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Langmuir 2007, 23, 2308-2310
Solubilization of Paclitaxel (Taxol) by Peptoad Self-Assemblies Fredric M. Menger,* Hailing Zhang, Jason de Joannis, and James T. Kindt* Department of Chemistry, Emory UniVersity, 1515 Dickey DriVe, Atlanta, Georgia 30322 ReceiVed NoVember 21, 2006. In Final Form: January 8, 2007 A molecular dynamics simulation was carried out involving a paclitaxel molecule, 987 peptoad molecules, and 35 938 water molecules (conditions shown experimentally to effect paclitaxel solubilization in water). The peptoads are shown to form large clumps, the centers of which are dry and thus favorable to hydrogen bonding between paclitaxel and peptoads. Hydrogen-bonding equilibrium among the peptoads themselves in the developing clumps is achieved in 2 ns. The number and position of hydrogen bonds between the paclitaxel and peptoads fluctuate randomly from two to six within a 2-5 ns time frame. Hydrophobic association between the peptoad chains and the apolar paclitaxel groups does not seem to be an important element of the solubilization. Instead, the hydrophobic chains of the peptoads encasing the paclitaxel extend outward into the dry interior of the peptoad clump where other chains in the clump are located. One hopes that studies such as this will ultimately allow rational predictions when designing new and specific drug solubilizers.
Recently, we synthesized a group of linear amphiphiles called peptoads (see peptoad-G below) and composed of a long hydrocarbon chain coupled to 3 amide groups.1 The water solubility of these compounds showed a remarkable sensitivity to the length of the chain. Thus, an analog of peptoad-G with 4 more carbons in the chain was many orders of magnitude less soluble. Although the highly water soluble peptoad-G has a chain length that is 5 to 11 carbons shorter than the 12 to18 carbons typical of most conventional surfactants, the peptoad nevertheless forms molecular assemblies in water. However, the peptoad assemblies grow continuously above 0.02 M, and in the absence of discrete aggregation numbers and a meaningful critical micelle concentration, as found with most long-chain amphiphiles, we preferred to call the assemblies “clumps” as opposed to “micelles”. One of the more interesting properties of the clumps is their ability to water-solubilize paclitaxel (Taxol), a well-known anticancer drug. This observation led us to investigate the location of paclitaxel within the clumps, the water content at this location, the relative role of hydrogen bonding and hydrophobic interactions, the identity of the paclitaxel/clump contact points, and the rates at which the contact points form and rearrange. As experimental methods cannot easily be applied to these issues, we called upon computer modeling for assistance. The hope was to collect data of a type that would ultimately permit a priori design of solubilizers for any particular drug structure.
GROMACS molecular dynamics simulation2-4 of the clumps (T ) 298 K; P ) 1 bar isotropically applied; 0.002 ps time step; 10 ns total time) utilized 108 peptoad-G molecules and 20 001 (1) Menger, F. M.; Zhang, H. Langmuir 2005, 21, 10428-110438. (2) Berendsen, H. J. C.; van der Spoel, D.; Vandrunen, R. Comput. Phys. Commun. 1995, 91, 43-56. (3) Lindahl, E.; Hess, B.; van der Spoel, D. J. Mol. Model. 2001, 7, 306-317. (4) van der Spoel, D.; Lindahl, E.; Hess, B.; Groenhof, G.; Mark, A. E.; Berendsen, H. J. C. J. Comput. Chem. 2005, 26, 1701-1718.
water molecules in a (8.8 nm)3 box. All peptoad-G atoms were treated explicitly (using a GROMOS force field) except for the aliphatic hydrogens that were handled via methyl and methylene sites. A simple point charge (SPC) model was used to describe water. Despite GROMACS flexibility, speed, and previous success with lipid systems,5,6 a single paclitaxel calculation nonetheless required a dedicated AMD 1.6 GHz single processor system operating for 2 months. Using this model, as opposed to merely subdividing the peptoads into head and tail “units”, allowed us to acquire the necessary degree of atomic resolution. Moreover, in contrast to Monte Carlo simulations often applied to surfactant self-assembly,7 molecular dynamics could provide the time dependency of the self-assembly and solubilization processes. In order to preclude unrealistic intermolecular repulsions at the outset, peptoad-G molecules were initially placed within their box in an arbitrarily stacked array. The system randomized within 0.2 ns, and after 5 ns, the molecules had compacted into an irregular sphere (radius ca. 20 Å) that changed little in shape or energy during the ensuing 5 ns, indicating that the simulations had converged. Figure 1 shows the final status of this 10 ns simulation of an aqueous peptoad-G clump with only the terminal dimethylamides depicted in colored CPK format. The main features of Figure 1 are as follows: (a) The clump contains 97 molecules arranged in a more-or-less uniform density (0.03 ( 0.01 atoms/Å3). Of the original 108 molecules, 11 remained in solution as monomers in dynamic equilbrium with the clump. (b) The terminal amides are located exclusively at the clump surface, near the water, whereas the hydrocarbon chains reside within the clump interior. Figure 2 shows the distribution of carbons-1 and -11 and nitrogen-19 from the center of the clump. (For ease of comparison, the data are presented in analog fashion rather than using multiple histograms.) (c) The clump is dry, apart from a few water molecules located 15-20 Å from the clump center (Figure 2). (d) The number of intermolecular hydrogen bonds (defined as having an acceptor/donor distance of e3.5 Å and an RH/D/A e 60°) reached a maximum of 125 per clump after 3 ns (Figure 3). An analog of peptoad-G, with nine carbons in the chain instead of seven, gave crystals suitable for X-ray analysis (Figure 4).1 (5) Marrink, S. J.; Mark, A. E. J. Am. Chem. Soc. 2003, 125, 15233-15242. (6) de Vries, A. H.; Mark, A. E.; Marrink, S. J. J. Am. Chem. Soc. 2004, 126, 4488-4489. (7) Firetto, A.; Floriano M. A.; Panagiotopoulos, A. Z. Langmuir 2006, 22, 6514-6522.
10.1021/la0633886 CCC: $37.00 © 2007 American Chemical Society Published on Web 02/03/2007
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Figure 1. A 10 ns MD simulation of a peptoad-G clump. The terminal dimethylamides are depicted in CPK format with CH3 in orange, N in blue, and O in red. The remaining segments of the molecules are drawn in line format.
Figure 2. Left Y axis: Distribution of carbons-1 and -11 and nitrogen19 in volume domains of 0-3 Å, 3-4 Å, 4-5 Å, and so forth, where 0 is the center of the spherical clump. Gaussian curves have been imposed upon the data of carbons-1 and -11 and nitrogen-19 intending to show the relative occupancy sites of the groups. Right Y axis: Number of water molecules as a function of distance.
Figure 3. Plot showing the attainment of the hydrogen-bonding equilibrium within a peptoad-G clump. The data give the total number of intermolecular peptoad-G hydrogen bonds in the developing clump vs time.
All-linear chains sit side-by-side in a plane stabilized by NHs OdC hydrogen bonding among the central amide linkages. The
Langmuir, Vol. 23, No. 5, 2007 2309
Figure 4. X-ray-deduced packing of a peptoad (the 9-carbon-chained analog of peptoad-G) in the crystalline state. The peptoad assembles in layers (shown one above another with two molecules in each) hydrogen-bonded to each other (dotted lines, arrows) by means of chain-proximal amide groups. Hydrogen bonding between central amide groups occurs among members of layers (dotted lines, arrows). CdO oxygen is colored red and NH nitrogen is colored blue. The small dots are water molecules from which the crystal was obtained.
Figure 5. Hydrogen bonds in the clump with a 3.5 Å distance and 60° angle cutoff. Peptoad-G is colored in silver, and hydrogen bonds are in red. The hydrogen bonds form a network on the periphery of the clump.
sheets of peptoad molecules are then stacked, chains above chains, in an arrangement stabilized by hydrogen bonding among the set of amide groups proximal to the chains. As seen in Figures 1 and 5, the chains in clumps show none of the solid-sate order. Figure 5 also shows the location of the hydrogen bonds near the surface of the clump. This hydrogen-bonding network explains why peptoad-G self-assembles in water at concentrations where conventional ionic surfactants of the same chain length are monomeric. Peptoad clumps have the benefit of hydrogenbonding attraction in contrast to the electrostatic headgroup repulsion in conventional ionic micelles. When the concentration of peptoad-G was doubled to 216 molecules/box, a rod (l/w ) 2.1) was formed. Sphere-to-rod transitions with increasing concentration are common with conventional surfactants.8 When two 97 molecule clumps were brought into contact, they fused into a rod (similar to that formed from the 216 single molecules) within 1 ns. (8) Holmberg, K.; Jo¨sson, B.; Kronberg, B.; Lindman, B. Surfactants and Polymer in Aqueous Solutions, 2nd ed.; Wiley & Sons: Chichester, England, 2003.
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Figure 6. A plot showing the fluctuating number of hydrogen bonds between peptoad-G and paclitaxel vs time. After 1.5 ns, the number of hydrogen bonds varies from 2 to 6.
In the paclitaxel dissolution studies, 1 molecule of paclitaxel (represented by 66 particles with every CH, CH2, and CH3 group treated as a single entity) was placed at the center of a (12 nm)3 box along with 987 peptoad-G molecules and 35 938 water molecules. This duplicates a concentration used in our experimental solubilization work.5 After 3.2 ns, a space-filling network was formed (accounting for the previously observed viscosity)1 in which paclitaxel was totally encased in peptoad-G. Importantly, the region surrounding the paclitaxel had expelled all water. Thus, water was completely excluded within 6 Å of the paclitaxel, creating an environment that favors paclitaxel hydrogen bonding with the peptoad-G. The hydrogen-bonding file given by GROMACS showed that after 1.5 ns the number of hydrogen bonds between paclitaxel and the peptoad-G fluctuates randomly from 0 to 6. Figure 6 depicts hydrogen-bonding frequency as a function of time. Not only does the number of hydrogen bonds vary with time, but the location of the hydrogen-bonding sites on the paclitaxel fluctuates as well. Figure 7 shows hydrogen-bonding configurations at three arbitrarily chosen times. In summary, peptoad-G functions as a paclitaxel solubilizer by creating clumps with water-free domains where two to six
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Figure 7. Hydrogen bonding between peptoad-G and paclitaxel configuration at three arbitrarily selected times: 4.15, 4.20, and 4.50 ns. Hydrogen bonds, depicted in red, show the H/A distances in angstroms; H-B acceptors or donors from peptoad-G molecules are depicted in blue.
fluxional hydrogen bonds interact with the drug. Direct hydrophobic contact between peptoad-G chains and apolar paclitaxel moieties does not seem to be an important element of the solubilization. Instead, the paclitaxel is sheathed with hydrogenbonded peptoad-G molecules, and it is their hydrocarbon chains that penetrate into the hydrophobic clump interior. No doubt the balance of forces between a drug and its solubilizer varies with the particular system, and much work will be required before reliable predictions will emerge in this important and timely subject.9 Acknowledgment. This work was supported by a National Institutes of Health grant to F.M.M. F.M.M. wrote the paper, H.Z. carried out all the calculations, and J.d.J. and J.T.K. provided expert guidance on the use of the MD method. LA0633886
(9) Stephenson, B. C.; Rangel-Yagui, C. O.; Pessoa, A., Jr.; Tavares, L. C.; Beers, K.; Blankschtein, D. Langmuir 2005, 22, 1514-1525.
