.5 Hz)] (4H, Hα), 5.11*, 5.26, 5.35 (1H, br s, H1″), 5.34, 5.39, 5.50* (2H, br s, NH), 6.34–6.47 (4H, m, H3′, 5′), 7.13–7.22 (2H, m, H6′). ¹³C-NMR (CDCl₃) δ: 21.99 (C2″, 3″), 40.03 (Cα), 55.25, 55.33 (OCH₃), 68.69*, 69.06 (C1″), 98.43 (C3′), 103.67 (C5′), 119.49*, 119.70 (C1′), 129.81, 130.10*, 130.25 (C6′), 158.50 (C2′), 160.29 (C4′), 166.95, 167.20* (C2, 4), 169.88*, 170.13 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 470 (M+H)⁺. HR-FAB-MS m/z: 470.2402 (Calcd for C₂₄H₃₂N₅O₅: 470.2403). Anal. Calcd for C₂₄H₃₁N₅O₅: C, 61.39; H, 6.65; N, 14.92. Found: C, 61.21; H, 6.72; N, 14.68.

N²,N⁴-Bis(benzo[d][1,3]dioxol-5-yllmethyl)-6-isopropoxy-1,3,5-triazin-2,4-diamine (7h) (Entry 7)

White solid. mp 143–145°C (from EtOAc). IR cm⁻¹: 3338, 3239, 3097 (NH), 1612, 1521 (C=N), 1246, 1232, 1102, 1038 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: 1.28, 1.32* (6H, d, J=6.2 Hz, H2″, 3″), 4.46, 4.50*, 4.51 (4H, br s, Hα), 5.14, 5.24* (1H, br s, H1″), 5.36, 5.42*, 5.58 (2H, br s, NH), 5.93 (4H, s, H2′), 6.70–6.85 (6H, m, H4′, 6′, 7′). ¹³C-NMR (CDCl₃) δ: 21.96 (C2″, 3″), 44.46, 44.58* (Cα), 69.14, 69.54* (C1″), 100.98 (C2′), 108.19 (C4′, 7′), 120.64 (C6′), 132.79*, 133.10 (C5′), 146.76 (C3a′), 147.79 (C7a′), 166.99*, 167.05 (C2, 4 or C6), 167.30 (C6 or C2, 4). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 438 (M+H)⁺. HR-FAB-MS m/z: 438.1779 (Calcd for C₂₂H₂₄N₅O₅: 438.1777). Anal. Calcd for C₂₂H₂₃N₅O₅: C, 60.40; H, 5.30; N, 16.01. Found: C, 60.42; H, 5.27; N, 16.00.

6-Isopropoxy-N²,N⁴-bis(3,4,5-trimethoxybenzyl)-1,3,5-triazine-2,4-diamine (7i) (Entry 8)

Colorless crystals. mp 148–150°C (from 2-PrOH). IR cm⁻¹: 3315, 3246 (NH), 1586, 1542 (C=N), 1225, 1159, 1334, 1099 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.29 (br s), 1.33 (d, J=4.8 Hz)] (6H, H2″, 3″), 3.82 (18H, br s, OCH₃), 4.53*, 4.56 (4H, m, Hα), [5.16 (0.25H, br s), 5.26* (0.75H, br s)] (H1″), [5.45*, 5.52 (1.5H), 5.67 (0.5H)] (br s, NH), 6.54 (4H, br s, H2′, 6′). ¹³C-NMR (CDCl₃) δ: 21.91 (C2″, 3″), 40.91, 44.93, 44.98, 45.09* (Cα), 56.07 (OCH₃ on C3′, 5′), 60.79 (OCH₃ on C4′), 69.24*, 69.60 (C1″), 104.39*, 104.70 (C2′, 6′), 134.48*, 134.66 (C1′), 137.16 (C3′, 5′), 153.34 (C4′), 167.11, 167.37* (C2, 4), 169.98*, 170.27 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 530 (M+H)⁺. HR-FAB-MS m/z: 530.2621 (Calcd for C₂₆H₃₆N₅O₇: 530.2615). Anal. Calcd for C₂₆H₃₅N₅O₇: C, 58.97; H, 6.66; N, 13.22. Found: C, 58.94; H, 6.74; N, 13.20.

N²,N⁴-Bis(3,4-dimethylbenzyl)-6-isopropoxy-1,3,5-triazine-2,4-diamine (7j) (Entry 9)

Colorless solid. mp 119–123°C (from EtOH). IR cm⁻¹: 3397, 3247, 3105 (NH), 1614, 1577, 1523 (C=N), 1166, 1146, 1104 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.28 (2H, br s), 1.32* (4H, d, J=5.5 Hz)] (H2″, 3″), 2.23 (12H, br s, CH₃), 4.49, 4.54* (4H, br s, Hα), [5.14 (0.3H), 5.25* (0.7H)] (br s, H1″), [5.38* (1.4H), 5.52 (0.6H)] (br s, NH), 7.02–7.12 (6H, m, H2′, 5′, 6′). ¹³C-NMR (CDCl₃) δ: 19.36 (CH₃ on C4′), 19.69 (CH₃ on C3′), 21.95 (C2″, 3″), 44.41, 44.52* (Cα), 68.99, 69.42* (C1″), 124.86, 125.04* (C6′), 128.74, 128.92* (C2′), 129.73 (C5′), 135.47 (C4′), 136.22*, 136.48 (C1′), 136.71 (C3′), 167.03, 167.33* (C2, 4), 169.93, 170.21* (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 406 (M+H)⁺. HR-FAB-MS m/z: 406.2594 (Calcd for C₂₄H₃₂N₅O: 406.2607). Anal. Calcd for C₂₄H₃₁N₅O: C, 71.08; H, 7.70; N, 17.27. Found: C, 71.03; H, 7.70; N, 17.27.

6-Isopropoxy-N²,N⁴-bis(2,4,6-trimethylbenzyl)-1,3,5-triazine-2,4-diamine (7k) (Entry 10)

Colorless crystals. mp 146–147°C (from EtOH). IR cm⁻¹: 3252, 3105, 2975 (NH), 1619, 1577, 1519 (C=N), 1172, 1017 (C–N and/or C–O). ¹H-NMR (CDCl₃) δ: [1.26 (d, J=5.5 Hz), 1.35*, 1.43 (br s)] (6H, H2″, 3″), 2.26, 2.27, 2.31* (6H, br s, CH₃ on C4′), 2.36, 2.37 (12H, br s, CH₃ on C2′, 6′), 4.42, 4.55, 4.58*, 4.66 (4H, br s, Hα), 4.68, 4.86* (2H, br s, NH), [5.09 (0.25H), 5.24* (0.6H), 5.43 (0.15H)] (br s, H1″), 6.86, 6.88 (4H, br s, H3′, 5′). ¹³C-NMR (CDCl₃) δ: 19.56 (CH₃ on C2′, 6′), 20.87 (CH₃ on C4′), 21.96 (C2″, 3″), 39.31*, 39.43 (Cα), 68.95, 69.38*, 69.79 (C1″), 129.06 (C3′, 5′), 131.16*, 131.41 (C1′), 137.35 (C2′, 4′, 6′), 166.48, 166.76, 167.10, 167.32* (C2, 4), 169.79, 170.17*, 170.70 (C6). (The observed ¹H- and ¹³C-signals assignable to the predominant tautomer are asterisked.) Positive-ion FAB-MS m/z: 434 (M+H)⁺. HR-FAB-MS m/z: 434.2905 (Calcd for C₂₆H₃₆N₅O: 434.2920). Anal. Calcd for C₂₆H₃₅N₅O: C, 72.02; H, 8.14; N, 16.15. Found: C, 71.81; H, 8.34; N, 16.00.

Antiviral Activity Assay and Cytotoxicity Assay of Synthesized TAZ Derivatives

The anti-HSV-1 activities (EC₅₀) of the synthesized TAZ derivatives were measured by using a plaque reduction assay⁹⁾ and their cytotoxicity against Vero cells (CC₅₀) was also evaluated as our previous description.⁵⁾ The results are summarized in Table 3 together with data for aciclovir.¹⁵⁾ Calculated log P values¹⁰⁾ for the compounds are also shown in Supplementary Materials (Fig. S1). There were few distinct correlations between log P values by using ChemBioDraw Ultra v.14.0 and anti-HSV-1 activity (EC₅₀).

Acknowledgment

The authors would like to thank Ms. Kanae Yamada, Ms. Shoko Tomonaga and Ms. Junko Matsuyama for their valuable technical assistance.

Conflict of Interest

The authors declare no conflict of interest.

Supplementary Materials

The online version of this article contains supplementary materials.

References and Notes

• 1) Clercq E. D., “Advances in Pharmacology Vol 67: Antiviral Agents,” Elsevier Academic Press, London, 2013, and related references cited therein.
• 2) Duran A. M., Meiler J., Comput. Struct. Biotechnol. J., 8, e201308004 (2013).
• 3) Contreras J.-M., Sippl W., “The Practice of Medicinal Chemistry” 3rd ed., ed. by Wermuth C. G., Elsevier Academic Press, London, 2008, pp. 380–414 and related references cited therein.
• 4) Mibu N., Yokomizo K., Koga A., Honda M., Mizokami K., Fujii H., Ota N., Yuzuriha A., Ishimaru K., Zhou J., Miyata T., Sumoto K., Chem. Pharm. Bull., 62, 1032–1040 (2014).
• 5) Mibu N., Yokomizo K., Yuzuriha A., Otsubo M., Kawaguchi Y., Sano M., Sakai I., Nakayama K., Zhou J.-R., Sumoto K., Heterocycles, 94, 1653–1677 (2017) and related references cited therein.
• 6) Some of the tri-substituted TAZ derivatives described in this paper showed a complicated ¹³C-NMR signal pattern because of their keto-enol tautomeric isomers in solutions. Compounds (3b, 4c–e, 6a, 7a, 7b, 7e–k) that have sec-benzylamino groups on a TAZ template are examples of TAZ derivatives with such a signal pattern. These compounds showed the presence of two tautomeric isomers in solution. Thus, signals that appeared in the ¹³C-NMR spectra of C₃-type compounds (4c–e) are fully consistent with the tautomeric mixture of two isomers shown below (A and B). We showed examples of ¹H- and ¹³C-NMR spectra of some tri-substituted TAZ derivatives (3–7) in Supplementary Materials (Fig. S2) for readers’ convenience.
• 7) For example, see: Cooke G., Rotello V. M., Chem. Soc. Rev., 31, 275–286 (2002) and also see the ref. 8.
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• 10) Calculated log P values were calculated by using ChemBioDraw Ultra v.14.0.
• 11) Mibu N., Yokomizo K., Murakami K., Ono Y., Ishimaru M., Otsubo M., Inao H., Ono Y., Zhou J.-R., Sumoto K., Chem. Pharm. Bull., 64, 1769–1780 (2016).
• 12) We have recently reported the results of calorimetric experiments on sugar recognition properties of this compound. J. Therm. Anal. Calorim., (2017), submitted.
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