Azetidin-2-ones: structures of antimitotic compounds based on the 1-(3,4,5-trimethoxyphenyl)azetidin-2-one core

Five azetidin-2-ones are described based on the 1-(3,4,5-trimethoxyphenyl)azetidin-2-one core with different substituents on the lactam 3 and 4 positions: (1) 3-(4-fluorophenyl)-4-(4-methoxyphenyl); (2) 3-(furan-2-yl)-4-(4-methoxyphenyl); (3) 3-(naphthalen-1-yl)-4-(4-methoxyphenyl); (4) 3-(3,4-dimethoxyphenyl)-4-(4-methoxyphenyl); (5) 4,4-bis(4-methoxyphenyl)-3-phenyl-1-(3,4,5-trimethoxyphenyl)azetidin-2-one.


Chemical context
-Lactam antibiotics e.g. penicillins, cephalosporins, carbapenems and monobactams, are based on a core -lactam ring structure and play a significant role in the clinical treatment of bacterial infections (Kong et al., 2010). Their mechanism of action is by targeting the transpeptidase enzymes (penicillinbinding proteins), which are required for bacterial cell-wall synthesis. However, because of extensive use, many bacteria have developed resistance to -lactam antibiotics. Additionally, the antiproliferative activity of compounds containing the -lactam (azetidin-2-one) ring structure has been investigated (Zhou et al. 2018;Galletti et al. 2014;Geesala et al., 2016;Arya et al., 2014;Fu et al., 2017). We have previously demonstrated the effectiveness of 1,4-diarylazetidin-2-ones in breast-cancer cell lines as tubulin-targeting antimitotic agents and selective estrogen-receptor modulators (SERMs; O'Boyle et al., 2014).
To further increase our library of -lactam antimitotic compounds, we have investigated the systematic synthesis and activity of a range of different -lactams based on the 1-(3,4,5trimethoxyphenyl) -lactam core (O'Boyle et al. 2010(O'Boyle et al. , 2011a. The five structurally characterized azetidin-2-ones reported herein have all been included in studies as tubulintargeting agents with mitotic catastrophe. They have all ISSN 2056-9890 displayed good antiproliferative effects in MCF-7 human breast-cancer cells, but tuning the substitution pattern in the aromatic ring C atoms has produced more efficacious azetidinones for further testing. The structural study of these compounds has been challenging, as the yields from synthesis were low, hence obtaining suitable crystalline samples was difficult. These structures will enable further modelling to improve the design of more effective -lactam antibiotics.

Structural commentary
Compound 1 crystallizes in the orthorhombic system, 2 and 4 in the monoclinic and 3 and 5 in the triclinic system. It is clear from the space group that these chiral molecules have crystallized as conformational racemates.
The molecules are shown in Figs. 1-5. Bond lengths and angles fall within reported limits. From Table 1 it can be seen that there are some commonalities in the structures, despite the differences in chirality and substituents. The common 3,4,5-trimethoxyphenyl rings and the carbonyl of the lactam display an intramolecular hydrogen bonds (C10Á Á ÁO17, see Table 2), which orient the A and B rings to be approximately co-planar with angles of 2.62 (13)-17.08 (9) between ring plane normals (see Table 1). The A and B rings can also twist and flex along the N1-C5 vector, which can also be seen in the C2-N1-C5-C10 torsion angle (see Table 1). See Fig. 6 for an overlay of similar conformations of 1-5 normal to the plane of the lactam.
It can also be seen that for both mono and di-substituted C4 lactams the angle between the lactam A and the C ring (C18-C23) is approximately orthogonal with angles ranging from 83.59 (9) to 89.56 (8) .
The conformations of the chiral centres at C3 in 1-5 are approximately eclipsed by geometry of the sp 3 carbons in the lactam ring with H3-C3-C4-C18 angles ranging from 0.98 in the R isomer of 5 (di-substituted in the 4 position -more steric requirements), to a wider 13.08 in 4. The conformation at C4 is also partially eclipsed with H4-C4-C3-C26 angles of 4.96 in 3 and the largest angle of 13.76 in 4 (see Table 1).
2.1. 3-(4-Fluorophenyl)-4-(4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)azetidin-2-one, 1 There is a single molecule of 1 in the asymmetric unit in the orthorhombic centrosymmetric space group Pbca, see Fig. 1. The compound is a racemate and the relative stereochemistry shown is 3S, 4R. In this compound, the A|D ring plane normal angle is close to 90 (Table 1). A similar, recently published structural isomer (Malebari et al., 2020; CSD refcode PUKNUH) is also a racemate and has two independent enantiomers in the asymmetric unit. The major structural difference between 1 and PUKNUH is the orientation of the trimethoxyphenyl ring plane to the lactam ring [8.87 (4) plane normals for lactam N1 in PUKNUH, with the same chirality]. It can also be seen in the C2-N1-C5-C10 torsion angle of À4.3 (3) for 1 (see Table 1) and 11.9 (2) for the N1 lactam in PUKNUH where, in spite of the hydrogen bond between the ring and the lactam carbonyl, the substituted B rings are orientated differently and the 4-methoxy groups on this ring are oriented in opposite directions (see Fig. S1 in the supporting information).
2.2. 3-(Furan-2-yl)-4-(4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)azetidin-2-one, 2 Compound 2 has two independent molecules in the asymmetric unit in the monoclinic centrosymmetric system P2 1 /c, and only one molecule is displayed in Fig. 2. In this racemic compound, both trans diastereomers are seen and the relative stereochemistry is 3S, 4S for the lactam with N1 and 3R, 4R for the lactam with N1A. See Table 1 for the geometric parameters. A comparison of the two independent molecules in 2 show similar differences as seen above -differences in the orientation of ring B to the lactam A ring (See Table 1 for A|B ring plane normals and the torsion angle C2-N1-C5 -C10) as well as the difference in orientation the 4-methoxy group position on the B ring (see Fig. S2 in the supporting information). The other notable difference is the orientation of the D rings to the lactam. In the N1 molecule (relative stereochemistry 3S, 4S) the A|D plane normals angle is approximately orthogonal (see Table 1). However in the other conformation (N1A, relative stereochemistry 3R, 4R) this A|D angle is much more acute. The twist of the group is also reflected in the torsion angles C2-C3-C26-O27 = À43.8 (3) and C2A-C3A-C26A-O27A = 180.0 (2) . There are no significant interactions to the furan directing this change.

Figure 3
Molecular structure of 3, relative stereochemistry 3S, 4R, with displacement ellipsoids drawn at the 50% probability level. Hydrogen atoms shown as spheres of arbitrary radius.

Figure 4
Molecular structure of 4, relative stereochemistry 3S, 4R, with displacement ellipsoids drawn at the 50% probability level. Hydrogen atoms shown as spheres of arbitrary radius.
C10 torsion angles are small, and the C and D rings are essentially orthogonal to the lactam (see Table 1). Showing all the commonalities described above, the main difference in 4 is seen in the dihedral angle along the C3-C4 vector, as this  molecule displays the largest angle for H3-C3-C4-C26/  H4-C4-C3-C26 (see Table 1).
2.5. 3-Phenyl-4,4-bis(4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)azetidin-2-one, 5 Compound 5, with two independent molecules, one of each enantiomer in the asymmetric unit in the triclinic space group P1, is a more unusual -lactam in that there are two identical substituents on the 4 position, see Fig. 5 where only one of the racemic molecules is shown. The torsion angles H3-C3-C4-C18 and C26-C4-C3-C34 [À5.62 (2) and 2.56 (2) ] in both enantiomers show that the arrangement is the most eclipsed among 1-5. Compound 5 also shows the largest A|B plane normal angles, indicating a bending along the N1-C5 vector and the trimethoxy ring and lactam are twisted as seen in the large C2-N1-C5-C10 torsion angles (Table 1). While showing all the common features outlined above, this molecule displays a conformational difference in the 4-methoxy group on the B ring between each enantiomer, also seen in 2 and shown in Fig. S3. This is the only example of a 4,4 0disubstituted 1-(3,4,5-trimethoxyphenyl) lactam. As a result of steric requirements, the 4 and 4 0 substituents in both molecules show a substantial difference in A|C plane normals. Other non-bicyclic 4,4 0 -disubstituted lactams are known (see Database survey, Table 3). Only AHERUA, which has phenyl substitutents, shows similar steric demands with equivalent C2-N1-C5-C10 torsion angles of ca 10.7 and A|C plane normal angles of 81.066 (1) and 61.454 (1) . RIFYIO has different steric requirements with methoxycarbonylphenylethyl and acetyl groups on N1 and C3 respectively. C4 is diphenyl substituted with A|C angles of 76.79 (5) and 66.21 (5) .

Supramolecular features
As well as the intramolecular hydrogen-bonding pattern described above, with the number of methoxy groups present, there are many weak C-HÁ Á ÁO intermolecular interactions in 1-5. The most significant are shown in Table 2. A motif seen in 1 is an association with two opposing methoxy groups, see Fig. 7, which form a 'buckle' to join two molecules together.
In 2 a 'double buckle' is present due to a bifurcated hydrogen bond between C12 and O11/O13 of an adjacent Molecular structure of one of the unique molecules in the asymmetric unit of 5, relative stereochemistry 3R, with displacement ellipsoids drawn at the 50% probability level. Hydrogen atoms shown as spheres of arbitrary radius.

Figure 6
Overlay of similar diastereomers of 1--5 normal to the plane of the lactam. The N1, C2 and O17 atoms were used as overlay centres. The flexibility in orientation of the B ring relative to the A ring (lactam) is clearly seen, as well as the substituents of the C and D rings.
In 4 the molecules do not associate via the 'buckle' and instead form an end-to-end hydrogen-bonded cyclic dimer with a bifurcated hydrogen bond, see Fig. 9.
The 'buckle' association of 1 is also seen in both enantiomers in 5, with further C--HÁ Á ÁO interactions by the carbon of the methoxy group of one enantiomer interacting with opposite enantiomer ketone (Table 2). Adjacent like enantiomers are also linked via C-HÁ Á ÁO interactions with the C phenyl ring and the lactam ketone, forming an interconnected sheet parallel to the bc plane, see Fig. 10  Dimer hydrogen-bonding motif seen in 4, linked via a bifurcated methoxy-methoxy C-HÁ Á ÁO interaction. Only hydrogen atoms involved in intra-and intermolecular hydrogen bonding are shown. Dotted lines indicate hydrogen-bonding interactions. [Symmetry code: (i) Àx + 1, Ày + 1, Àz + 1].
Compound 3, with the naphthyl substituent, does not display the same supramolecular features. A weaker C-HÁ Á ÁO interaction from the chiral centre C4 to the oxygen on the central methoxy group, B ring, links the molecules into a cyclic dimer. These dimers are associated via further C-HÁ Á ÁO hydrogen bonding (Table 2)

Funding information
A postgraduate research award from Trinity College Dublin is gratefully acknowledged.

sup-1
Acta Cryst. For all structures, data collection: APEX3 (Bruker, 2017); cell refinement: SAINT (Bruker, 2016); data reduction: SAINT (Bruker, 2016); program(s) used to solve structure: SHELXT (Sheldrick, 2015a); program(s) used to refine structure: SHELXL (Sheldrick, 2015b); molecular graphics: OLEX2 (Dolomanov et al., 2009); software used to prepare material for publication: OLEX2 (Dolomanov et al., 2009).  Special details Geometry. All esds (except the esd in the dihedral angle between two l.s. planes) are estimated using the full covariance matrix. The cell esds are taken into account individually in the estimation of esds in distances, angles and torsion angles; correlations between esds in cell parameters are only used when they are defined by crystal symmetry. An approximate (isotropic) treatment of cell esds is used for estimating esds involving l.s. planes.

Fractional atomic coordinates and isotropic or equivalent isotropic displacement parameters
Hydrogen-bond geometry (Å, º)