Cooperative Recruitment of Amphotericin B Mediated by a

Article pubs.acs.org/JPCC

Cooperative Recruitment of Amphotericin B Mediated by a Cyclodextrin Dimer Jia He,†,‡ Christophe Chipot,§,∥,▽ Xueguang Shao,†,‡,⊥ and Wensheng Cai*,†,‡ †

Collaborative Innovation Center of Chemical Science and Engineering, Tianjin 300072, China Research Center for Analytical Sciences, College of Chemistry, Nankai University, Tianjin, 300071, China ⊥ State Key Laboratory of Medicinal Chemical Biology, Nankai University, Tianjin, 300071, China § Laboratoire International Associé Centre National de la Recherche Scientifique et University of Illinois at Urbana−Champaign, Université de Lorraine, Unité Mixte de Recherche No. 7565, B.P. 70239, 54506 Vandœuvre-lès-Nancy cedex, France ∥ Theoretical and Computational Biophysics Group, Beckman Institute, University of Illinois at Urbana−Champaign, Urbana, Illinois 61801, United States ▽ Department of Physics, University of Illinois at Urbana−Champaign, 1110 West Green Street, Urbana, Illinois 61801, United States ‡

S Supporting Information *

ABSTRACT: γ-Cyclodextrin (γ-CD) and hydroxypropyl-γ-CD (HP-γ-CD) improve the bioavailability of amphotericin B (AmB) while reducing its toxicity. In a recent study, AmB was found to possess two sites within its prolonged macrolide ring, binding to γ-CD. In the present contribution, cooperative binding of AmB to a γ-CD dimer, a hydroxypropyl-γ-CD (HP-γ-CD) dimer and a hybrid dimer formed by the latter two cyclic oligosaccharides was examined by molecular dynamics simulations and freeenergy calculations in an aqueous solution. The potentials of mean force (PMFs) characterizing the dimerization of the CDs on the macrolide ring of AmB were determined for four different spatial arrangements, namely head-to-head (H−H), headto-tail (H−T), tail-to-head (T−H), and tail-to-tail (T−T). The PMFs allowed the most stable supramolecular organization to be identified along the transition coordinate for every possible orientation of the participating cyclic oligosaccharides. To estimate the absolute binding free energy of each spatial arrangement, alchemical transformations were carried out using free-energy perturbation. T−H corresponds to the most stable orientation for the γ-CD dimer, whereas for the HP-γ-CD and hybrid dimers, the H−T motif is preferred. Our simulations also indicate that, among the three different dimers, the hybrid γ-CD/HP-γ-CD possesses the highest binding affinity toward AmB, in line with experiment. Hydrogen-bonding interactions and spatial matching of the host:guest complex play an important role in the cooperative binding of AmB to CD dimers. The difference in the propensity of the three CD dimers to bind AmB can rationalize the experimental observation that the hybrid γ-CD/HP-γ-CD dimer is a better carrier to enhance the bioavailability of AmB.

■

INTRODUCTION Cyclic polysaccharide-based carriers, such as cyclodextrins (CDs), CD-based copolymers and lipid formulations, have been acknowledged to be ideal candidates for improving the bioavailability of amphotericin B (AmB), a polyene macrolide antifungal drug.1−4 The inclusion properties of AmB and γ-CD, or its derivatives, have been investigated in great detail.5−9 The complexation efficiency and the stability constant of AmB and γ-CD, or hydroxypropyl-γ-CD (HP-γ-CD), in 1:1 and 1:2 stoichiometries, were measured by means of phase-solubility diagrams.3,4 Results show that CD dimer possesses a higher binding affinity toward AmB, relative to the CD monomer. In addition, enhancement of aqueous solubility of AmB by native γ-CDs proved to be more significant than that by HP-γ-CD, irrespective of the stoichiometry.3,4 Surprisingly enough, the hybrid γ-CD/HP-γ-CD dimer possesses a higher binding affinity toward AmB than γ-CD or HP-γ-CD, in a 1:2 © 2014 American Chemical Society

stoichiometry. The free-energy changes characterizing the inclusion of AmB in a CD have been estimated in a previous investigation,10 revealing that AmB can form stable complexes at two well-defined binding sites of its macrolide ring, implying the possibility for concomitant binding of two cyclic polysaccharides. The detail of the underlying mechanism for cooperative binding of AmB to CD dimers remains, however, in large measure fragmentary. To design effective CD-based carriers for AmB, therefore, a complete understanding of the intermolecular interactions at play would be desirable. Theoretical approaches have proven to constitute an appealing route to gain atomic-level insights into recognition and association phenomena.10−17 Many studies have hitherto Received: July 22, 2014 Revised: September 11, 2014 Published: September 19, 2014 24173

dx.doi.org/10.1021/jp507325j | J. Phys. Chem. C 2014, 118, 24173−24180

The Journal of Physical Chemistry C

Article

focused on the relative stability of CD dimers in head-to-head (H−H), head-to-tail (H−T), and tail-to-tail (T−T) orientations. Head refers to the secondary hydroxyl groups, that is, the wide rim of the CD, while tail refers to the primary hydroxyl groups, that is, narrow rim. In the early work of Harada et al., the most favorable conformation of the polyrotaxane (α-CD)nPEG was determined experimentally to be an H−H arrangement.18,19 Similar results were obtained by Mattice and coworkers using molecular dynamics simulations.15 We also examined the dimerization of α-CDs threaded onto a PEG chain theoretically. It was found that the H−H motif is preferred over H−T and T−T, and dimerization is primarily driven by hydrogen bonding between adjacent α-CDs. Moreover, dissociation of β-CD dimers was investigated by means potential of mean force (PMF) calculations, from which it was concluded that formation of the dimer depends on the properties of both the guest and the solvent.12,13 At the experimental level, both H−H and H−T orientations are observed in the crystal packing of CDs.18−21 The main thrust of the present contribution is to explore the cooperative binding of AmB to CD dimers. A number of key aspects, namely the structural features, the underlying thermodynamics, and the determinants for the most stable orientation will be investigated in ample detail. Toward this end, the inclusion of AmB with a γ-CD dimer, an HP-γ-CD dimer, and a hybrid γ-CD/HP-γ-CD dimer in different orientations were dissected at an atomic resolution by MD simulations and free-energy calculations in an aqueous solution. The present work provides a cogent rationalization of the recognition and association of CD dimers with AmB.

of AmB could be included in the cavity of a γ-CD monomer.10 It can, therefore, be conjectured that dimerization of CDs could only occur on the polyene macrolide. Due to the asymmetry of the guest molecule, four dimerization orientations, namely, H− H, H−T, T−H and T−T, were investigated, as illustrated in Scheme 2. In the case of the hybrid γ-CD/HP-γ-CD dimer,

METHODS Molecular Dynamics Simulations. The initial coordinates of AmB, the native γ-CD and the HP-γ-CD were taken from previously published work.10,22 On the experimental front, hydroxypropylated γ-CD is a mixture of structurally related compounds. The average number of substituents for HP-γ-CD used in the experiments is about five. It is possible to construct a structure of HP-γ-CD with similar number of substituents through random substituents at O2 and O6 positions of each glucose unit. However, the previous MD simulations have shown that complexation may vary significantly among the isomers with the same degree of substitution, albeit different substitution sites.23 Therefore, in this study, the O2 and O6 positions of each glucose unit of HP-γ-CD were fully substituted by 2-hydroxypropyl groups (see Scheme 1). A previous investigation reported that only the polyene macrolide

there are by and large two distinct ways to include the guest molecule, that is, (i) the HP-γ-CD first encapsulates AmB from the end of the ring, followed by the inclusion by the native CD, namely, AmB:HP-γ-CD/γ-CD, and, conversely, (ii) the γ-CD encapsulates AmB prior to the HP-γ-CD, namely, AmB:γ-CD/ HP-γ-CD. As a result, 16 initial spatial arrangements were constructed and subsequently immersed in a periodic box of water. Each molecular assembly contains one CD dimer, one AmB, and approximately 3700 water molecules in a simulation cell of initial dimensions equal to 50 × 50 × 50 Å3. The MD simulations were performed with NAMD 2.9,24 using the particle mesh Ewald algorithm25 to handle long-range electrostatic interactions and the r-RESPA multiple time-step integrator26 with time steps of 2 and 4 fs for short- and longrange interactions. A 14 Å cutoff was utilized to truncate the short-range van der Waals and electrostatic interactions. Chemical bonds involving hydrogen atoms were constrained to their equilibrium lengths by means of the Shake/Rattle/ Settle algorithms.27−29 The temperature and the pressure were maintained at 300 K and 1 atm, respectively, using Langevin dynamics and the Langevin piston method.30 Periodic boundary conditions were applied in the three directions of Cartesian space. The carbohydrate solution force field31 and available parameters of the CHARMM27 force field32 were employed to describe inter- and intramolecular interactions. The parameters and charges of AmB were taken from a previous study.10 The analyses of the simulations were performed with the VMD visualization program.33 The hydrogen-bonding criteria used to analyze the number of hydrogen bonds between the two CD monomers are the angle O−H···O > 135° and the distance O···O < 3.5 Å. The contact area between the CD dimers and AmB is given by the

Scheme 2. Schematic Representation of the Initial Structure for the Four Possible Orientations in the Inclusion Complexesa

The CD close to the polar head of AmB is labeled “CD1”, while the other, close to the tail of the macrolide ring, is labeled “CD2”. H or T orientation indicates whether CD faces its partner from its wide or narrow rim.

a

■

Scheme 1. Structure of AmB and CD

24174



Article

following: contact area = [(Sdimer + SAmB) − Scomplex]/2, where Sdimer and SAmB are the solvent-accessible surface areas of the isolated CD dimer and AmB, respectively, and Scomplex is that of the host/guest complex. Free-Energy Calculations. ABF Calculations. The PMFs delineating dimerization of the CDs threaded onto the polyene macrolide of AmB were determined along a unidimensional transition coordinate, that is, the Euclidian distance between the centers of mass (COM) of the glycosidic oxygen atoms of the two the CDs, as shown in Scheme 2. To obtain these profiles, the adaptive biasing force (ABF) algorithm34−36 implemented within the Collective Variables module37 of NAMD24 was utilized. For efficiency, the reaction pathway was broken down into short, 1 Å wide consecutive windows. In each window, at least a 5 ns MD trajectory was generated, corresponding to an aggregate simulation time in excess of 580 ns for the 16 systems. Instantaneous values of the force were stored in 0.1 Å wide bins. The conformational space available to AmB was limited by means of geometrical restraints enforced on the root mean-square deviation (RMSD) relative to the native structure. The geometrical restraint imposed on the conformational space available to AmB by means of the RMSD relative to the native conformation is weak and essentially decoupled from the transition coordinate. Its contribution is, therefore, not accounted for in the PMFs. Since our ABF calculations primarily aim at obtaining the global minimum along the transition coordinate rather than the actual binding free energy, we contend that a simple separation PMF is reasonable here. FEP Calculations. To measure the absolute binding free energy for the different orientations, reversible coupling of AmB to its environment, either in the bound or in the unbound state,38,39 was performed using free-energy perturbation (FEP) (see Scheme 3). The initial structures of the complexes correspond to the minima of the PMFs determined using the ABF algorithm. For both the bound state and the unbound

state, the alchemical transformation was carried out bidirectionally.40 To lower the variance of the estimator, the statistical data accrued in the coupling and decoupling simulations was combined by means of the Bennett acceptance ratio (BAR) algorithm.41,42 In all alchemical transformations, the model pathway was stratified in 26 windows of uneven width. Each stratum consisted of 400000 data-collection steps preceded by 100000 equilibration steps, representing an aggregate simulation time of 884 ns for the 16 chemical systems investigated here. During the above free-energy perturbation calculations, AmB was restrained to its equilibrated conformation by means of an RMSD collective variable. In addition, for the reversible coupling of the bound state, two positional restraints, that is, the Euclidian distances separating the COMs of the polyene macrolide from the two CDs, were enforced. The contribution of these geometrical restraints was determined via independent thermodynamic-integration simulations.43 A stratification strategy of 27 individual points was used. At each point, the gradient of the potential energy with respect to the collective variable was measured. Scaling of the force constant was performed in bidirectional simulations. A total of 400000 data-collection steps were generated at each point after a period of equilibration of 100000 MD steps, representing an aggregate simulation time of 918 ns for the 16 chemical systems. To calculate the standard binding free energy, ΔG0b, the contribution due to the positional restraints ought to be included. Confining AmB in a spherical volume to prevent it from escaping as it is decoupled from its environment is tantamount to a loss of translational entropy, which can be evaluated analytically by ΔΔGt0 = −

1 ln(C 0ΔV ) β −3

(1)

where C = 1/1661 Å , the standard concentration. ΔV is the effective volume sampled by the guest, here approximated as the volume of a sphere, the radius of which is the distance between the center of mass of AmB and that of the CD dimer. The standard binding free energy, thus, writes44,45 0

Scheme 3. Complete Thermodynamic Cycle Used to Estimate the Standard Binding Free Energy of AmB/(CD)2 Complexes, ΔG0ba

ΔG b0 = ΔΔGint + ΔΔGt0 + ΔΔGc

44

(2)

Here, ΔΔGc = ΔGbulk − ΔGsite c c represents the reversible work to maintain AmB in its native conformation. ΔΔGint = ΔGsite int − ΔGbulk int is the net alchemical free-energy change, based on the contributions of the bound and the unbound states.

■

RESULTS AND DISCUSSION Exploring Stable Inclusion Complexes of AmB and CD Dimers. In order to explore the stability and the structural features of the inclusion complexes, the PMFs for the association of the two CDs with the guest were determined and gathered in Figures 1 and 2. All four possible dimerization spatial arrangements were considered. It is worth noting that the PMFs span different ranges of the transition coordinate on account of the distinct intermolecular interactions at play and the distinct supramolecular organizations of the CDs threaded onto AmB − over the chosen ranges of the transition coordinate, the free-energy change is less or equal to 3 kcal/ mol. AmB:(γ-CD)2. As shown in Figure 1A−D, the H−H spatial arrangement features one global minimum around 7.5 Å and two local minima around 12−14 Å. The most stable structure is

“AmB0” denotes an unrestrained AmB molecule, whereas “AmB*” refers to the guest restrained in its native conformation and position in the complex. “nothing*” symbolically stands for the guest decoupled and ΔGbulk refer to the from the solvent or the CD dimer. ΔGbulk c t contributions to the binding free energy of the conformational and positional restraints in the unbound state−marked by the subscript site “bulk”; ΔGsite c and ΔGt correspond to their counterpart in the bound bulk state−marked by the subscript “site”. Here, ΔΔGint = ΔGsite int − ΔGint , represents the net alchemical free-energy change, based on the − contributions of the bound and unbound states. ΔΔGc = ΔGbulk c ΔGcsite is the reversible work to maintain AmB in its native conformation. a

24175



Article

Figure 2. Free-energy profiles for the dimerization of CDs along the model reaction coordinate for AmB:HP-γ-CD/γ-CD in orientation H−H (A), H−T (B), T−H (C), and T−T (D) and AmB:γ-CD/HP-γCD in orientation H−H (E), H−T (F), T−H (G), and T−T (H).

Figure 1. Free-energy profiles for the dimerization of the CDs along the model reaction coordinate for AmB:(γ-CD)2 in its H−H (A), H− T (B), T−H (C), and T−T (D) orientations and for AmB:(HP-γCD)2 in its H−H (E), H−T (F), T−H (G), and T−T (H) orientations.

AmB:(HP-γ-CD)2. In the case of the HP-γ-CD dimer, the PMFs obtained in the four orientations (see Figure 1E−H) all exhibit a single minimum emerging around 14−16 Å. The corresponding most stable structures are depicted in the SI. The substituents located at the tail of the ring occupy the cavity of CD2, while the scaffold is deeply inserted in CD1. Among the four possible spatial arrangements, it is apparent that CD monomers organized in an H−T orientation are the closest to each other. The observation that the distance between the HPγ-CD monomers is generally greater than that between γ-CDs (see Figure 1) stems from intermolecular repulsive interactions of the HP groups. The two HP-γ-CD monomers at the minimum in the H−H spatial arrangement are farther away than in H−T and T−H arrangements, which is at variance with the native CD dimer in the same orientation. This observation can be ascribed to the disruption of the hydrogen-bonding network by the substituents. AmB:γ-CD Hybrids. As depicted in Figure 2, the PMFs exhibit a single free-energy minimum along the reaction pathway. The structures of the complexes around the minima are gathered in the SI. The scaffold and the tail of the macrolide ring embrace the cavity of the two CDs, respectively. The CD monomers in the H−T spatial arrangement are closer than in any of the other orientations, both for AmB:HP-γ-CD/γ-CD and AmB:γ-CD/HP-γ-CD. Preferred Inclusion Complex and Relative Stability of the Different Dimers. The PMFs revealed the stable forms of the inclusion complexes along the chosen transition coordinate for every possible orientation of the cyclic polysaccharides.

depicted in the Supporting Information (SI). The scaffold of the macrolactone moiety is almost completely encapsulated by the H−H CD dimer, while the substituents located at the tail of the ring are somewhat exposed to the solvent. The structures of the two local minima are similar to those of the other orientations (see below) and are, therefore, not shown. In the case of the other spatial arrangements, the PMFs exhibit only one free-energy minimum, located around 12−14 Å, wherein the scaffold of the macrolide ring is deeply included in the cavity of CD1, while the tail of prolonged ring is inserted in CD2 (see SI). The steric hindrances of the aminosugar and the carboxylic acid moiety prevent AmB from penetrating further in the cavity of CD1. As CD2 moves close to CD1, the free energy increases due to repulsive forces between the two cyclic polysaccharides. As CD2 separates from CD1, the free energy also increases because of the dissociation from the tail of AmB. It is apparent that the γ-CD monomers organized in an H−H motif are closer than in other orientations. This geometry is favored by the fact that the secondary hydroxyl groups of the two CDs form a tight intermolecular hydrogen-bonding network and the interactions of the primary hydroxyl groups are weak−a result in agreement with other studies, wherein two CDs in an H−H spatial arrangement attract each other, leading to closer cavities.12,13 Hydrogen bonds are also observed in the alternate orientations. In the 1:2 stoichiometry, the relative position of the two CDs with respect to the binding sites of AmB is very similar to that found for the different poses in the 1:1 stoichiometry inclusion complex.10 24176



Article

Table 1. Binding Free Energies and Their Contributions (in kcal/mol) of AmB and CD Dimers Estimated Using the FEP Method, Based on the Thermodynamic Cycle Sketched in Scheme 3 system AmB:(γ-CD)2

AmB:(HP-γ-CD)2

AmB:HP-γ-CD/γ-CD

AmB:γ-CD/HP-γ-CD

H−H H−T T−H T−T H−H H−T T−H T−T H−H H−T T−H T−T H−H H−T T−H T−T

ΔΔGint

ΔΔGc

ΔG0t

ΔG0b

−7.6 ± 1.0 −10.2 ± 2.0 −12.3 ± 0.8 −7.1 ± 1.7 −9.2 ± 2.3 −11.6 ± 1.0 −8.6 ± 1.3 −7.9 ± 1.1 −9.4 ± 1.1 −13.5 ± 1.2 −8.4 ± 1.5 −8.5 ± 1.1 −11.7 ± 0.8 −11.6 ± 1.3 −9.9 ± 0.9 −8.0 ± 0.9

−0.8 ± 0.4 +0.6 ± 0.8 −0.2 ± 0.2 −0.1 ± 0.2 +0.9 ± 0.2 +0.0 ± 0.8 −0.3 ± 0.6 +1.2 ± 1.5 −0.1 ± 0.3 −0.6 ± 0.5 +0.3 ± 0.4 +2.2 ± 0.7 +1.8 ± 1.2 +0.5 ± 0.3 +1.5 ± 1.1 +0.5 ± 0.9

+3.5 +3.2 +3.1 +3.5 +3.4 +3.4 +3.2 +3.2 +3.2 +3.5 +3.2 +3.4 +3.1 +3.2 +3.4 +3.1

−4.9 ± 1.1 −6.4 ± 2.2 −9.4 ± 0.8 −3.7 ± 1.7 −4.9 ± 2.3 −8.2 ± 1.3 −5.7 ± 1.4 −3.5 ± 1.9 −6.3 ± 1.1 −10.6 ± 1.3 −4.9 ± 1.6 −2.9 ± 1.3 −6.8 ± 1.4 −7.9 ± 1.3 −5.0 ± 1.4 −4.4 ± 1.3

Based on these structures, the underlying host/guest absolute binding free energies were determined in FEP calculations, following the thermodynamic cycle depicted in Scheme 3. The free-energy differences and the standard deviations are gathered in Table 1. AmB:(γ-CD)2. In a previous study of AmB:γ-CD in a 1:1 stoichiometry,10 AmB was shown to form stable complexes with a γ-CD monomer at two distinct binding sites, which we refer here to CD1 and CD2 (see Scheme 2). The energetically most favored inclusion mode corresponds to the T orientation at the site occupied by CD2. Conversely, at the site occupied by CD1, the H and T orientations exhibit an identical binding propensity. If the interaction of the CD monomers were ignored, the most favorable binding mode would be that characterizing the T−T or the H−T spatial arrangement. If, on the contrary, we only consider the interaction of the two CDs, then, the energetically most favored inclusion orientation is expected to be H−H, on account of its tight hydrogen-bonding network. However, as presented in Table 1, the T−H orientation (see Figure 3A) is mirrored in a higher binding affinity, from whence it can be deduced that the stability of the complex is modulated by cooperative interactions of the two CDs and AmB. Further analyses, such as those of hydrogen bonds between the two CDs, of the contact area between the CD dimer and AmB, and of the distance between the two CDs, are presented in Figure 4. In the H−H spatial arrangement, the number of hydrogen bonds between the CD monomers is greater than that in its T− H counterpart (see Figure 4A), but the contact area with the guest molecule of the former is lesser than that of the latter by about 50 Å2 (see Figure 4B), leading to less favorable van der Waals, dispersion interactions. As discussed in our previous work, van der Waals forces are the main contributors to the binding free energy.10 The T−H spatial arrangement, therefore, possesses a higher stability than its H−H counterpart. Hydrogen bonds between the CDs in the H−T and the T− T orientations were found to be less than in the T−H orientation, leading to the observed relatively low stability of the former complexes. AmB:(HP-γ-CD)2. As indicated in Table 1, the energetically preferred inclusion orientation is H−T (see Figure 3B), a result that can be chiefly ascribed to favorable hydrogen-bonding

Figure 3. Snapshots of the inclusion structures of AmB:(γ-CD)2 (A), AmB:(HP-γ-CD)2 (B), AmB:HP-γ-CD/γ-CD (C), and AmB:γ-CD/ HP-γ-CD (D) near the global minima of the PMFs for the most favorable spatial arrangement of the partners. To distinguish between the H and T sides of the CDs, the hydrogen atoms on the O2 position (on the H side) are colored in orange. For clarity, water molecules are omitted.

interactions between the CDs. In addition, CD1 in the H orientation leaves to its partner, CD2, more space to slide on the tail of the macrolide ring. Compared with the H orientation, CD2, in the T orientation, forms with AmB a greater contact area. Last, it is worth noting that among all orientations, the CDs in the H−T spatial arrangement are the closest (see Figure 4C). AmB:γ-CD Hybrids. From Table 1, the energetically preferred inclusion orientation is the H-T spatial arrangement for both the AmB:HP-γ-CD/γ-CD and AmB:γ-CD/HP-γ-CD complex (see Figure 3C, D), which arises primarily from favorable hydrogen-bonding interactions of the two CDs. The distance between the CDs in the H−T orientation is also the shortest, compared to the alternate orientations (see Figure 4C). Furthermore, AmB:HP-γ-CD/γ-CD is preferred over AmB:γ-CD/HP-γ-CD, an observation that can be rationalized by the analysis in Figure 4A,B. In the former host/guest complex, the number of hydrogen bonds and the contact area 24177



Article

complex play an important role in the cooperative binding of AmB to CD dimers. As presented in Table 1, the most favorable binding free energy for AmB:(γ-CD)2 is −9.4 ± 0.8 kcal/mol. As a basis of comparison, at the experimental level, the preferred spatial arrangement corresponds to a binding free energy of −7.2 kcal/ mol.4 For AmB:(HP-γ-CD)2, the most stable spatial arrangement corresponds to a binding free energy of −8.2 ± 1.3 kcal/ mol, somewhat more favorable than the experimental estimate of −7.0 kcal/mol.4 For AmB:γ-CD hybrids, the binding free energy is equal to −10.6 ± 1.3 kcal/mol, again overestimating in absolute value the experimentally determined binding affinity of −7.3 kcal/mol.4 While the theoretical binding free energies reported here systematically overshoot in absolute value the experimental estimates, ranking between the different CD dimers is correct, namely, the hybrid γ-CD/HP-γ-CD < (γCD)2 < (HP-γ-CD)2.

■

CONCLUSIONS The inclusion of AmB in a γ-CD dimer, an HP-γ-CD dimer, and a hybrid γ-CD/HP-γ-CD dimer in different orientations was examined at an atomic resolution by means of MD simulations combined with free-energy calculations in explicit solvent. The PMFs revealed the stable forms of the inclusion complexes along the model reaction pathway for every possible spatial arrangement of the cyclic polysaccharides. Based on these structures, the absolute binding free energies were determined in FEP calculations. For AmB:(γ-CD)2, the energetically preferred inclusion orientation is T−H. For AmB:(HP-γ-CD)2 and AmB:γ-CD hybrids, the energetically preferred inclusion orientation is H−T. Among the three different CD dimers, the hybrid γ-CD/HP-γ-CD dimer and the HP-γ-CD dimer possess, respectively, the highest and lowest binding affinity toward AmB. Further analyses show that cooperative binding of AmB to the CD dimers is primarily driven by spatial matching in the host/guest complex and hydrogen-bonding interactions of the CDs. The difference in the propensity of the three CD dimers to bind AmB can rationalize the experimental observation that the hybrid γ-CD/ HP-γ-CD dimer is a better choice. The results and the analysis presented herein contribute to improve our understanding of the molecular mechanisms that underlie the formation of CDmediated drug carriers.

Figure 4. Number of hydrogen bonds between the CD monomers (A), contact area between the CD dimers and AmB (B), and COM-toCOM distance between the two CD monomers (C) averaged over additional 5 ns MD simulations at thermodynamic equilibrium. n-S, where n = I, ..., IV, refers to the most stable orientation in each complex.

■

ASSOCIATED CONTENT

S Supporting Information *

Snapshots of the inclusion structures of AmB:(γ-CD)2, AmB: (HP-γ-CD)2, AmB:HP-γ-CD/γ-CD, and AmB:γ-CD/HP-γ-CD near the global minima of the PMFs for each favorable orientation. Distances between the two CDs and the tail of the AmB. Complete ref 32. This material is available free of charge via the Internet at http://pubs.acs.org.

are greater than in the latter, both in their most favorable H−T orientation. Relative Stability of the Three Different Dimers. In a closer examination of the free-energy profiles for the different CD dimers, we note that the hybrid γ-CD/HP-γ-CD dimer possesses the highest binding affinity toward AmB. Comparing AmB:HP-γ-CD/γ-CD with AmB:(HP-γ-CD)2, it can be seen that the number of intermolecular hydrogen bonds between the two CDs is greater in the former than that in the latter (see Figure 4A). In addition, the contact area between the hybrid HP-γ-CD/γ-CD and AmB is much greater than that in AmB:(γCD)2 (see Figure 4B). Among the three different dimers in their preferred spatial arrangement, the distance between the CDs in AmB:HP-γ-CD/γ-CD is the shortest (see Figure 4C). In light of these observations, it can be asserted that hydrogenbonding interactions and spatial matching in the host/guest

■

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS This study is supported by National Natural Science Foundation of China (No. 21373117) and Natural Science 24178



Article

(20) Chatziefthimiou, S. D.; Yannakopoulou, K.; Mavridis, I. M. βCyclodextrin Trimers Enclosing an Unusual Organization of Guest: The Inclusion Complex β-Cyclodextrin/4-Pyridinealdazine. CrystEngComm 2007, 9, 976−979. (21) Saenger, W.; Jacob, J.; Gessler, K.; Steiner, T.; Hoffmann, D.; Sanbe, H.; Koizumi, K.; Smith, S. M.; Takaha, T. Structures of the Common Cyclodextrins and Their Larger Analogues Beyond the Doughnut. Chem. Rev. 1998, 98, 1787−1802. (22) Yu, Y. M.; Chipot, C.; Cai, W. S.; Shao, X. G. Molecular Dynamics Study of the Inclusion of Cholesterol into Cyclodextrins. J. Phys. Chem. B 2006, 110, 6372−6378. (23) Yong, C. W.; Washington, C.; Smith, W. Structural Behaviour of 2-Hydroxypropyl-β-Cyclodextrin in Water: Molecular Dynamics Simulation Studies. Pharm. Res. 2008, 25, 1092−1099. (24) Phillips, J. C.; Braun, R.; Wang, W.; Gumbart, J.; Tajkhorshid, E.; Villa, E.; Chipot, C.; Skeel, R. D.; Kalé, L.; Schulten, K. Scalable Molecular Dynamics with NAMD. J. Comput. Chem. 2005, 26, 1781− 1802. (25) Darden, T.; York, D.; Pedersen, L. Particle Mesh Ewald: an N· Log(N) Method for Ewald Sums in Large Systems. J. Chem. Phys. 1993, 98, 10089−10092. (26) Tuckerman, M.; Berne, B. J.; Martyna, G. J. Reversible Multiple Time Scale Molecular Dynamics. J. Chem. Phys. 1992, 97, 1990−2001. (27) Ryckaert, J. P.; Ciccotti, G.; Berendsen, H. J. C. Numerical Integration of the Cartesian Equations of Motion of a System with Constraints: Molecular Dynamics of N-alkanes. J. Comput. Phys. 1977, 23, 327−341. (28) Andersen, H. C. Rattle: A “Velocity” Version of the Shake Algorithm for Molecular Dynamics Calculations. J. Comput. Phys. 1983, 52, 24−34. (29) Miyamoto, S.; Kollman, P. A. SETTLE: An Analytical Version of the SHAKE and RATTLE Algorithms for Rigid Water Models. J. Comput. Chem. 1992, 13, 952−962. (30) Feller, S. E.; Zhang, Y.; Pastor, R. W.; Brooks, B. R. Constant Pressure Molecular Dynamics Simulation: The Langevin Piston Method. J. Chem. Phys. 1995, 103, 4613−4621. (31) Kuttel, M.; Brady, J. W.; Naidoo, K. J. Carbohydrate Solution Simulations: Producing a Force Field with Experimentally Consistent Primary Alcohol Rotational Frequencies and Populations. J. Comput. Chem. 2002, 23, 1236−1243. (32) MacKerell, A. D., Jr.; Bashford, D.; Bellott, M.; Dunbrack, R. L.; Evanseck, J. D.; Field, M. J.; Fischer, S.; Gao, J.; Guo, H.; Ha, S.; et al. All-Atom Empirical Potential for Molecular Modeling and Dynamics Studies of Proteins. J. Phys. Chem. B 1998, 102, 3586−3616. (33) Humphrey, W.; Dalke, A.; Schulten, K. VMD: Visual Molecular Dynamics. J. Mol. Graphics 1996, 14, 33−38. (34) Darve, E.; Pohorille, A. Calculating Free Energies Using Average Force. J. Chem. Phys. 2001, 115, 9169−9183. (35) Hénin, J.; Chipot, C. Overcoming Free-Energy Barriers Using Unconstrained Molecular Dynamics Simulations. J. Chem. Phys. 2004, 121, 2904−2914. (36) Hénin, J.; Fiorin, G.; Chipot, C.; Klein, M. L. Exploring Multidimensional Free Energy Landscapes Using Time-Dependent Biases on Collective Variables. J. Chem. Theory Comput. 2010, 6, 35− 47. (37) Fiorin, G.; Klein, M. L.; Hénin, J. Using Collective Variables to Drive Molecular Dynamics Simulations. Mol. Phys. 2013, 111, 22−23. (38) Gilson, M. K.; Given, J. A.; Bush, B. L.; McCammon, J. A. The Statistical-Thermodynamic Basis for Computation of Binding Affinities: A Critical Review. Biophys. J. 1997, 72, 1047−1069. (39) Hermans, J.; Wang, L. Inclusion of Loss of Translational and Rotational Freedom in Theoretical Estimates of Free Energies of Binding. Application to a Complex of Benzene and Mutant T4 Lysozyme. J. Am. Chem. Soc. 1997, 119, 2707−2714. (40) Pohorille, A.; Jarzynski, C.; Chipot, C. Good Practices in FreeEnergy Calculations. J. Phys. Chem. B 2010, 114, 10235−10253. (41) Bennett, C. H. Efficient Estimation of Free Energy Differences with Monte Carlo Data. J. Chem. Phys. 1976, 22, 245−268.

Foundation of Tianjin, China (No. 13JCYBJC18800). The CINES, Montpellier, France, is gratefully acknowledged for provision of generous amounts of CPU time. The Cai Yuanpei Program is also appreciatively acknowledged for its support of the international collaboration between the research groups of C.C. and W.C.

■

REFERENCES

(1) Gallis, H. A.; Drew, R. H.; Pickard, W. W.; Amphotericin, B. 30 Years of Clinical Experience. Rev. Infect. Dis. 1990, 12, 308−329. (2) Walsh, T. J.; Viviani, M. A.; Arathoon, E.; Chiou, C.; Ghannoum, M.; Groll, A. H.; Odds, F. C. New Targets and Delivery Systems for Antifungal Therapy. Med. Mycol. 2000, 38, 335−347. (3) Jansook, P.; Loftsson, T. CDs as Solubilizers: Effects of Excipients and Competing Drugs. Int. J. Pharm. 2009, 379, 32−40. (4) Jansook, P.; Kurkov, S. V.; Loftsson, T. Cyclodextrins as Solubilizers: Formation of Complex Aggregates. J. Pharm. Sci. 2010, 99, 719−729. (5) Kajtar, M.; Vikmon, M.; Morlin, E.; Szejtli, J. Aggregation of Amphotericin B in the Presence of γ-Cyclodextrin. Biopolymers 1989, 28, 1585−1596. (6) Jansook, P.; Moya-Ortega, M. D.; Loftsson, T. Effect of Selfaggregation of γ-Cyclodextrin on Drug Solubilization. J. Inclusion Phenom. Macrocyclic Chem. 2010, 68, 229−236. (7) Vikmon, M.; Stadler-Szoke, A.; Szejtli, J. Solubilization of Amphotericin B with γ-Cyclodextrin. J. Antibiot. 1985, 38, 1822−1824. (8) Kim, Y. T.; Shin, B. K.; Garripelli, V. K.; Kim, J. K.; Davaa, E.; Jo, S.; Park, J. S. A Thermosensitive Vaginal Gel Formulation with HPγCD for the pH-dependent Release and Solubilization of Amphotericin B. Eur. J. Pharm. Sci. 2010, 41, 399−406. (9) Rajagopalan, N.; Chen, S. C.; Chow, W. S. A Study of the Inclusion Complex of Amphotericin-B with γ-Cyclodextrin. Int. J. Pharm. 1986, 29, 161−186. (10) He, J.; Chipot, C.; Shao, X. G.; Cai, W. S. CyclodextrinMediated Recruitment and Delivery of Amphotericin B. J. Phys. Chem. C 2013, 117, 11750−11756. (11) Liu, P.; Chipot, C.; Shao, X.; Cai, W. How Do α-Cyclodextrins Self-Organize on a Polymer Chain? J. Phys. Chem. C 2012, 116, 17913−17918. (12) Zhang, H.; Tan, T.; Feng, W.; van der Spoel, D. Molecular Recognition in Different Environments: β-Cyclodextrin Dimer Formation in Organic Solvents. J. Phys. Chem. B 2012, 116, 12684− 12693. (13) Zhang, H. Y.; Tan, T. W.; Hetényi, C.; Lv, Y. Q.; van der Spoel, D. Cooperative Binding of Cyclodextrin Dimers to Isoflavone Analogs Elucidated by Free Energy Calculations. J. Phys. Chem. C 2014, 118, 7163−7173. (14) Zheng, X. Y.; Wang, D.; Shuai, Z. G.; Zhang, X. Molecular Dynamics Simulations of the Supramolecular Assembly between an Azobenzene-Containing Surfactant and α-Cyclodextrin: Role of Photoisomerization. J. Phys. Chem. B 2012, 116, 823−832. (15) Pozuelo, J.; Mendicuti, F.; Mattice, W. L. Inclusion Complexes of Chain Molecules with Cycloamyloses. 2. Molecular Dynamics Simulations of Polyrotaxanes Formed by Poly(ethylene glycol) and αCyclodextrins. Macromolecules 1997, 30, 3685−3690. (16) Wallace, S. J.; Kee, T. W.; Huang, D. M. Molecular Basis of Binding and Stability of Curcumin in Diamide-Linked γ-Cyclodextrin Dimers. J. Phys. Chem. B 2013, 117, 12375−12382. (17) Mieda, S.; Ikeda, A.; Shigeri, Y.; Shinoda, W. Thermodynamic Stability of [60]Fullerene and γ-Cyclodextrin Complex in Aqueous Solution: Free Energy Simulation. J. Phys. Chem. C 2014, 118, 12555− 12561. (18) Harada, A.; Li, J.; Kamachi, M. The Molecular Necklace: A Rotaxane Containing Many Threaded α-Cyclodextrins. Nature 1992, 356, 325−327. (19) Harada, A.; Okada, M.; Li, J.; Kamachi, M. Prearation and Characterization of Inclusion Complexes of Poly(propylene glycol) with Cyclodextrins. Macromolecules 1995, 28, 8406−8411. 24179



Article

(42) Liu, P.; Dehez, F.; Cai, W. S.; Chipot, C. A Toolkit for the Analysis of Free-Energy Perturbation Calculations. J. Chem. Theory Comput. 2012, 8, 2606−2616. (43) Gumbart, J. C.; Roux, B.; Chipot, C. Standard Binding Free Energies from Computer Simulations: What Is the Best Strategy? J. Chem. Theory Comput. 2013, 9, 794−802. (44) Deng, Y. Q.; Roux, B. Calculation of Standard Binding Free Energies: Aromatic Molecules in the T4 Lysozyme L99A Mutant. J. Chem. Theory Comput. 2006, 2, 1255−1273. (45) Deng, Y. Q.; Roux, B. Computations of Standard Binding Free Energies with Molecular Dynamics Simulations. J. Phys. Chem. B 2009, 113, 2234−2246.

24180


Cooperative Recruitment of Amphotericin B Mediated by a

Recommend Documents