A molecular reactivity template for cannabinoid activity

Journal of Molecular Structure (Theochem), 149 (1987) 331-343 Elsevier Science Publishers B.V., Amsterdam - Printed in The Netherlands

A MOLECULAR REACTIVITV TEMPLATE FOR CANNABINOID ACTIVITY

PATRICIA ‘Department

H. REGGIO’**

and ALEKSANDER

P. MAZUREK’

of Chemistry, Kennesaw College, Marietta, Georgia 30061

(U.S.A.)

aDepartment of Pharmacology, Mount Sinai School of Medicine of the City University of New York, New York 10029 (U.S.A.) (Received 27 June 1986)

ABSTRACT Methods of theoretical chemistry are used to characterize the molecular structure and some reactivity properties of the major psychopharmacologically active component of cannabis, (-)-trans- A 9tetrahydrocannabinol (A 9 -THC). This characterization is part of an exploration of the molecular parameters that could serve to unify considerations of structure-activity relationships in disparate classes of cannabinoids. Our hypothesis is that there are two components of the A9-THC structure that confer upon the molecule reactivity characteristics crucial to activity: the lone pairs of electrons of the phenol oxygen, and the orientation of the carbocyclic ring. The conformation of A9-THC is calculated using molecular mechanics. Since the position of the phenolic OH is central to the working hypothesis, the rotational energy behavior of the phenolic OH is studied. Results from the calculations identify two minimum energy conformations of the OH in A9-THC. Results from ab-initio calculations of the OH rotation on a model fragment of A9-THC agree well with the molecular mechanics results. The molecular electrostatic potential (MEP) of A9 -THC is calculated for the energetically optimal conformations. The results indicate that the two faces of the A’-THC molecule are distinguishable. The MEP of the bottom face of A9-THC (in each of the two minimum energy conformations of the phenolic OH), along with the conformational results for the orientation of the carbocyclic ring relative to the phenol group form a reactivity template to be used in comparisons with properties of active and inactive cannabinoids. INTRODUCTION

Hashish and marihuana are each derived from the Indian hemp, Cannabis Both substances have been used for centuries for their medicinal and psychotomimetic effects. Little is known, however, about the mechanism of action of the major psychopharmacologically active principle, (-)-transA9-tetrahydrocannabinol (A 9-THC) [ 1, 21. Fig. 1 shows the numbering systems commonly employed in the literature for A’-THC. At present there is an increasing amount of information available about various pharmacological and biochemical effects of A9-THC and other cannabinoids, which may or may not be related to their psychopharmacological activity (e.g. see reviews [ 3, 41). Some of the metabolites of A9-THC are themselves psycho-

satiua.

0166-1280/87/$03.50

0 1987 Elsevier Science Publishers B.V.

332

A’-THC

= A’-THC

Fig. 1. An illustration of the two numbering systems which are commonly employed for the cannabinoids.

pharmacologically active. To what extent they contribute to the behavioral actions of A’-THC has not been established [5]. It has been shown, however, that A’-THC itself can exert behavioral effects without conversion to one of its active metabolites [ 61. Several hypotheses concerning the molecular mechanism of action of A’THC exist. One states that A’-THC might act on one or several of the known neurotransmitter/receptor systems [7]. Another major view is that the cannabinoids interact with their own specific receptor in the central nervous system [8, 91. Still another view is that the high lipophilicity of the cannabinoids allows them to produce their effects by perturbation of phosphohpid ordering in membranes [lo]. Many studies of structure- activity relationships (SAR’s) in the cannabinoid series have been published (e.g. see review [ 111 and more recently [ 12-141). An immediate observation from a review of all of these studies is that some cannabinoids can be structurally dissimilar as in A’-THC and Abbott 40656 [15] and yet have similar activities, whereas others, which show only slight structural differences as in lo- O-OH- A8-THC and lo- O-OH- A’-THC [ 161 can exhibit dramatic activity differences (Fig. 2). This observation emphasizes that a common basis for the activity of the compounds must be sought in their chemical properties and reactivity characteristics rather than merely in their three dimensional structures. Traditionally, cannabinoid SAR’s have focused almost exclusively on the independent contribution of certain structural groups (“functional groups”) of the molecules and have been compiled into extensive “lists of requirements” [ 171. Various investigators have alluded to the possible importance of certain regions or conformations of the molecules. These regions include: a methyl group in the plane of the aromatic ring, a free phenol and a free C-4(C-5’) aromatic position [ 161, the presence of the phenolic hydroxyl group [18], the side chain [ 191, non-planarity [20], and planarity of the molecule [21]. Stereoselectivity in the activity of the tetrahydrocannabinols has been reported [22]. This type of approach in SAR studies often carries

333

Abbott

40656

(-)-A’-THC Active

Active (A)

IO-a-OH-n8-THC

pseudoequotonol

IO-P-OH-A’

THC pseudooxlal Less active

Active (El

Fig. 2. Cannabinoids (a) which are structurally dissimilar yet exhibit similar activities, and (b) which are structurally similar yet exhibit dramatically different activities.

tacit assumptions: that functional groups must react directly with specific sites in the receptor, that modification of one group does not affect the reactivity of another, and, consequently, that geometric and stereometric factors such as distance and spatial relationships between functional groups are all important. This focus on isolated aspects of the cannabinoids ignores the fact that the molecular properties that are directly responsible for the molecular interactions (that lead to the pharmacological effect) are encoded in the entire molecular structure. To date, no attempt to characterize the entire molecule and to correlate this characterization with its activity has been attempted in the cannabinoid field. Previous theoretical studies on cannabinoids have not focused on the calculation of molecular reactivity characteristics. Instead, these studies have concentrated almost exclusively on the calculation of conformation in an attempt to predict the most probable geometry (lowest energy conformer) and its relation to activity. The structures and energies of A9-THC and three other cannabinoids obtained by Westheimer calculations and extended Huckel molecular orbital calculations have been reported [23]. Theoretical studies of cannabinoids with analgetic and anticonvulsant activity have also been performed [24, 251. The work presented here reaches beyond the considerations which have prevailed in theoretical studies of the cannabinoids and aims to reveal elements of molecular reactivity that are related to activity.

334

In order to explore the molecular determinants for cannabinoid activity, a template molecule must be chosen. Such a molecule should have demonstrated cannabinoid activity, so that its molecular reactivity characteristics can be used as criteria for cannabinoid activity in future comparisons. Since A9-THC has been reported to be the major psychoactive component of cannabis [l, 21 and since studies indicate that it can exert behavioral effects without metabolic activation [6], we have chosen A9-THC as the template molecule. We report here the formulation and first test of a hypothesis that emerged from our review of the SAR literature. According to this hypothesis there are two components of the A9-THC structure that confer upon the molecule ‘reactivity characteristics crucial to activity. These components are: the lone pairs of electrons of the phenol oxygen (these lone pairs generate reactivity properties dependent on the orientation of the OH bond relative to the carbocyclic ring), and the orientation of the carbocyclic ring (this ring and its orientation generates hydrophobic properties). The spatial arrangements of the reactivity characteristics generated by these structural elements of A9-THC are the components of a reactivity template to which those of other cannabinoids will subsequently be compared. METHODS

Conformational study of A9-THC The first step in the characterization was the conformational analysis of A9-THC. The force field or molecular mechanics method as encoded in the program MM2 [26] was used in this analysis. The force field available in MM2 has been parameterized for oxygen-containing molecules and has proven satisfactory for the calculation of structures and energies of oxygencontaining compounds [27]. The X-ray structure of A9-THC Acid B [28] was used to obtain an input geometry, i.e. bond lengths, bond angles, and dihedral angles for the fused ring skeleton of A9-THC. Necessary atoms were added to this skeleton at standard bond lengths and bond angles [29] to produce an input geometry for A9-THC in the MM2 calculation. Since the position of the phenol was very important to our working hypothesis, two studies of the molecular energy as a function of rotation about the C1-0 axis were conducted. In the first study, the dihedral driver option in the MM2 program [26] was used. Rotations in 36 steps were made about the C1-0 axis. In a second study, an SCF calculation was performed for a model fragment of A9-THC (see Fig. 3). The GAUSSIAN 80 system of programs and the STO-3G basis set were employed here [30]. Initially all atoms in the fragment were frozen in their optimized positions. (These positions were determined by MM2 calculations for the full molecule.) Rotations of the phenolic hydrogen in 18 steps were made about the C1-0 axis.

335

Fig. 3. Model fragment for A ‘-THC.

Molecular electrostatic potential A very direct indication of the nature and extent of electrostatic drugreceptor interactions was obtained by mapping the potential generated by A’-THC. The procedure used to calculate these maps was as follows: (1) molecular orbital calculations were performed using the GAUSSIAN 80 package of programs [30] and the LP-3G basis set [31] for each of the minimum energy conformations identified in the studies above. (2) Each conformation together with its corresponding LP-3G wave function was then input to POLYPOT, the electrostatic potential mapping program [32] to obtain an MEP map. The MEP’s at 1.5 A above and below the plane of the aromatic ring were calculated for each of the minimum energy conformations of the molecule. Because A’-THC was too large a molecule to be accommodated by GAUSSIAN 80, an MM2 optimized version of A’-THC with a propyl, instead of a pentyl side chain was used for these calculations. Such a modification was acceptable since the focus of our study was on the fused ring structure and the phenol group and not on the side chain. The LP-3G basis set was used in the Coreless HartreeFock Effective Potential (CHFEP) scheme developed by Topiol [33, 341. It has been shown that the MEP is well represented by wave functions obtained with an LP-3G basis set [35, 361. RESULTS AND DISCUSSION

Conforma tional analysis Figure 4 shows the minimum energy conformer of (-)-trans-A’-THC as obtained from the MM2 calculation. The results of this calculation compare very favorably with that of the MM1 calculation performed by Archer et al. [23]. Previous studies have predicted that the cyclohexene ring (ring A) of A 9-THC should .exist predominantly in a half-chair conformation [ 37-391. The results obtained here with MM2 are consistent with this expectation. As a result of the interaction between the C6 methyl groups and Hha, ring B assumes a conformation such that the axial C6 methyl group is on the same

336

Fig. 4. Conformation

of A9-THC (with propyl side chain) as determined by MM2.

side of the molecule as HIOa and is much closer to H,,, than is the other methyl group. The substituents on C6 and C,, are staggered with respect to one another (optimized C10a-C6aC6-05 dihedral angle is 63”). NMR results for A9-THC corroborate this conformation of ring B [23] . The Cr hydroxyl group is subject to steric interaction with the Cl0 proton. This interaction causes the 0 -C,-C 1o,, bond angle to open slightly (121”), and the hydroxyl oxygen to bend slightly out of the plane of the benzene ring. The hydroxyl proton optimizes at a position away from ring A (see study of hydroxyl proton position below). NMR studies of A9-THC confirm this conformational preference of the phenolic OH [ 23 J . Optimized bond lengths are within 0.01 a of the following values: C,z-C 2 1.40 A, C,2-C,3 1.51 A, C,3-CfP3 1.53 A, C,z--0 1.36 A, C,3-0 $2 A, (&Q--H 1.10 A, &3--H 1.11 A, and O-H .94 A. One notable difference between the MM1 results reported by Archer et al. [23] and the MM2 results reported here is the C2-C1-O-H dihedral angle, 7. Our calculations. indicate a 7 of --lo, whereas Archer et al. reported a range of 30-50” for the cannabinoids studied. The X-ray structure of A9-THC Acid B shows r = -1” [28]. It is clear from Fig. 4 that A9-THC is a non-planar molecule, a fact that is often ignored. Fig. 5 shows a perspective of the molecule viewed in the direction parallel to the vector from CZ to Glob. Since this conformation will be used as a template for comparisons, it is important to note the relationship between ring A (the carbocyclic ring) as shown in Fig. 5 and ring C (the aromatic ring) with its OH substituent. From this perspective, the conformation of ring A causes the top part of the ring to move to the left, thus permitting no protrusion of ring A into the bottom face of the molecule. It is part of our working hypothesis that this orientation is necessary for cannabinoid activity. The results of the MM2 study of the molecular energy as a function of rotation about the C 1-O axis are summarized in Fig. 6. There are local minima at 7 = -1” and 155” separated by a 2.6 kcal barrier. The SCF results for the model fragment of A9-THC agree very well with those above (see Fig. 7). The

337

Fig. 5. Conformation of A9 -THC as determined by MM2: here the perspective of ring A is viewed in the direction parallel to the vector from C!, to Glob.

3.5

0

180 00 220 240 250 280 300 320 340= 20 40 60 80 100 1M 140 160kg?

TAU

Fig. 6. Rotational energy behavior of the phenolic OH group of a9-THC as determined by an MM2 calculation.

SCF method pinpointed two minimum energy conformers 7 = -1” and 7 = 154” separated by a 4.3 kcal barrier. The results of these studies of molecular energy as a function of rotation

Fig. 7. Rotational energy behavior of the phenolic OH group of a model fragment of ‘-THC as determined by an SCF calculation.

A

about the C1-O axis indicate that two minimum energy conformations of the phenolic OH exist. In Conformation I, the phenolic OH is only slightly bent out of the plane of the aromatic ring with the hydrogen pointing away from Ring A (7 = -1”). In Conformation II, the phenolic hydrogen is below the plane of the aromatic ring and is pointing towards ring A (T = 155”). The conformation of ring A itself remains unchanged. Since the conformational preference of the phenolic hydroxyl is important to our working hypothesis (which concerns the orientation of the phenolic OH relative to ring A), and since both of these minima may be important, both of these conformations were considered in our attempt to relate structure to activity. Molecular electrostatic potential The calculated electrostatic potential pattern generated by A’-THC in the two minimal energy conformers are shown in Figs. 8-11. These patterns are composed of lines of equipotential energy, in K&/mole, that would be felt by a test positive charge placed at any specified point on the map. Figs. 8-11 illustrate the regions of negative potential in planes parallel to the benzene ring at distances of 1.5 A above and below the plane. Because A’-THC is a non-planar molecule, the MEP’s will differ depending upon whether the molecule is viewed from above (“top face”, Figs. 8, 10) or below (“bottom face”, Figs. 9, 11). Comparison of MEP’s Top face The MEP’s of the top face of A’-THC in both of its minimum energy conformations (Conformation I, Fig. 8 and Conformation II, Fig. 10) are very similar. Both exhibit minima located over the aromatic ring and over the

339 cl ::

a

2 D

0 a

L.-Y 0

0 d 0 + D 0’ -15.0

O A + x 0 I

I

-10.0

-5.0

I

I

I

1

0.0

5.0

10.0

15.0

-44.0 -40.0 -35.0 -30.0 -20.0 -10.00

Fig. 8. Molecular electrostatic potential of the top face of A~-THC (Conformation I). Contours represent the electrostatic potential in a plane 1.5 A above the aromatic ring in the molecule. Values are in kcal mol-I.

Fig. 9. Molecular electrostatic potential of the bottom face of A~-THC (Conformation I). Contours represent the electrostatic potential in a plane 1.5 A below the aromatic ring in the molecule. Values are in kcal mol-‘.

oxygen of ring B. The region of negative potential near the phenol oxygen in Conformation II is slightly larger than that of Conformation I (i.e., the potential extends further from the molecule in this region). Bottom face The MEP’s for the bottom face of A9-THC in Conformation I (Fig. 9) and Conformation II (Fig. 11) exhibit a notable difference. Both MEP’s possess

340

0

u; I3 0 * + x O

0 d 0 + 0 s! ’

-15.0

I

-10.0

1 -5.0

I 0.0

I 5.0

Fig. 10. Molecular electrostatic See Fig. 8 for details.

I 10.0

-4.5.0 -40.0 -35.0 -30.0 -20.0 -10.00

I 15.0

potential of the top face of a9-THC (Conformation

II).

-41.0 O -40.0 q

* + x O

Fig. 11. Molecular electrostatic tion II). See Fig. 9 for details.

potential

-35.0 -30.0 -20.0

-10.00

of the bottom

face of A9-THC (Conforma-

minima associated with the aromatic ring and a less pronounced minimum associated with the ring B oxygen. Each MEP exhibits a minimum associated with the phenol oxygen. The location of this minimum however, and the regions into which the negative potential extends are distinguishable. In Conformation I the phenolic hydrogen is oriented away from ring A. Consequently, the MEP of Conformation I in Fig. 9 is characteristic of the lone pairs of electrons of the phenol oxygen pointing towards ring A. In Conformation II the phenolic hydrogen is oriented towards ring A. Consequently, the MEP of Conformation II in Fig. 11 is characteristic of the lone

341

pairs of electrons pointing away from ring A. Thus the difference in the position of the phenolic hydrogen (and consequently of the lone pairs) causes a distinguishable difference in the MEP’s of the two conformers. CONCLUSIONS

In the light of the present findings, our working hypothesis can be refined somewhat to state that there are two components of the A’-THC structure that confer upon the molecule reactivity characteristics crucial to activity. These components are: the orientation of the carbocyclic ring (ring A) as illustrated in Fig. 5 such that this ring moves out of the bottom face of the molecule (this conformation is likely to allow for nondirectional hydrophobic interactions), and the lone pairs of electrons of the phenol oxygen (these generate directional reactivity properties dependent on the orientation of the OH bond relative to the carbocyclic ring). A consideration of the MEP patterns of the top and the bottom faces of A’-THC (in either of its pertinent OH positions) reveals that the faces should be distinguishable on a molecular level. Our results indicate that the MEP of the bottom face of the A’-THC molecule is sensitive to the position of the phenolic OH group of ring C. We therefore propose that it is the MEP of the bottom face of the molecule that must be recognized at the site of action. At this stage of the investigation it is impossible to identify which conformation of the phenolic OH of A’-THC (I or II) is more relevant at the site of action. Such a conclusion will depend on subsequent comparisons of the A’THC reactivity template established here, with the properties of active and inactive cannabinoids. Hence the MEP’s shown in Figs. 9 and 11 along with the conformation of ring A as illustrated in Fig. 5 form our preliminary set of molecular reactivity characteristics for cannabinoid activity. ACKNOWLEDGEMENTS

The authors thank Dr. Hare1 Weinstein for many helpful discussions. This work was supported in part by grants from the National Institute on Drug Abuse DA-03934 and DA-01875. A generous grant of computer time from the University Computing Center of the City University of New York is gratefully acknowledged. Data analysis and curve fitting were done on the PROPHET computer system, a national computational resource sponsored by the NIH through the Division of Research Resources. One of us (P. H. R.) also wishes to acknowledge support from the Kennesaw College Faculty Development Grant Program. REFERENCES 1 Y. Gaoni and R. Mechoulam, 2 Y. Gaoni and R. Mechoulam,

J. Am. Chem. Sot., 86 (1964) J. Am. Chem. Sot., 93 (1971)

1646. 217.

342 3 H. G. Pars, R. K. Razdan and J. F. Howes, in N. J. Harper and A. B. Simmonds (Eds.), Advances in Drug Research, Academic Press, London, 1977, p. 97. 4 S. Burstein and S. Hunter, Rev. Pure Appl. Pharm. Sci., 2 (1981) 155. 5 G. R. Handrick, R. P. Duffley, G. Lambert, J. G. Murphy, H. C. Dalzell, J. F. Howes, R. K. Razdan, B. R. Martin, L. S. Harris and W. C. Dewey, J. Med. Chem., 25 (1982) 1447. 6 J. M. Carney, R. C. Balster, B. R. Martin and L. S. Harris, J. Pharmacol. Exp. Ther., 210 (1979) 399. 7 M. K. Poddar and W. L. Dewey, J. Pharmacol. Exp. Ther., 214 (1980) 63. 8 M. Binder and 1. Franke, in F. Hucho (Ed.), Neuroreceptors, Walter de Gruyter, Berlin, 1982, p. 151. 9M. Binder, F. J. Witteler, B. Schmidt and I. Franke, in S. Agurell, W. L. Dewey and R. E. Willette (Eds.), The Cannabinoids: Chemical, Pharmacologic, and Therapeutic Aspects, Academic Press, New York, 1984, p. 709. 10 C. J. Hillard, A. R. Harris and A. S. Bloom, J. Pharmacol. Exp. Ther., 232 (1985) 579. 11 R. D. Sofia, in L. L. Iverson, S. D. Iverson and S. H. Snyder (Eds.), Handbook of Psychopharmacology, Vol. 12, Drugs of Abuse, Plenum Press, New York, 1978, p. 319. 12 B. R. Martin, M. J. Kallman, G. F. Kaempf, L. S. Harris and W. L. Dewey, Pharmacol. Biochem. Behav., 21 (1984) 61. 13 B. R. Martin, L. S. Harris and W. L. Dewey, in S. Agurell, W. L. Dewey, and R. E. Willette (Eds.), The Cannabinoids: Chemical, Pharmacologic, and Therapeutic Aspects, Academic Press, New York, 1984, p. 523. 14 H. Edery, G. Porath, R. Mechoulam, N. Lander, M. Srebnik and N. Lewis, J. Med. Chem., 27 (1984) 1370. 15 S. Rosell, U. Bjorkroth, S. Agurell, K. Leander, A. Ohlsson, B. Martin, and R. Mechoulam, in G. G. Nahas and W. D. M. Paton (Eds.), Marihuana Biological Effects: Analysis, Metabolism, Cellular Responses, Reproduction, and Brain, Pergamon Press, Oxford, 1979, p. 63. 16 H. Edery, Y. Grunfeld, Z. Ben-Zvi and R. Mechoulam, Ann. N.Y. Acad. Sci., 191 (1971) 40. 17 R.‘K. Razdan, in S. Agurell, W. L. Dewey and R. E. Willette (Eds.), The Cannabinoids: Chemical, Pharmacologic, and Therapeutic Aspects, Academic Press, New York, 1984, p. 63. 18 D. B. Uliss, H. D. Calzell, G. R. Handrick, J. F. Howes and R. K. Razdan, J. Med. Chem., 18 (1975) 213. 19 P. F. Osgood, J. F. Howes, R. K. Razdan and H. G. Pars, J. Med. Chem., 2 (1978) 809. 20 M. Binder, H. Edery and G. Porath, in G. G. Nahas and W. D. M. Paton (Eds.), Marihuana Biological Effects: Analysis, Metabolism, Cellular Responses, Reproduction, and Brain, Pergamon Press, Oxford, 1979, p. 71. 21 S. Burstein, 9. A. Hunter and K. Ozman, Mol. Pharm., 23 (1983) 121. 22 W. Dewey, B. Martin and E. May, in D. E. Smith (Ed.), CRC Handbook of Stereoisomers: Drugs in Psychopharmacology, CRC Press, Boca Raton, FL, 1984, p. 317. 23 R. A. Archer, D. B. Boyd, P. V. Demarco, I. J. Tyminski and N. L. Allinger, J. Am. Chem. Sot., 92 (1970) 5200. 24 M. R. Johnson, L. S. Melvin and G. M. Milne, Life Sci., 31 (1982) 1703. 25 I. Tamir, R. Mechoulam and A. Meyer, J. Med. Chem., 23 (1980) 223. 26 N. L. Allinger and Y. H. Yuh, “MM2”, Quantum Chemistry Program Exchange, 13 (1980) 395. 27 N. L. Allinger, S.-H. Chang, D. H. Glaser and H. Honig, Isr. J. Chem., 20 (1980) 51. 28 E. Rosenquist and T. Ottersen, Acta Chem. Stand., 1329 (1975) 379. 29 A. D. Mitchell and L. C. Cross (Eds.), Tables of Interatomic Distances and Configurations in Molecules and Ions, Special Publication No. 11, The Chemical Society, London, 1958; Special Publication No. 18, 1965.

343 30 E. J. Fluder, H. B. Schlegel and S. W. Topiol, GAUSSIAN 80 (IBM version), unpublished, 1980. 31 S. Topiol and R. Osman, J. Chem. Phys., 73 (1980) 5191. 32 H. Weinstein, S. Srebrenik and R. Pauncz, POLYPOT, unpublished (1973). 33 S. Topiol, J. W. Moskowitz and C. F. Melius, J. Chem. Phys., 68 (1978) 2364. 34 S. Topiol, J. W. Moskowitz and C. F. Melius, J. Chem. Phys., 70 (1979) 3008. 35 H. Weinstein, R. Osman, S. Topiol and J. P. Green, Ann. N.Y. Acad. Sci., 367 (1981) 434. 36 S. Topiol, R. Osman and H. Weinstein, Ann. N.Y. Acad. Sci., 367 (1981) 17. 37 N. L. Allinger, J. A. Hirsch, M. A. Miller and I. J. Tyminski, J. Am. Chem. Sot., 90 (1968) 5773. 38 F. R. Jensen and C. H. Brushweller, J. Am. Chem. Sot., 91 (1969) 5774. 39 J. F. Chiang and S. H. Bauer, J. Am. Chem. Sot., 91 (1969) 1898.