Synthesis and Properties of 1,3-Disubstituted Ureas Containing (Adamantan-1-yl)(phenyl)methyl Fragment Based on One-Pot Direct Adamantane Moiety Inclusion

A one-stage method for the preparation of 1-[isocyanato(phenyl)methyl]adamantane containing a phenylmethylene fragment located between the adamantane fragment and the isocyanate group, and 1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane with additional methyl groups at the nodal positions of adamantane, with a yield of 95% and 89%, respectively, is described. The method includes the direct inclusion of an adamantane moiety through the reaction of phenylacetic acid ethyl ester with 1,3-dehydroadamantane or 3,5-dimethyl-1,3-dehydroadamantane followed by the hydrolysis of the obtained esters. The reaction of 1-[isocyanato(phenyl)methyl]adamantane with fluorine(chlorine)-containing anilines gave a series of 1,3-disubstituted ureas with 25–85% yield. 1-[Isocyanato(phenyl)methyl]-3,5-dimethyladamantane was involved in the reactions with fluorine(chlorine)-containing anilines and trans-4-amino-(cyclohexyloxy)benzoic acid to obtain another series of ureas with a yield of 29–74%. The resulting 1,3-disubstituted ureas are promising inhibitors of the human soluble epoxide hydrolase (hsEH).


Introduction
Lipophilic fragments of inhibitors of soluble epoxidhydrolase (sEH, E. C. 3.3.2.10), an enzyme located in the arachidonic cascade [1][2][3][4] and involved in the metabolism of epoxy fatty acids (arachidonic acid metabolites) to the corresponding vicinal diols by catalytic addition of water molecules, usually contain adamantane [2] or aromatic fragments [5] in their structure. Inhibition sEH allows for successfully fighting against kidney diseases [6], cardiovascular diseases, and diabetes [7]. However, there are no references in the literature on sEH inhibitors, the lipophilic part of which contains both adamantane and an aromatic moiety at the same time. Vazquez et al. considered the replacement of the adamantane fragment with compounds containing polycyclic hydrocarbons smaller or larger than adamantane. Inhibitory activity values (IC 50 ) for these compounds were 0.4-21.7 nM, indicating that sEH is able to accommodate inhibitors of very different sizes. However, it has been noted that the human liver microsomal stability of diamantane-containing inhibitors is lower than that of their corresponding adamantane counterparts [8].
One of our studies was devoted to the synthesis of symmetric 1,3-disubstituted diureas containing both an adamantane fragment and an aromatic ring in the lipophilic part [10]. Compounds containing bulky lipophilic fragments can be used to study features of inhibition between soluble epoxide hydrolases of different species due to the differences in the protein structures [11]. However, compounds containing in the right part the halogen-containing (F, Cl) anilines have not been previously obtained.
1,3-disubstituted ureas 8a, 8b and 10a-c were synthesized by method B from (±)-1-[adamantyl(phenyl)methyl]amine hydrochloride 6a and aromatic isocyanates 9a-d, as well as cyclohexyl isocyanate 9e, as the closest analog trans-4-amino-(cyclohexyloxy)benzoic acid 7a devoid of an oxophenylcarboxylic fragment (Scheme 4).  usually insoluble in ether while most of the amines and isocyanates as well as triethylamine are soluble. For the cases when the starting material is insoluble in ether, DMF is used. As for the water and alcohols, they cannot be used as solvents for this reaction because they will react with the isocyanates. Most of the other polar solvents such as ethyl acetate can dissolve the resulting ureas which makes isolation more difficult. Inorganic bases are also insoluble in most of the solvents suitable for this reaction. mine are soluble. For the cases when the starting material is insoluble in ether, DMF is used. As for the water and alcohols, they cannot be used as solvents for this reaction because they will react with the isocyanates. Most of the other polar solvents such as ethyl acetate can dissolve the resulting ureas which makes isolation more difficult. Inorganic bases are also insoluble in most of the solvents suitable for this reaction. used. As for the water and alcohols, they cannot be used as solvents for this reaction because they will react with the isocyanates. Most of the other polar solvents such as ethyl acetate can dissolve the resulting ureas which makes isolation more difficult. Inorganic bases are also insoluble in most of the solvents suitable for this reaction. In 1 H NMR spectra of compounds 8a-e obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a, the chemical shift of protons 1 NH is within the range of 6.64-6.92 ppm, and the proton signals 3 NH bound to anilines shift to a weaker field of 8.40-8.71 ppm, which is probably due to the close location of the electron-withdrawing phenyl substituent to the NH-group. In 1 H NMR spectra of compounds 8f-j obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane 5b, the signals of 1 NH proton shift to a strong field of 4.05 ppm compared to compounds 8a-e, which is probably due to the presence of electron-donating methyl substituents in the nodal positions of adamantane. The proton signals 3 NH bound to the phenyl substituent stay at the same range of 8.36-8.59 ppm, as for the compounds 8a-e. For the compound 10с obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a and cyclohexyl isocyanate 9e, in the absence of a phenyl substituent, the proton signal 3 NH shifts to a strong field of 6.37 ppm. Similarly to compounds 8a-j, the signal of a proton 1 NH shifts to a strong field of 5.73 ppm.
The calculated lipophilicity coefficient LogP for the series of ureas 8a-k is in the range of 5.96-6.93, which somewhat exceeds the allowable limits according to the Lipinski rule [17]. For a series of ureas 8f-j obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane 5b, the lipophilicity coefficient is 0.12 units higher than that of ureas 8a-e obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a (Table 1). Based on the literature data [11], such compounds will have a higher inhibitory activity, but have a lower solubility in water and are more susceptible to metabolism in vivo. Comparing urea 8k obtained from trans-4-amino-(cyclohexyloxy)benzoic acid with analogs, it can be seen that the lipophilicity coefficient also became 0.12 units higher than for its analog 11 not containing methylene substituents in the nodal positions of adamantane. Comparing urea 8k with previously obtained analogs 12, 13 containing a fragment of trans-4-amino-(cyclohexyloxy)benzoic acid, it can be seen that the introduction of substituents in the nodal positions or in the bridge separating the adamantane fragment and the ureide group leads to an increase in the lipophilicity coefficient by 1.70 units ( Table 1).
The introduction of methyl substituents in the nodal positions of adamantane made it possible to reduce the melting temperatures of the ureas 8f-j (51-196 °C) obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane 5b by 37-198 °C, in comparison with the melting temperatures of similar urea products 8a-e (233-270 °C) derived from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a. The general rule that lowering melting point increases solubility is based on the simplified solubility equation proposed by Wouters and Quéré [18]. Reduced melting point is also a positive factor for drug candidates as it simplifies preparation of drug dosage forms by hot-melt extrusion [19]. For Synthesis of 1,3-disubstituted urea 8a-k and 10a-c was carried out in an anhydrous diethyl ether medium for 12 h at room temperature in the presence of an equimolar amount of triethylamine (Table 1). Diethyl ether and triethylamine were chosen as the solvent and the base, respectively, for this reaction for a number of reasons. Ureas are usually insoluble in ether while most of the amines and isocyanates as well as triethylamine are soluble. For the cases when the starting material is insoluble in ether, DMF is used. As for the water and alcohols, they cannot be used as solvents for this reaction because they will react with the isocyanates. Most of the other polar solvents such as ethyl acetate can dissolve the resulting ureas which makes isolation more difficult. Inorganic bases are also insoluble in most of the solvents suitable for this reaction.
In 1 H NMR spectra of compounds 8a-e obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a, the chemical shift of protons 1 NH is within the range of 6.64-6.92 ppm, and the proton signals 3 NH bound to anilines shift to a weaker field of 8.40-8.71 ppm, which is probably due to the close location of the electron-withdrawing phenyl substituent to the NH-group. In 1 H NMR spectra of compounds 8f-j obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane 5b, the signals of 1 NH proton shift to a strong field of 4.05 ppm compared to compounds 8a-e, which is probably due to the presence of electron-donating methyl substituents in the nodal positions of adamantane. The proton signals 3 NH bound to the phenyl substituent stay at the same range of 8.36-8.59 ppm, as for the compounds 8a-e. For the compound 10c obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a and cyclohexyl isocyanate 9e, in the absence of a phenyl substituent, the proton signal 3 NH shifts to a strong field of 6.37 ppm. Similarly to compounds 8a-j, the signal of a proton 1 NH shifts to a strong field of 5.73 ppm.
The calculated lipophilicity coefficient LogP for the series of ureas 8a-k is in the range of 5.96-6.93, which somewhat exceeds the allowable limits according to the Lipinski rule [17]. For a series of ureas 8f-j obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5dimethyladamantane 5b, the lipophilicity coefficient is 0.12 units higher than that of ureas 8a-e obtained from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a (Table 1). Based on the literature data [11], such compounds will have a higher inhibitory activity, but have a lower solubility in water and are more susceptible to metabolism in vivo. Comparing urea 8k obtained from trans-4-amino-(cyclohexyloxy)benzoic acid with analogs, it can be seen that the lipophilicity coefficient also became 0.12 units higher than for its analog 11 not containing methylene substituents in the nodal positions of adamantane. Comparing urea 8k with previously obtained analogs 12, 13 containing a fragment of trans-4-amino-(cyclohexyloxy)benzoic acid, it can be seen that the introduction of substituents in the nodal positions or in the bridge separating the adamantane fragment and the ureide group leads to an increase in the lipophilicity coefficient by 1.70 units ( Table 1).
The introduction of methyl substituents in the nodal positions of adamantane made it possible to reduce the melting temperatures of the ureas 8f-j (51-196 • C) obtained from (±)-1-[isocyanato(phenyl)methyl]-3,5-dimethyladamantane 5b by 37-198 • C, in comparison with the melting temperatures of similar urea products 8a-e (233-270 • C) derived from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a. The general rule that lowering melting point increases solubility is based on the simplified solubility equation proposed by Wouters and Quéré [18]. Reduced melting point is also a positive factor for drug candidates as it simplifies preparation of drug dosage forms by hot-melt extrusion [19]. For urea 8k obtained from isocyanate 5b and trans-4-amino-(cyclohexyloxy)benzoic acid, the addition of methylene substituents to the nodal positions of adamantane leads to an increase in the melting temperature by 11 • C. Thus, the melting temperature of urea 8k is 174 • C, and for urea 11 obtained from isocyanate 5a, it is 163 • C. An increase in the melting temperatures by 58 • C is also observed in urea products obtained from 1-isocyanatomethyl-3,5-dimethyladamantane 12 and 1-isocyanatomethyladamantane 13 (Table 1). However, when comparing urea 8k obtained from isocyanate 5b with urea 12 obtained from 1-isocyanatomethyl-3,5-dimethyladamantane, the melting point decreases by 66 • C when introducing a phenyl substituent into the structure of isocyanate. A similar decline of melting temperatures by 19 • C is observed in urea 13 obtained from 1-[isocyanato(phenyl)methyl]adamantane 11 and 1-isocyanatomethyladamantane 8k (Table 1, Figure 1).
Molecules 2023, 28, x FOR PEER REVIEW 7 of 15 from (±)-1-[isocyanato(phenyl)methyl]adamantane 5a. The general rule that lowering melting point increases solubility is based on the simplified solubility equation proposed by Wouters and Quéré [18]. Reduced melting point is also a positive factor for drug candidates as it simplifies preparation of drug dosage forms by hot-melt extrusion [19]. For urea 8k obtained from isocyanate 5b and trans-4-amino-(cyclohexyloxy)benzoic acid, the addition of methylene substituents to the nodal positions of adamantane leads to an increase in the melting temperature by 11 °C. Thus, the melting temperature of urea 8k is 174 °C, and for urea 11 obtained from isocyanate 5a, it is 163 °C. An increase in the melting temperatures by 58 °C is also observed in urea products obtained from 1-isocyanatomethyl-3,5-dimethyladamantane 12 and 1-isocyanatomethyladamantane 13 (Table 1). However, when comparing urea 8k obtained from isocyanate 5b with urea 12 obtained from 1-isocyanatomethyl-3,5-dimethyladamantane, the melting point decreases by 66 °C when introducing a phenyl substituent into the structure of isocyanate. A similar decline of melting temperatures by 19 °C is observed in urea 13 obtained from 1-[isocyanato(phenyl)methyl]adamantane 11 and 1-isocyanatomethyladamantane 8k (Table 1, Figure 1).

Equipment
Purification of the obtained adamantyl-containing derivatives of phenylacetic acid ethyl ester 3a and 3b was performed on a Pure C-815 Flash Advanced chromatographic system (Buchi Labortechnik AG, Flawil, Switzerland).
Hydrolysis of the obtained adamantyl-containing derivatives of phenylacetic acid ethyl ester 3a and 3b was carried out on a Monowave 450 microwave laboratory reactor (Anton Paar GmbH, Graz, Austria).
The structure of the obtained compounds was confirmed by 1 H, 13 C, and 19 F NMR spectroscopy, chromatography-mass spectrometry, and elemental analysis. Mass spectra were recorded on an Agilent GC 7820A/MSD 5975 chromatography-mass spectrometer (Agilent Technologies, Santa Clara, CA, USA) in fullscan (EI) mode. 1

Conclusions
The one-stage insertion of an adamantane moiety through the reaction of 1,3dehydroadamantane and its 3,5-dimethyl homolog allowed us to obtain isocyanates containing a phenylmethylene fragment located between the adamantane fragment and the isocyanate group with a yield of 95% and 89%, respectively. The reaction of synthesized isocyanates with fluorine(chlorine)-containing anilines and trans-4-amino-(cyclohexyloxy)benzoic acid gave a series of 1,3-disubstituted ureas with 29-74% yield. The reaction of 1-[isocyanato-(phenyl)methyl]adamantane with fluoro(chlorine)-containing anilines gave a series of 1,3-disubstituted ureas with 25-85% yield. Inhibitory activity against sEH and other biochemical data for the synthesized compounds will be published in a further manuscript as soon as it can be acquired.