Abstract
The crystal structure of 8-((dimethylamino)methyl)-1-(3-(dimethylamino)propyl)-1,7,8,9-tetrahydrochromeno[5,6-d]imidazol-2-amine (1), an inhibitor of the hepatitis C virus internal ribosome entry site, is described and compared to the structure of the compound in complex with the viral RNA target. Compound 1 crystallized by pentane vapor diffusion into dichloroethane solution. It crystallized in the monoclinic system, P21/c space group with unit cell parameters a = 15.7950(5) Å, b = 14.0128(4) Å, c = 8.8147(3) Å, β = 94.357(2)° and a cell volume of 1945.34(11) A−3. Packing interactions in the small molecule crystal lattice correspond to key interactions of the compound with the viral RNA target.
Keywords: antiviral, benzimidazole, hepatitis C virus, RNA target, translation inhibitor
Introduction
Compounds containing the 2-aminobenzimidazole scaffold have been shown to inhibit the hepatitis C virus (HCV) by binding to the internal ribosome entry site (IRES) RNA and inducing a conformational change which selectively blocks viral protein synthesis [1]. The title compound 1, which is a potent inhibitor of HCV translation, has previously been used for co-crystallization with the viral RNA target and structure determination by X-ray crystallography [2]. The benzimidazole inhibitor 1 was synthesized over 9 steps from 2-chloro-6-hydroxybenzaldehyde (2) following a recently established route (Scheme 1) [3]. The title compound 1 was obtained as a racemic mixture whose crystal structure is described here and compared to the structure of the inhibitor in complex with the viral RNA target.
Scheme 1.
Synthesis of compound 1. Abbreviations: DABCO: 1,4-diazabicyclo[2.2.2]octane, EDC: 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide, DCM: dichloromethane, TFA: trifluoroacetic acid, NMP: N-methylpyrrolidone, THF: tetrahydrofurane.
Experimental
The title compound 1 was synthesized following a recently established route [3].
X-ray Crystallography
X-ray diffraction data of the title compound were collected on a Bruker Kappa diffractometer equipped with an APEX CCD II area detector using graphite-monochromated Cu Ka radiation (λ=1.54178 Å). A colorless plate (0.2 × 0.3 × 0.5mm) was mounted on a cryoloop with paratone oil. Data were collected in a nitrogen gas stream at 173(2) K using phi and omega scans. The data were integrated using the Bruker SAINT [4] software and scaled using the SADABS [5] program. Solution by direct methods (SHELXS) [6] produced complete phasing models consistent with the proposed structures which were refined by least square methods on F2 using the SHELXL-97 [6] program package. The refinement was continued until the maximum shift/e.s.d was 0.000. The final difference map was featureless with maximum and minimum electron densities at 0.192 and −0.167 eÅ−3. The crystal data, intensity collection conditions and refinement parameters are presented in Table 1. All non-hydrogen atoms were refined anisotropically by full-matrix least-squares. Selected bond lengths and bond angles are given in Tables 2 and 3. Hydrogen atoms H(14A) and H(14B) were allowed to refine freely. All other H atoms were located geometrically, with C–H = 0.95–1.0 Å and treated using a riding model, with isotropic U set to 1.2 times the isotropic equivalent of that of the attached parent atom. For the methyl group, the C–C–H angles and C–H distances were fixed, but the CH3 group was allowed to rotate about the C–CH3 bond.
Table 1.
Crystal data and structure refinement for 1.
| CCDC | 869034 | |
| Empirical formula | C18 H29 N5 O | |
| Formula weight | 331.46 | |
| Temperature | 173(2) K | |
| Wavelength | 1.54178 Å | |
| Crystal system | Monoclinic | |
| Space group | P2(1)/c | |
| Unit cell dimensions | a = 15.7950(5) Å | a= 90°. |
| b = 14.0128(4) Å | b= 94.357(2)°. | |
| c = 8.8147(3) Å | γ = 90°. | |
| Volume | 1945.34(11) Å3 | |
| Z | 4 | |
| Density (calculated) | 1.132 Mg/m3 | |
| Absorption coefficient | 0.577 mm−1 | |
| F(000) | 720 | |
| Crystal size | 0.50 × 0.30 × 0.20 mm3 | |
| Theta range for data collection | 2.81 to 67.31°. | |
| Index ranges | −17<=h<=18, −16<=k<=15, −8<=l<=10 | |
| Reflections collected | 9004 | |
| Independent reflections | 3209 [R(int) = 0.0283] | |
| Completeness to theta = 67.31° | 92.0 % | |
| Absorption correction | None | |
| Max. and min. transmission | 0.7529 and 0.6384 | |
| Refinement method | Full-matrix least-squares on F2 | |
| Data/restraints/parameters | 3209/0/261 | |
| Goodness-of-fit on F2 | 1.040 | |
| Final R indices [I>2sigma(I)] | R1 = 0.0525, wR2 = 0.1365 | |
| R indices (all data) | R1 = 0.0652, wR2 = 0.1497 | |
| Largest diff. peak and hole | 0.192 and −0.167 e.Å−3 | |
Table 2.
Bond lengths [Å] for 1.
| O(7)-C(8) | 1.380(3) | C(11)-C(10) | 1.384(3) |
| O(7)-C(5) | 1.383(3) | C(2)-C(8) | 1.399(3) |
| O(7)-C(6) | 1.610(11) | C(2)-C(3) | 1.513(3) |
| N(14)-C(13) | 1.349(3) | C(10)-C(9) | 1.381(3) |
| N(14)-H(14A) | 0.90(3) | C(9)-C(8) | 1.403(3) |
| N(14)-H(14B) | 0.89(3) | C(3)-C(4) | 1.514(3) |
| N(19)-C(20) | 1.465(3) | N(24)-C(26) | 1.376(4) |
| N(19)-C(21) | 1.464(3) | N(24)-C(25) | 1.409(4) |
| N(19)-C(18) | 1.468(3) | N(24)-C(22) | 1.468(4) |
| N(15)-C(13) | 1.381(2) | N(24)-C(23) | 1.500(14) |
| N(15)-C(1) | 1.404(2) | N(24)-C(27) | 1.829(13) |
| N(15)-C(16) | 1.469(2) | C(4)-C(6) | 1.286(11) |
| C(1)-C(2) | 1.399(3) | C(4)-C(22) | 1.460(5) |
| C(1)-C(11) | 1.408(3) | C(4)-C(5) | 1.510(4) |
| N(12)-C(13) | 1.317(2) | C(4)-C(23) | 1.639(14) |
| N(12)-C(11) | 1.395(3) |
Table 3.
Bond angles [°] for 1.
| C(8)-O(7)-C(5) | 113.8(2) | N(12)-C(13)-N(14) | 124.97(19) |
| C(8)-O(7)-C(6) | 116.5(4) | N(12)-C(13)-N(15) | 113.37(18) |
| C(5)-O(7)-C(6) | 39.8(4) | N(14)-C(13)-N(15) | 121.60(18) |
| C(13)-N(14)-H(14A) | 121.2(16) | C(10)-C(9)-C(8) | 120.5(2) |
| C(13)-N(14)-H(14B) | 117.0(17) | O(7)-C(8)-C(2) | 122.1(2) |
| C(13)-N(15)-C(1) | 106.39(16) | O(7)-C(8)-C(9) | 115.08(19) |
| C(13)-N(15)-C(16) | 123.76(17) | C(2)-C(8)-C(9) | 122.8(2) |
| C(1)-N(15)-C(16) | 128.11(16) | C(2)-C(3)-C(4) | 113.01(19) |
| C(2)-C(1)-N(15) | 132.33(19) | C(6)-C(4)-C(22) | 123.2(5) |
| C(2)-C(1)-C(11) | 122.89(19) | C(6)-C(4)-C(5) | 42.8(5) |
| N(15)-C(1)-C(11) | 104.68(17) | C(22)-C(4)-C(5) | 116.2(3) |
| C(13)-N(12)-C(11) | 104.93(16) | C(6)-C(4)-C(3) | 125.8(5) |
| C(10)-C(11)-N(12) | 129.27(19) | C(22)-C(4)-C(3) | 110.8(2) |
| C(10)-C(11)-C(1) | 120.1(2) | C(5)-C(4)-C(3) | 109.9(2) |
| N(12)-C(11)-C(1) | 110.61(17) | C(6)-C(4)-C(23) | 116.2(7) |
| C(1)-C(2)-C(8) | 114.9(2) | C(5)-C(4)-C(23) | 131.1(5) |
| C(1)-C(2)-C(3) | 124.47(18) | C(3)-C(4)-C(23) | 114.0(5) |
| C(8)-C(2)-C(3) | 120.5(2) | O(7)-C(5)-C(4) | 113.6(3) |
| C(9)-C(10)-C(11) | 118.6(2) | C(4)-C(6)-O(7) | 113.0(8) |
Results and Discussion
Benzimidazole Inhibitor Crystal Structure
Depending on the crystallization conditions, compound 1 crystallized either in the monoclinic P21/c space group with one molecule in the asymmetric unit or in the orthorhombic Pna21 space group with two molecules in the asymmetric unit. An ORTEP drawing of the asymmetric unit with atom numbering scheme is shown in Fig. 1. The core bicyclic ring system of compound 1 is essentially planar with the mean deviation from the plane for all non-hydrogen atoms of 0.0124 Å. The pyran ring in 1 is structurally disordered with a 3:1 ratio in favor of the R-enantiomer. As a consequence of this disorder, the dimethylamine alkyl chain on the pyran ring is also disordered. Hydrogen atoms H(14A) and H(14B) were easily located from the electron density map. The dimethylamine-propyl chain folds in such a conformation that it allows for the formation of an intramolecular hydrogen bond of N(19) with H14A [H(14A) •••N(19) = 2.14(3) Å]. H(14B) and N(12) form two additional hydrogen bonds [H(14B) •••N(12) = 2.03(3) Å] with another molecule of 1, forming a dimeric unit. The hydrogen bonds of the aminoimidazole moiety involved in dimer association (Table 4) correspond to the key interactions that occur when 1 binds to the Hoogsteen edge of a guanine residue in the viral RNA target (see next section) [2]. The dimers are positioned within a plane as shown in Fig. 2. Planes stack along the c-axis as shown on Fig. 3. Crystals of compound 1 in the orthorhombic crystal system were obtained by slow evaporation from a methanol solution. Compound 1 crystallized in the Pna21 space group with a racemate of two molecules (R- and S-enantiomer) in the asymmetric unit. While the S-form (1S) is more planar (mean deviation from the plane is 0.0084 Å), the R-form (1R) is less planar (mean deviation from the plane is 0.0193 Å) then the enantiomers of 1 in the P21/c space group. 1R and 1S exhibit the same hydrogen bonding pattern as 1 and form 1R-1S dimeric units. These dimers form a zigzag pattern along the c-axis.
Figure 1.
ORTEP plot of compound 1. Thermal ellipsoids are shown at 50% probability.
Table 4.
Hydrogen bonds for 1 [Å and °].
| D-H…A | d(D-H) | d(H…A) | d(D…A) | <(DHA) |
| N(14)-H(14B)…N(12)#1 | 0.89(3) | 2.03(3) | 2.918(3) | 172(2) |
| N(14)-H(14A)…N(19) | 0.90(3) | 2.14(3) | 3.012(3) | 166(2) |
Symmetry transformations used to generate equivalent atoms:
#1 −x,−y+2,−z+1
Figure 2.
Hydrogen bonds (1) are shown as dashed lines. The hydrogen atoms involved in the hydrogen-bonding network are shown, while the others are omitted for clarity.
Figure 3.
Stacking of dimeric units of 1.
Comparison of the crystal structure of the small molecule free and in complex with the viral RNA target
Packing interactions of compound 1 in the crystal lattice are found analogously in key interactions of the inhibitor with its viral RNA target (Fig. 4). Hydrogen bonds between the amino-imidazole moiety in dimers of both enantiomers of 1 in the small molecule crystal correspond to bonds formed by docking of 1 at the Hoogsteen edge of G110 in the HCV IRES. In the RNA complex, the benzimidazole 1 partially intercalates between two nucleotide bases, G52 and A53. Similarly, the compound 1 is stacking in a co-planar fashion between neighboring molecules in the small molecule crystal lattice. Interestingly, the conformation of both exocyclic dimethylamino alkyl substituents of 1 is similar both in the small molecule crystal and the RNA complex, suggesting that binding to the viral target requires little conformational adaptation of the ligand.
Figure 4.
Comparison of packing interactions of 1 in the small molecule crystal structure (a) with key interactions of the compound with its viral RNA target (b). Compound 1 is shown in yellow stick representation. Neighboring molecules (a) and RNA nucleotides (b) are shown as white sticks. Heteroatoms are colored accordingly. Nucleotides in the RNA target structure are labeled according to the viral genome numbering.
Supplementary Material
Acknowledgments
This work was supported in part by the National Institutes of Health, Grant No. AI72012. Support of the NMR facility by the National Science Foundation is acknowledged (CRIF grant CHE-0741968).
Footnotes
CCDC 869034 (P21/c space group) and 869035 (Pna21 space group) contain the supplementary crystallographic data for this paper. These data can be obtained free of charge by e-mailing data_request@ccdc.cam.ac.uk, or by contacting The Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK, Fax: +44(0)1223–336033.
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