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Neuromuscular blocking agents: Structure and activity
Chem.-BioL Interactions, 6 (1973) 351-365
351
© Elsevier Scientific Publishing Company, Amsterdam--Printed in The Netherlands
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY*
PETER PAULING ANDTREVOR J. PETCHER** William Ramsay, Ralph Forster and Christopher Ingold Laboratories, University College London Gower Street, London WC 1 (Great Britain)
(Received February 7th, 1973)
SUMMARY Correlation of crystal structure analyses and model building of rigid molecules indicate the structural requirements for curariform activity and the structural differences between depolarizing and non-depolarizing neuromuscular blocking agents.
INTRODUCTION We discuss here the correlation between three-dimensional structure and physiological activity of potent neuromuscular blocking agents 1-4. The structures considered have been determined by X-ray diffraction analysis 5-7 or derived from known crystal structures by use of atomic models and the application of accepted stereochemical principles. Some structural rules for potent neuromuscular blocking activity are presented and some proposals are made for the synthesis of new neuromuscular blocking agents. There is a significant structural difference between depolarizing and non-depolarizing blocking agents which provides information about the mechanism of the cholinergic neuromuscular receptors. The structural correlation has been based on the crystal structure of O,O',N-trimethyl-(÷)-tubocurarine di-iodide 8 and we describe the structural features of other potent neuromuscular blocking agents in terms of the pharmacodynamic groups of this molecule. Tubocurarine
The conformation of T M T C , in crystals of the di-iodide a is shown in Fig. 1. The molecule is sufficiently rigid that it is likely that no change in conformation can result in going from the crystal to solution, or upon interaction with the hypothetical receptor substance. The forces involved in maintaining the molecular rigidity are for * This work was supported by the Medical Research Council of Great Britain. ** Present address: Research Laboratories for Pharmaceutical Chemistry, Sandoz Ltd., Basle (Switzerland). Abbreviations: CPK, Corey, Pauling, Koltun; EPMR, equipotent molar ratio; TMTC, O,O',Ntrimethyl-(+)-tubocurarine.
352
P. PAULING, T. J. PETCHER
C H 3 0 ~
O
CH 2
CH2
OCH30
/
\
v
v
-OCH3
Fig. 1. The conformation of O,O',N-trimethyl-(+)-tubocurarine as observed in crystals o f the diiodide s. The molecule is projected onto the plane o f N1, N2, 02, 04.
the most part repulsive interactions between hydrogen atoms. The N + - N + distances in the two independent molecules of T M T C in the unit cell are observed to be 1060 and 1075 pm*. The conformations of the two independent molecules in the unit cell are essentially identical. The most striking feature of the gross conformation of the molecule is that there is a fold about the line in which lie the six oxygen atoms which endows the two major faces of the molecule with entirely different properties. One face, the upper in Fig. 1, is largely hydrophilic, is convex, and has all the lone pairs of electrons on the oxygen atoms directed outwards from the ridge which they occupy. The reverse face, in contrast, is concave and almost entirely hydrophobic. These features immediately suggest that the receptor area responsible for binding curare-like molecules might have, in addition to two anionic sites separated by approximately 1080 pm to bind ammonium groups, a hydrophilic cleft capable of dipole-dipole interaction with the oxygen atoms and a hydrophobic ridge opposite this cleft capable of interacting with the concave hydrophobic side of the molecule. T M T C was formerly known as O,O'-dimethyltubocurarine but it has recently been shown 9 that the previously accepted formulation of (+)-tubocurarine is incorrect. The correct formula is (I). (+)-Tubocurarine contains one quaternary nitrogen atom and one tertiary nitrogen atom. T M T C is approximately 10 times more potent than tubocurarine and this increase, which was previously ascribed entirely to O-methylation 1° is presumably partially due to quaternisation of the tertiary nitrogen atom, which in tubocurarine is largely protonated at physiological pH. * 100 picometers (pm, 10. * 2 m) equals 1 Angstrom.
353
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
CH30~
+
0
CH2
Form. I - Tubocurarine. Pancuronium The crystal structure o f this steroid derivative (3~,17fi-diacetoxy-2fl,16fl-di(Nm e t h y l p i p e r i d i n o ) - 5 ~ - a n d r o s t a n e ) has recently been d e t e r m i n e d 11, a n d the absolute configuration is k n o w n b y relation to that o f the p a r e n t steroid. The c o n f o r m a t i o n is shown in Fig. 2. This molecule is a p o t e n t n e u r o m u s c u l a r b l o c k i n g agent o f clinical i m p o r t a n c e since it a p p e a r s to be largely free o f the undesirable side effects such as
b
Fig. 2. The conformation of pancuronium in crystals of the bromide 11, projected onto the backbone chain between the nitrogen atoms. The crystals also contain solvent molecules of water and methylene chloride.
354
P. PAULING, T. J. PETCHER
tachycardia 1,2 associated with conventional blocking agents such as gallamine and is only one-tenth as potent as hexamethonium at ganglia 12. In man, the EPMR is approximately 0.2 with respect to (q-)-tubocurarine 12. The molecule is entirely rigid because repulsive interactions between the piperidine rings and the steroid nucleus effectively preclude any boatchair interconversion in the androstane fragment. The N + - N + separation is 1108 pm, the steroid and piperidine rings form an entirely hydrophobic mass down the line joining the two nitrogens but the oxygen atoms of the two acetoxy groups are not similarly disposed to those of TMTC. One interesting feature of pancuronium is the eight-carbon chain held rigidly in an almost antiplanar conformation joining the two charged nitrogen atoms. We shall return to this point in the discussion. Calabash curarines
All potent neuromuscular blocking agents obtained from the alkaloids of calabash curare contain two of the same fused five-ring units (II) joined together in different ways 13. The conformation but not the absolute configuration of the basic building unit has been established in the crystal structure of caracurine lI (Fig. 3) (ref. 5) and one may build space-filling molecular models of these compounds with some confidence, since there are only restricted ways of joining together the two basic units. The absolute configuration of the subunit has been established 14 by relation to that of (--)-strychnine and corresponds to the published atomic co-ordinates of caracurine II. This is the absolute configuration that we have used. CH 3
Form. II - Subuni t o f c a l a b a s h curarine.
CH 3
HOCH2 J 3,CH
CH II II
2'
CH I
J 4CH2 OH
J pe
CH~
Form. l l I - Toxiferin I.
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
355
i?
CH3 Fig. 3. T h e c o n f o r m a t i o n o f caracurine II in crystals o f the dimethiodide 5, d e m o n s t r a t i n g the configuration o f the basic s u b u n i t o f calabash curarines, projected down the two-fold axis o f molecular symmetry.
CH3 N
CH3
I
H .....
CH
..... H
J
CH3
CH 3
Fig. 4. T h e c o n f o r m a t i o n o f toxiferin II in crystals o f the di-iodide 7, projected down the axis o f twofold molecular s y m m e t r y .
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P. PAULING, T. J. PETCHER
The conformation of the potent neuromuscular blocking agent toxiferin II (C-calebassine) has been determined in crystals of the di-iodide 7, and is shown in Fig. 4. The N + - N ÷ distance is approximately 1060 p m and the molecule has a convex hydrophilic face, a concave hydrophobic face, and folds about a line through the two oxygen atoms which are separated by approximately 480 pm. The conformation of the five-ring subunit is identical to that observed in the structure of caracurine II (ref. 5).
F o r m . IV - D i a z a o c t a d i e n e ring in toxiferin I.
One of the most potent molecules in this group is toxiferin I (III) (ref. 15). The two subunits are joined by - N - C = bonds to form a 1,5-diazaocta-3,7-diene ring (IV) at the centre of the molecule. There is only one way of joining the two subunits so that the resulting molecule correlates with TMTC. The two uncharged nitrogen atoms are trigonal and the diazaoctadiene ring is in the boat conformation which gives the molecule a fold across the middle as in TMTC. It is reasonable to assume that both C H 2 O H groups are either cis or trans to the carbon atom C1 because the molecule as a whole possesses a two-fold axis of symmetry and this geometrical isomerism, together with the two torsion angles about the = C - C H 2 O H bonds, constitute the only variable parameters in the molecular conformation. The conformation of toxiferin I is thus a two-parameter problem. These two parameters are the torsion angles* 31 = C 1 - C 2 - C 3 - C 4 and 32 = C2-C3-C4-01 (ref. 16). The former of these must be either 0 or 180 ° because of the double bond, and the latter may assume a general value. In toxiferin II and the methyl ether of anhydrocalebassin-C 17, 31 is antiplanar, 180 °. Examination of a space-filling C P K molecular model of the proposed structure indicates that when 31 ~ 180 °, 32 may be only + 60 or 180°: other conformations are disallowed because of steric hindrance. The ease of ring closure in this series to produce such molecules as strychnine and caracurine II suggests that the preferred conformation is that in which the hydroxy groups most closely approach the central diazaoctadiene ring. This is the conformation with r2 = -}-60° and is the conformation illustrated in Fig. 5. The similarity in size, shape, N + - N + distance and distribution of hydrophilic groups of this molecule and those of T M T C is very marked. We calculate that the N + - N + separation is approximately 1060 pm and that the O - O distance is approximately 950 pm. The oxygen atoms correspond very closely in separation and disposition relative to the two charged nitrogen atoms and the rest of the molecule with 0 2 and 0 4 in TMTC. This proposed conformation of toxiferin I results in a * The torsion angle of the the planes A X Y and X A X an d negative 0 ~ termed syn(peri)planar,
b o n d e d group A - X - Y - B is the angle in the N e w m a n proj e c t i on between Y B. It is positive 0 ~ -u180 ° if clockwise from A X to Y B viewed from - 180 ° if counterclockwise. Values of ~ = 0, zk 60, ~ 120 and 180 ° are ± synclinal, ± anticlinal and a nt i (pe ri )pl a na r, respectively. See ref. 16.
NEUROMUSCULAR BLOCKING AGENTS" STRUCTURE AND ACTIVITY
357
CH 3
CH I CH2OH
J
HC
I
CH2OH
c.II
I
CH 3
Fig. 5. The proposed conformation of toxiferin I projected in the same way as caracurine II and toxiferin II, down the two-fold axis of molecular symmetry. molecule which is again convex-hydrophilic and concave-hydrophobic and folds about a line drawn through the two oxygen atoms. Of the remaining alkaloids in this group of which the chemical constitutions are knov,;n, only two are more potent than (+)-tubocurarine; the other two are rather weak blocking agents. The two more potent ~3 are mono- and di-deoxytoxiferin I (C-alkaloids H and K respectively) which must have the same conformation as toxiferin I. They are both less potent than toxiferin I, the approximate E P M R s with respect to (÷)-tubocurarine for complete block on cat gastrocnemius being 0.13, 0.44 and 0.06 respectively 13. Since the completely deoxygenated molecule (C-alkaloid K) is still a very potent blocking agent, it is probable that the principal interaction with the receptor after the quaternary a m m o n i u m groups is hydrophobic and that the oxygen atoms in the other molecules serve only to increase the probability of correct spatial relation to the receptor by an orientation effect rather than by subsidiary binding. The two remaining alkaloids of known chemical structure, which are rather weak blocking agents ~3, are caracurine I and C-alkaloid D. In both of these, hydrogenation and 3,7-fusion of the diazaoctadiene ring of toxiferin I has occurred which decreases the N ÷ - N + separation to 850 pm 5 and accounts for the low potency.
Strychnine and dihydro-fl-erythroidine The crystal structures of these substances have been determined, that of strychnine in two crystals of different salts ~8.~9 and the absolute configuration of dihydro-
358
P. PAULING, T. J. PETCHER
fl-erythroidine determined 6. The conformations of strychnine in the two crystals are essentially identical. Strychnine is better known for its powerful effect on the central nervous system but N-methyl strychnine is a neuromuscular blocking agent (EPMR z 18 relative to (-t-)-tubocurarine on the frog sartorius 2° and N-methyl strychnidine, in which there is no keto oxygen atom, is more potent; E P M R 3.5 (ref. 20). fl-Erythroidine is one of the few tertiary bases with appreciable neuromuscular blocking activity (EPMR z 85) (ref. 13). The dihydro derivative is more potent and has an E P M R of 8.5 on the cat gastrocnemius 21, but N-methylation appears to decrease potency 22,23. The molecules of strychnine and dihydro-fl-erythroidine are shown in Figs. 6 and 7. Strychnine is very closely related to the basic subunit of the alkaloids of calabash curare and it is easy to see that it could interact with half of the receptor area occupied by a curare molecule. The ether oxygen atom makes one face of the molecule more hydrophilic than the other and this characteristic is more marked in strychnidine. The N÷-ring oxygen distance is 479 pm, which is similar to the N1÷-O3 separation in TMTC. The molecule of dihydro-fl-erythroidine correlates quite well with one half of the TMTC molecule. If one superposes the methoxy group of dihydro-fl-erythroidine with O2-C72 of TMTC and the remaining two oxygen atoms with 05, 06, the nitrogen atom occupies a similar position to that of N2 in TMTC. The fit is not as good as
Fig. 6. The conformation of strychnine as observed in crystals of the hydrobromide dihydrate I s. The observed conformation in crystals of the sulfate pentahydrate 19 is almost identical.
0 -/
"O"
Fig. 7. The conformation of dihydro-fl-erythroidine as observed in crystals of the hydrobromide 6.
N E U R O M U S C U L A R B L O C K I N G A G E N T S : S T R U C T U R E A N D ACTIVITY
359
that of strychnine with the alkaloids of calabash curare but accounts satisfactorily for the small potency. The activity constant of fl-erythroidine increases with concentration of the antagonist 24 suggesting that more than one molecule is involved per receptor site. The data are not sufficient, however, to show a square law dose-response which would indicate two molecules per receptor. Gallamine
This trisonium compound (V) was introduced clinically as a neuromuscular blocking agent in 1949 (ref. 25). In ability to reduce the flexing power of the hand in man, it is rather less active than (+)-tubocurarine (EPMR = 4.2) (ref. 26) but it is a true non-depolarizing antagonist of acetylcholine and its effects are reversed by, for example, neostigmine. The conformational analysis involves only the conformations of the side chains and their relative disposition. Various studies of the conformation of the N ÷ - C - C - O group clearly indicate that the preferred conformation is that with the N ÷ - C - C - O torsion angle synclinal and the N + - O distance approximately 325 pm (ref. 27-30). The preferred conformation of trimethoxy-substituted aromatic rings has been discussed elsewhere 31 and one of the results may be extended to apply to this molecule. If an aromatic ring is trimethoxy-substituted at the 1, 2, 3 positions only, then the preferred conformation is that with the methyl substituents at positions 1 and 3 in the plane of the ring and antiplanar to each other and the remaining methyl group out of the plane of the ring. The 1-2-O-Me torsion angle of the central methoxy group must be about 90 °. To apply this argument to gallamine we substitute triethylammoniumethyl for methyl in the above argument bearing in mind that the N + - C C-O group is a group with a preferred conformation. By applying these stereochemical rules, the conformation of gallamine is fixed within certain limits and is shown in Fig. 8. The N1 ÷-N3 + separation is 1060 pm, very similar to that observed in TMTC. We suggest that it is the two nitrogen atoms in meta position which are required for blocking activity and that the triethylammoniumethyl substitution at position 2 could well be substituted. Any suitably bulky group would serve just as well to bring about the necessary relative orientation of N1 and N3.
~
4-
OCHaCH2N (Calls) 3 O CH2CH2 I~ (C2H5) 3
~'~v'/~'~OCH2CH21~(C2H5)3
Form. V - Ga||amine.
THE TEMPLATE FOR CURARIFORM DRUGS
In Fig. 9 we have superposed similar projections of all potent curariform drugs discussed in the previous sections, showing only the charged nitrogen atoms and the oxygen atoms. The N + - N + distances are sufficiently similar that we show only the mean positions, whereas all oxygen atoms are included. We have also superposed an eight-carbon chain in the antiplanar extended conformation between the nitrogen
360
P. PAULING, T. J. PETCHER
+
(C2Hs)3NCH2CH2Q (C2Hsl3NCH2CH20 (C2Hs)31qCH2CH20" Fig. 8. The proposed conformation of gallamine triethiodide. Two equally probable conformations of the central side chain are shown. The N + - C - C - O torsion angles in the other side chains may equally well be positive or negative synclinal but the figure shows that conformation which correlates best with the other molecules.
atoms. All the oxygen atoms are in or above the plane containing the two charged nitrogen atoms and lie in a band approximately 600 pm wide running diagonally across the line joining the two charged nitrogen atoms. Using the cartesian co-ordinate system indicated in the figure (positive Z comes up out of the page) all the oxygen atoms in this composite of potent drugs lie in the two octants of three-dimensional space defined by O ~
O~Y>~--600, O ~
O ~
(+X,--Y,+Z) (--X, + Y , + Z )
where the figures are distances in pm. The oxygen atoms are thus clearly implicated in determining the stereospecificity of curariform drugs since a major difference between, for example, (÷)-tubocurarine and (--)-tubocurarine is the absolute configuration of the distribution of oxygen atoms relative to the nitrogen atoms; the N + - N + separation is necessarily the same but the oxygen atoms of (--)-tubocurarine would appear in the ( + X , + Y , ÷ Z ) and (--X, - - Y , ÷ Z ) octants of Fig. 9. Fig. 9 describes the relationship between three-dimensional structure and potent curariform activity and can be used as a template to synthesise new neuromuscular blocking agents. The structural requirements of any such molecules may be summarised thus:
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
361
YI° \\\\
@
0
@
0
' 2
&_)
.-.. -~
-6"--~ i \,k.
i
f'~_~,.-~_.l
--Q_)4-
"\\\\ ,
,
IOOpm
~"
--
- -
~
~
"~-
"
s
x
I \\~
"~. -5
C
-6"
Fig. 9. A superposition of all the molecules discussed in the text, showing nitrogen and oxygen atoms only. The axial system is graduated in Angstrom units (1 A. = 100 pm) and positive Z comes up out of the page. The oxygen atoms of TMTC are labelled.
(a) Two quaternary nitrogen atoms separated by 1080 ± 30 pm; (b) Rigidity of molecular conformation; (e) A sufficient degree of lipophilicity on the concave side to promote subsidiary hydrophobic interaction with the receptor in the area between the cationic centres; (d) Oxygen atoms suitably disposed about the molecule to aid orientation. The easiest way of achieving (a) is to have the two nitrogen atoms separated by an eight-carbon chain in the antiplanar extended conformation. One of the simplest ways of ensuring the rigidity, (b), and extension of such a chain would be a bis-onium compound based on trans octa(1,3,5,7)tetraene. Alternatively the eight-carbon chain could be part of a fused ring system retaining the unsaturation, or part of a fused saturated ring system which is not free to undergo internal conformational change such as the androstane fragment of pancuronium. One way to satisfy (e) is to determine partition coefficients between octanol and water in the usual way for the tertiary bases of some known curariform drugs and to tailor any new molecules to suit by altering the hydrocarbon content. The best way to satisfy (d) is to superpose a model of the intended molecule on Fig. 9 and to introduce hydroxy, methoxy or acetoxy groups in such a way that the oxygen atoms appear in the appropriate octants of three-dimensional space. In the Appendix we have listed three classes of molecule which should satisfy the criteria listed above.
362
P. PAULING, T. J. PETCHER
DEPOLARIS1NG BLOCKING AGENTS
The outstanding structural feature of depolarising blocking agents such as decamethonium and succinylcholine is their flexibility which contrasts with the rigidity of curariform drugs. The N + - N ÷ separation in decamethonium is 1370 pm in the fully extended antiplanar conformation as observed in the crystal structure 3z but it is known that polymethylene chains curl up in the gas phase 33 and it is probable that this occurs to some extent in solution. GILL34" has suggested probable N + - N ÷ separations for polymethylene bisonium chains in solution from tetra- to octamethonium, and extrapolation of these results gives a value of between 950 and 1100 pm for decamethonium. The structure of the rapidly hydrolysed depolarizing muscle blocking agent succinylcholine has been analysed in crystals of three different salts. In the perchlorate 3~ and the picrate 36 the molecule is crystallographically centrosymmetric, but in the iodide 37 the observed conformation is far from centrosymmetric. The three observed conformations are each different from the others 36, showing that the molecule is highly flexible. The observed N + - N ÷ distances are 1188 pm in the perchlorate, 1064 pm in the picrate and 784 pm in the iodide. Since the observed range of N + - N + distances of the depolarizing muscle blocking agents include the fixed N ÷ - N ÷ distance of the non-depolarising curare agents and their pharmacological actions are related, it is likely that the two types of compounds interact with the same pair of receptor anionic sites. The main requirement for depolarizing blocking activity is a flexible link between the two charged ammonium groups which allows the N ÷ - N + distance to change. While hydrophilic groups appear to be important in curariform agents, they do not seem to be essential to depolarizing blocking agents and a relatively hydrophobic region between the two ammonium groups is acceptable and possibly desirable. THE RECEPTOR
It is likely that the curares and the depolarizing decamethonium-like blocking agents interact with the same pair of anionic sites because of the similar N ÷ - N + separations and related actions. The difference in mode of action has been explained. Decamethonium produces blockade accompanied by depolarisation of the membrane which is identical to the effect of excess ACh, while the curares produce blockade with no change in the membrane resting potential. It is possible that depolarisation is brought about by some conformational change in the receptor 38'39. Decamethonium, succinylcholine and similar molecules are flexible and do not hinder conformational change in the receptor upon interaction of the - N ÷ groups with anionic sites. The curares, however, are rigid and prevent conformational change in the receptor. They thus provide information about the resting separation of anionic sites in the receptor (1080 i 30 pm) and the distribution of hydrophobic and hydrophilic groups in the immediately surrounding area. The complement of Fig. 9 would show the relationships. We propose that with depolarization, the anionic sites of the
NEUROMUSCULARBLOCKINGAGENTS: STRUCTUREAND ACTIVITY
363
receptor move closer together, to about 800-900 pm apart. It is likely that one or both anionic sites of the blocking receptor are the sites of action of ACh. If so, this correlation of the structural features of neuromuscular blocking agents provides information about the local environment and spatial distribution of ACh receptors. APPENDIX Possible new neuromuscular blocking agents. Type 1. Substituted 1,2,5,6,6a,7,8,11,12,12a-decahydrochrysenes. y, x
~
X'
Y X,X' is any of-N+(CH3)s, Me
Me
Y,Y' is any of-OH, -OCHa, -O'CO'CHa substituted at any of positions 6, 6a, 7, 8 combined with any of positions 12, 12a, 1, 2. The oxygen-containing substituents should both be above the plane of the ring system as drawn if possible.
Type 2. Substituted 1,2,3,5,6,6a,7,7a,8,9-decahydro benz[a]anthracenes. yI
X ~ Y
X~
Substitution as in type 1 ; --Y substitution in any combination of positions 7, 7a, 8, 9 with 1, 2, 3 and the same conditions pertaining.
Type 3. Steriods, 5~ series: Gonane, oestrane, androstane.
X'
X Y
364
P. PAULING, T. J. PETCHER X : 2fl,16fl-di( N - m e t h y l p i p e r i d i n o ) - o r 2 f l , 1 6 f l - d i ( N - m e t h y l p y r r o l i d i n o ) - . Y , Y ' : as i n t y p e s 1 a n d 2 in a n y c o m b i n a t i o n o f p o s i t i o n s 3, 4, 5, 6 w i t h l l , 12,
(13), 17. L a r g e r s u b s t i t u t i o n t h a n M e a t p o s i t i o n s l 0 a n d 13 w h e r e a p p r o p r i a t e m i g h t b e u s e f u l i n m a i n t a i n i n g c o n f o r m a t i o n a l inflexibility. Pancuronium
is 3 t 3 , 1 7 / 3 - d i a c e t o x y - 2 f l , 1 6 f l - d i - ( N - m e t h y l
piperidino)-5ct-andro-
stane. ACKNOWLEDGEMENTS W e t h a n k t h e M e d i c a l R e s e a r c h C o u n c i l f o r s u p p o r t , M r . JOHN CRESSWELL f o r t h e figures, M i s s M . DELLOW a n d M i s s P. BRENNAN f o r a s s i s t a n c e a n d D r . BIRTHE JENSEN f o r c o m m u n i c a t i o n
of her results in advance of publication.
REFERENCES 1 H . R . ING, The curariform action of onium salts, Physiol. Rev., 16 (1936) 527. 2 P.G. WASER, in D. BOVET(Ed.), Curare and Curare-like Agents, Elsevier, Amsterdam, 1959, p. 244. 3 P . G . WASER, Calebassen-Curare, Helv. Physiol. Pharm. Acta, 11, Suppl. 8 (1953) 4 A . R . BATTERSBYAND H. F. HODSON, Alkaloids of calabash-curare and strychnos species, Quart. Rev. Chem. Soc., 14 (1960) 77. 5 A . T . McPH~IL AND G. A. SIM, The structure of curacurine-lI: X-ray analysis of curacurine-ll dimethiodide, J. Chem. Soc., (1965) 1663. 6 A . W . HANSON, The crystal structure of dihydro-fl-erythroidine hydrobromide, Acta Cryst., 16, (1963) 939. 7 M. FEHLMANN, H. KOYAMA AND A. NIGGLI, Die Struktur des Alkaloids C-Calebassin, Helv. Chim. Acta, 48 (1965) 303. 8 H . M . SOBELL,T. D. SAKORE,S. S. TAVALE,F. G. CANEPA,P. J. PAULING AND T. J. PETCHER, Stereochemistry of a curare alkaloid: O,O, N-trimethyl-D-tubocurarine, Proc. Natl. Acad. Sci. (U.S.), 69. (1972) 2212. 9 A . J . EVERETT, L. A. LAUE AND S. WILKINSON, Revision of the structures of (÷)-tubocurarine chloride and (+)-chondrocurine, Chem. Commun., 0970) 1020. l0 R. B. BARLOW, in Introduction to Chemical Pharmacology, 2nd ed., Methuen, London, 1968, p. 123. 11 A . F . CAMERON, G. FERGUSON, C. HANNAWAY, I. R. MACKAY AND D. S. SAVAGE,Molecular structure of pancuronium bromide (3ct, 17fl-diacetoxy-2fl, 16fl-dipiperidino-5ct-androstane dimethobromide), a neuromuscular blocking agent. Cry~tal and molecular structure of the water: methylene chloride solvate, J. Chem. Soc. (B), (1971) 410. 12 W. L. M. BAIRD AND m. M. REID, The neuromuscular blocking properties of a new steroid compound, pancuronium bromide, Brit. J. Anaesth., 39 (1967) 775. 13 R . B . BARLOW, in Introduction to Chemical Pharmacology, 2nd ed., Methuen, London, 1968, p. 131f. 14 A. R. BATTERSBY,personal communication (1970) 15 W . O . M . PATON AND W. L. M. PERRY, The pharmacology of toxiferines, Brit. J. Pharmacol., 6 (1951) 299. 16 W. KLYNE AND V. PRELOG, Description of steric relationships across single bonds, Experientia, 16 (1960) 521. 17 K . W . GEMMEE, J. MONTEATH ROBERTSON, G. A. SIM, K. BERNAUER, A. GUGGlSBERG, M. HESSE, H. SCHMIDT AND P. KARRER, Die Struktur des Anhydro-isocalebassinmethyl~tthers; s/iurekatalysierte Umlagerungen des Curare Alkaloids C-Calebassin, Helv. Chim. Acta, 52. (1969) 689. 18 J. H. ROBERTSON ANO C. A. BEEVERS, The crystal structure of strychnine hydrogen bromide, Acta Cryst., 4 (1951) 270. 19 C. BOKHOVEN, J. C. SCHOONE AND J. M. BIJVOET, The Fourier synthesis of the crystal structure of strychnine sulphate pentahydrate, Acta Cryst., 4 (1951) 275.
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
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P. KARRER, C. H. EUGSTER AND P. WASER, U b e r C u r a r e w i r k u n g einiger Strychnidin- u n d Dihydrostrychnidin-chlor-Alkylate, Heir. Chim. Acta, 32 (1949) 2381. 21 S. SALAMA AND S. WRIGHT, Action o f calabash curare and related curariform substances on the central n e r v o u s system o f the cat, Brit. J. Pharmacol., 6 (1951) 459. 22 K . R . UNNA AND J. G. GRESLIN, P h a r m a c o l o g i c action of erythrina alkaloids, I i. Free, liberated a n d c o m b i n e d alkaloids, J. Pharmacol., 80 (1944) 53. 23 K . R . UNNA, M. KNIASUK AND J. G. GRESLIN, P h a r m a c o l o g i c action o f erythrina alkaloids, I fl-Erythroidine and substances derived from it, J. Pharmacol., 80 (1944) 39. 24 E. F. VAN MAANEN, T h e a n t a g o n i s m between acetylcholine a n d the curare alkaloids, D-tubocurarine, c-curarine-I, c-toxiferine-ll a n d fl-erythroidine in the rectus a b d o m i n i s o f the frog, J. Pharm. Exptl. Therap., 99 (1950) 255. 25 R . B . BARLOW, Introduction to ChemicalParmacology, 2rid ed., Methuen, L o n d o n , 1968, p. 125. 26 W . W . MUSHIN, R. WIEN, D. F. J. MASON AND G° T. LANGSTON, Curare-like actions oftri(diethylaminoethoxy)-benzene triethyliodide, Lancet, 1 (1949)726. 27 F . G . CANEPA, P. J. PAULING AND H. S()RUM, Structure o f acetylcholine a n d other substrates of cholinergic systems, Nature, 210 (1966) 907. 28 M. SUNDARALINGAM, C o n f o r m a t i o n a l relationship between the O - C - C - N + system o f phospholipids and substrates o f muscarinic and cholinergic systems, Nature, 217 (1968) 35. 29 R . W . BAKER, C. H. CHOTHIA, P. J. PAULING AND Z. J. PETCHER, Structure and activity o f m u s carinic stimulants, Nature, 230 (1971) 439. 30 P. PARTINGTON, J. FEENEY AND A. S. V. BURGEN, T h e c o n f o r m a t i o n of acetylcholine and related c o m p o u n d s in a q u e o u s solution as studied by nuclear magnetic resonance spectroscopy: Mol. Pharmacol., 8 (1972) 269. 31 C . H . CHOTHIA AND P. J. PAULING, O n the c o n f o r m a t i o n s o f hallucinogenic molecules a n d their correlation, Proc. Natl. Acad. Sci. (U.S.), 63 (1969) 1063. 32 K. LONSDALE, H. J. MILEEDGE AND L. M. PANT, Structural studies o f hexa- and d e c a m e t h o n i u m bromides, ( C H a ) a N ( C H 2 ) n N ( C H a ) a • 2 Br' 2 H 2 0 , in relation to their p h a r m a c o l o g i c a l action, Acta Cryst., 19 (1965) 827. 33 L. S. BARTELL AND O. A. KOHL, Structure and rotational isomerization in free h y d r o c a r b o n chains, Acta Cryst., 16 (1963) A55. 34 E. W. GILL, Inter-quaternary distance and ganglion-blocking activity in bis-quaternary comp o u n d s , Proc. Roy. Soc., B, 150 (1959) 381. 35 B. JENSEN, T h e crystal structure o f succinylcholine perchlorate, Acta Chem. Scand., 25, (1971) 3388. 36 B. JENSEN, T h e crystal structure o f succinylcholine picrate, Acta Chem. Scand., 27 (1973) in press. 37 B. JENSEN, T h e crystal structure o f succinylcholine iodide, Acta Chem. Scand., 24 (1970) 2517. 38 J. MONOD, J. CHANGEUX AND F. JACOB, Allosteric proteins a n d cellular control systems, J. Mol. Biol., 6, (1963) 306. 39 J. MONOD, J. WYMAN AND J. CHANGEUX, O n the n a t u r e o f allosteric transitions: A plausible model, J. Mol. Biol., 12, (1965) 88.
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© Elsevier Scientific Publishing Company, Amsterdam--Printed in The Netherlands
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY*
PETER PAULING ANDTREVOR J. PETCHER** William Ramsay, Ralph Forster and Christopher Ingold Laboratories, University College London Gower Street, London WC 1 (Great Britain)
(Received February 7th, 1973)
SUMMARY Correlation of crystal structure analyses and model building of rigid molecules indicate the structural requirements for curariform activity and the structural differences between depolarizing and non-depolarizing neuromuscular blocking agents.
INTRODUCTION We discuss here the correlation between three-dimensional structure and physiological activity of potent neuromuscular blocking agents 1-4. The structures considered have been determined by X-ray diffraction analysis 5-7 or derived from known crystal structures by use of atomic models and the application of accepted stereochemical principles. Some structural rules for potent neuromuscular blocking activity are presented and some proposals are made for the synthesis of new neuromuscular blocking agents. There is a significant structural difference between depolarizing and non-depolarizing blocking agents which provides information about the mechanism of the cholinergic neuromuscular receptors. The structural correlation has been based on the crystal structure of O,O',N-trimethyl-(÷)-tubocurarine di-iodide 8 and we describe the structural features of other potent neuromuscular blocking agents in terms of the pharmacodynamic groups of this molecule. Tubocurarine
The conformation of T M T C , in crystals of the di-iodide a is shown in Fig. 1. The molecule is sufficiently rigid that it is likely that no change in conformation can result in going from the crystal to solution, or upon interaction with the hypothetical receptor substance. The forces involved in maintaining the molecular rigidity are for * This work was supported by the Medical Research Council of Great Britain. ** Present address: Research Laboratories for Pharmaceutical Chemistry, Sandoz Ltd., Basle (Switzerland). Abbreviations: CPK, Corey, Pauling, Koltun; EPMR, equipotent molar ratio; TMTC, O,O',Ntrimethyl-(+)-tubocurarine.
352
P. PAULING, T. J. PETCHER
C H 3 0 ~
O
CH 2
CH2
OCH30
/
\
v
v
-OCH3
Fig. 1. The conformation of O,O',N-trimethyl-(+)-tubocurarine as observed in crystals o f the diiodide s. The molecule is projected onto the plane o f N1, N2, 02, 04.
the most part repulsive interactions between hydrogen atoms. The N + - N + distances in the two independent molecules of T M T C in the unit cell are observed to be 1060 and 1075 pm*. The conformations of the two independent molecules in the unit cell are essentially identical. The most striking feature of the gross conformation of the molecule is that there is a fold about the line in which lie the six oxygen atoms which endows the two major faces of the molecule with entirely different properties. One face, the upper in Fig. 1, is largely hydrophilic, is convex, and has all the lone pairs of electrons on the oxygen atoms directed outwards from the ridge which they occupy. The reverse face, in contrast, is concave and almost entirely hydrophobic. These features immediately suggest that the receptor area responsible for binding curare-like molecules might have, in addition to two anionic sites separated by approximately 1080 pm to bind ammonium groups, a hydrophilic cleft capable of dipole-dipole interaction with the oxygen atoms and a hydrophobic ridge opposite this cleft capable of interacting with the concave hydrophobic side of the molecule. T M T C was formerly known as O,O'-dimethyltubocurarine but it has recently been shown 9 that the previously accepted formulation of (+)-tubocurarine is incorrect. The correct formula is (I). (+)-Tubocurarine contains one quaternary nitrogen atom and one tertiary nitrogen atom. T M T C is approximately 10 times more potent than tubocurarine and this increase, which was previously ascribed entirely to O-methylation 1° is presumably partially due to quaternisation of the tertiary nitrogen atom, which in tubocurarine is largely protonated at physiological pH. * 100 picometers (pm, 10. * 2 m) equals 1 Angstrom.
353
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
CH30~
+
0
CH2
Form. I - Tubocurarine. Pancuronium The crystal structure o f this steroid derivative (3~,17fi-diacetoxy-2fl,16fl-di(Nm e t h y l p i p e r i d i n o ) - 5 ~ - a n d r o s t a n e ) has recently been d e t e r m i n e d 11, a n d the absolute configuration is k n o w n b y relation to that o f the p a r e n t steroid. The c o n f o r m a t i o n is shown in Fig. 2. This molecule is a p o t e n t n e u r o m u s c u l a r b l o c k i n g agent o f clinical i m p o r t a n c e since it a p p e a r s to be largely free o f the undesirable side effects such as
b
Fig. 2. The conformation of pancuronium in crystals of the bromide 11, projected onto the backbone chain between the nitrogen atoms. The crystals also contain solvent molecules of water and methylene chloride.
354
P. PAULING, T. J. PETCHER
tachycardia 1,2 associated with conventional blocking agents such as gallamine and is only one-tenth as potent as hexamethonium at ganglia 12. In man, the EPMR is approximately 0.2 with respect to (q-)-tubocurarine 12. The molecule is entirely rigid because repulsive interactions between the piperidine rings and the steroid nucleus effectively preclude any boatchair interconversion in the androstane fragment. The N + - N + separation is 1108 pm, the steroid and piperidine rings form an entirely hydrophobic mass down the line joining the two nitrogens but the oxygen atoms of the two acetoxy groups are not similarly disposed to those of TMTC. One interesting feature of pancuronium is the eight-carbon chain held rigidly in an almost antiplanar conformation joining the two charged nitrogen atoms. We shall return to this point in the discussion. Calabash curarines
All potent neuromuscular blocking agents obtained from the alkaloids of calabash curare contain two of the same fused five-ring units (II) joined together in different ways 13. The conformation but not the absolute configuration of the basic building unit has been established in the crystal structure of caracurine lI (Fig. 3) (ref. 5) and one may build space-filling molecular models of these compounds with some confidence, since there are only restricted ways of joining together the two basic units. The absolute configuration of the subunit has been established 14 by relation to that of (--)-strychnine and corresponds to the published atomic co-ordinates of caracurine II. This is the absolute configuration that we have used. CH 3
Form. II - Subuni t o f c a l a b a s h curarine.
CH 3
HOCH2 J 3,CH
CH II II
2'
CH I
J 4CH2 OH
J pe
CH~
Form. l l I - Toxiferin I.
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
355
i?
CH3 Fig. 3. T h e c o n f o r m a t i o n o f caracurine II in crystals o f the dimethiodide 5, d e m o n s t r a t i n g the configuration o f the basic s u b u n i t o f calabash curarines, projected down the two-fold axis o f molecular symmetry.
CH3 N
CH3
I
H .....
CH
..... H
J
CH3
CH 3
Fig. 4. T h e c o n f o r m a t i o n o f toxiferin II in crystals o f the di-iodide 7, projected down the axis o f twofold molecular s y m m e t r y .
356
P. PAULING, T. J. PETCHER
The conformation of the potent neuromuscular blocking agent toxiferin II (C-calebassine) has been determined in crystals of the di-iodide 7, and is shown in Fig. 4. The N + - N ÷ distance is approximately 1060 p m and the molecule has a convex hydrophilic face, a concave hydrophobic face, and folds about a line through the two oxygen atoms which are separated by approximately 480 pm. The conformation of the five-ring subunit is identical to that observed in the structure of caracurine II (ref. 5).
F o r m . IV - D i a z a o c t a d i e n e ring in toxiferin I.
One of the most potent molecules in this group is toxiferin I (III) (ref. 15). The two subunits are joined by - N - C = bonds to form a 1,5-diazaocta-3,7-diene ring (IV) at the centre of the molecule. There is only one way of joining the two subunits so that the resulting molecule correlates with TMTC. The two uncharged nitrogen atoms are trigonal and the diazaoctadiene ring is in the boat conformation which gives the molecule a fold across the middle as in TMTC. It is reasonable to assume that both C H 2 O H groups are either cis or trans to the carbon atom C1 because the molecule as a whole possesses a two-fold axis of symmetry and this geometrical isomerism, together with the two torsion angles about the = C - C H 2 O H bonds, constitute the only variable parameters in the molecular conformation. The conformation of toxiferin I is thus a two-parameter problem. These two parameters are the torsion angles* 31 = C 1 - C 2 - C 3 - C 4 and 32 = C2-C3-C4-01 (ref. 16). The former of these must be either 0 or 180 ° because of the double bond, and the latter may assume a general value. In toxiferin II and the methyl ether of anhydrocalebassin-C 17, 31 is antiplanar, 180 °. Examination of a space-filling C P K molecular model of the proposed structure indicates that when 31 ~ 180 °, 32 may be only + 60 or 180°: other conformations are disallowed because of steric hindrance. The ease of ring closure in this series to produce such molecules as strychnine and caracurine II suggests that the preferred conformation is that in which the hydroxy groups most closely approach the central diazaoctadiene ring. This is the conformation with r2 = -}-60° and is the conformation illustrated in Fig. 5. The similarity in size, shape, N + - N + distance and distribution of hydrophilic groups of this molecule and those of T M T C is very marked. We calculate that the N + - N + separation is approximately 1060 pm and that the O - O distance is approximately 950 pm. The oxygen atoms correspond very closely in separation and disposition relative to the two charged nitrogen atoms and the rest of the molecule with 0 2 and 0 4 in TMTC. This proposed conformation of toxiferin I results in a * The torsion angle of the the planes A X Y and X A X an d negative 0 ~ termed syn(peri)planar,
b o n d e d group A - X - Y - B is the angle in the N e w m a n proj e c t i on between Y B. It is positive 0 ~ -u180 ° if clockwise from A X to Y B viewed from - 180 ° if counterclockwise. Values of ~ = 0, zk 60, ~ 120 and 180 ° are ± synclinal, ± anticlinal and a nt i (pe ri )pl a na r, respectively. See ref. 16.
NEUROMUSCULAR BLOCKING AGENTS" STRUCTURE AND ACTIVITY
357
CH 3
CH I CH2OH
J
HC
I
CH2OH
c.II
I
CH 3
Fig. 5. The proposed conformation of toxiferin I projected in the same way as caracurine II and toxiferin II, down the two-fold axis of molecular symmetry. molecule which is again convex-hydrophilic and concave-hydrophobic and folds about a line drawn through the two oxygen atoms. Of the remaining alkaloids in this group of which the chemical constitutions are knov,;n, only two are more potent than (+)-tubocurarine; the other two are rather weak blocking agents. The two more potent ~3 are mono- and di-deoxytoxiferin I (C-alkaloids H and K respectively) which must have the same conformation as toxiferin I. They are both less potent than toxiferin I, the approximate E P M R s with respect to (÷)-tubocurarine for complete block on cat gastrocnemius being 0.13, 0.44 and 0.06 respectively 13. Since the completely deoxygenated molecule (C-alkaloid K) is still a very potent blocking agent, it is probable that the principal interaction with the receptor after the quaternary a m m o n i u m groups is hydrophobic and that the oxygen atoms in the other molecules serve only to increase the probability of correct spatial relation to the receptor by an orientation effect rather than by subsidiary binding. The two remaining alkaloids of known chemical structure, which are rather weak blocking agents ~3, are caracurine I and C-alkaloid D. In both of these, hydrogenation and 3,7-fusion of the diazaoctadiene ring of toxiferin I has occurred which decreases the N ÷ - N + separation to 850 pm 5 and accounts for the low potency.
Strychnine and dihydro-fl-erythroidine The crystal structures of these substances have been determined, that of strychnine in two crystals of different salts ~8.~9 and the absolute configuration of dihydro-
358
P. PAULING, T. J. PETCHER
fl-erythroidine determined 6. The conformations of strychnine in the two crystals are essentially identical. Strychnine is better known for its powerful effect on the central nervous system but N-methyl strychnine is a neuromuscular blocking agent (EPMR z 18 relative to (-t-)-tubocurarine on the frog sartorius 2° and N-methyl strychnidine, in which there is no keto oxygen atom, is more potent; E P M R 3.5 (ref. 20). fl-Erythroidine is one of the few tertiary bases with appreciable neuromuscular blocking activity (EPMR z 85) (ref. 13). The dihydro derivative is more potent and has an E P M R of 8.5 on the cat gastrocnemius 21, but N-methylation appears to decrease potency 22,23. The molecules of strychnine and dihydro-fl-erythroidine are shown in Figs. 6 and 7. Strychnine is very closely related to the basic subunit of the alkaloids of calabash curare and it is easy to see that it could interact with half of the receptor area occupied by a curare molecule. The ether oxygen atom makes one face of the molecule more hydrophilic than the other and this characteristic is more marked in strychnidine. The N÷-ring oxygen distance is 479 pm, which is similar to the N1÷-O3 separation in TMTC. The molecule of dihydro-fl-erythroidine correlates quite well with one half of the TMTC molecule. If one superposes the methoxy group of dihydro-fl-erythroidine with O2-C72 of TMTC and the remaining two oxygen atoms with 05, 06, the nitrogen atom occupies a similar position to that of N2 in TMTC. The fit is not as good as
Fig. 6. The conformation of strychnine as observed in crystals of the hydrobromide dihydrate I s. The observed conformation in crystals of the sulfate pentahydrate 19 is almost identical.
0 -/
"O"
Fig. 7. The conformation of dihydro-fl-erythroidine as observed in crystals of the hydrobromide 6.
N E U R O M U S C U L A R B L O C K I N G A G E N T S : S T R U C T U R E A N D ACTIVITY
359
that of strychnine with the alkaloids of calabash curare but accounts satisfactorily for the small potency. The activity constant of fl-erythroidine increases with concentration of the antagonist 24 suggesting that more than one molecule is involved per receptor site. The data are not sufficient, however, to show a square law dose-response which would indicate two molecules per receptor. Gallamine
This trisonium compound (V) was introduced clinically as a neuromuscular blocking agent in 1949 (ref. 25). In ability to reduce the flexing power of the hand in man, it is rather less active than (+)-tubocurarine (EPMR = 4.2) (ref. 26) but it is a true non-depolarizing antagonist of acetylcholine and its effects are reversed by, for example, neostigmine. The conformational analysis involves only the conformations of the side chains and their relative disposition. Various studies of the conformation of the N ÷ - C - C - O group clearly indicate that the preferred conformation is that with the N ÷ - C - C - O torsion angle synclinal and the N + - O distance approximately 325 pm (ref. 27-30). The preferred conformation of trimethoxy-substituted aromatic rings has been discussed elsewhere 31 and one of the results may be extended to apply to this molecule. If an aromatic ring is trimethoxy-substituted at the 1, 2, 3 positions only, then the preferred conformation is that with the methyl substituents at positions 1 and 3 in the plane of the ring and antiplanar to each other and the remaining methyl group out of the plane of the ring. The 1-2-O-Me torsion angle of the central methoxy group must be about 90 °. To apply this argument to gallamine we substitute triethylammoniumethyl for methyl in the above argument bearing in mind that the N + - C C-O group is a group with a preferred conformation. By applying these stereochemical rules, the conformation of gallamine is fixed within certain limits and is shown in Fig. 8. The N1 ÷-N3 + separation is 1060 pm, very similar to that observed in TMTC. We suggest that it is the two nitrogen atoms in meta position which are required for blocking activity and that the triethylammoniumethyl substitution at position 2 could well be substituted. Any suitably bulky group would serve just as well to bring about the necessary relative orientation of N1 and N3.
~
4-
OCHaCH2N (Calls) 3 O CH2CH2 I~ (C2H5) 3
~'~v'/~'~OCH2CH21~(C2H5)3
Form. V - Ga||amine.
THE TEMPLATE FOR CURARIFORM DRUGS
In Fig. 9 we have superposed similar projections of all potent curariform drugs discussed in the previous sections, showing only the charged nitrogen atoms and the oxygen atoms. The N + - N + distances are sufficiently similar that we show only the mean positions, whereas all oxygen atoms are included. We have also superposed an eight-carbon chain in the antiplanar extended conformation between the nitrogen
360
P. PAULING, T. J. PETCHER
+
(C2Hs)3NCH2CH2Q (C2Hsl3NCH2CH20 (C2Hs)31qCH2CH20" Fig. 8. The proposed conformation of gallamine triethiodide. Two equally probable conformations of the central side chain are shown. The N + - C - C - O torsion angles in the other side chains may equally well be positive or negative synclinal but the figure shows that conformation which correlates best with the other molecules.
atoms. All the oxygen atoms are in or above the plane containing the two charged nitrogen atoms and lie in a band approximately 600 pm wide running diagonally across the line joining the two charged nitrogen atoms. Using the cartesian co-ordinate system indicated in the figure (positive Z comes up out of the page) all the oxygen atoms in this composite of potent drugs lie in the two octants of three-dimensional space defined by O ~
O~Y>~--600, O ~
O ~
(+X,--Y,+Z) (--X, + Y , + Z )
where the figures are distances in pm. The oxygen atoms are thus clearly implicated in determining the stereospecificity of curariform drugs since a major difference between, for example, (÷)-tubocurarine and (--)-tubocurarine is the absolute configuration of the distribution of oxygen atoms relative to the nitrogen atoms; the N + - N + separation is necessarily the same but the oxygen atoms of (--)-tubocurarine would appear in the ( + X , + Y , ÷ Z ) and (--X, - - Y , ÷ Z ) octants of Fig. 9. Fig. 9 describes the relationship between three-dimensional structure and potent curariform activity and can be used as a template to synthesise new neuromuscular blocking agents. The structural requirements of any such molecules may be summarised thus:
NEUROMUSCULAR BLOCKING AGENTS: STRUCTURE AND ACTIVITY
361
YI° \\\\
@
0
@
0
' 2
&_)
.-.. -~
-6"--~ i \,k.
i
f'~_~,.-~_.l
--Q_)4-
"\\\\ ,
,
IOOpm
~"
--
- -
~
~
"~-
"
s
x
I \\~
"~. -5
C
-6"
Fig. 9. A superposition of all the molecules discussed in the text, showing nitrogen and oxygen atoms only. The axial system is graduated in Angstrom units (1 A. = 100 pm) and positive Z comes up out of the page. The oxygen atoms of TMTC are labelled.
(a) Two quaternary nitrogen atoms separated by 1080 ± 30 pm; (b) Rigidity of molecular conformation; (e) A sufficient degree of lipophilicity on the concave side to promote subsidiary hydrophobic interaction with the receptor in the area between the cationic centres; (d) Oxygen atoms suitably disposed about the molecule to aid orientation. The easiest way of achieving (a) is to have the two nitrogen atoms separated by an eight-carbon chain in the antiplanar extended conformation. One of the simplest ways of ensuring the rigidity, (b), and extension of such a chain would be a bis-onium compound based on trans octa(1,3,5,7)tetraene. Alternatively the eight-carbon chain could be part of a fused ring system retaining the unsaturation, or part of a fused saturated ring system which is not free to undergo internal conformational change such as the androstane fragment of pancuronium. One way to satisfy (e) is to determine partition coefficients between octanol and water in the usual way for the tertiary bases of some known curariform drugs and to tailor any new molecules to suit by altering the hydrocarbon content. The best way to satisfy (d) is to superpose a model of the intended molecule on Fig. 9 and to introduce hydroxy, methoxy or acetoxy groups in such a way that the oxygen atoms appear in the appropriate octants of three-dimensional space. In the Appendix we have listed three classes of molecule which should satisfy the criteria listed above.
362
P. PAULING, T. J. PETCHER
DEPOLARIS1NG BLOCKING AGENTS
The outstanding structural feature of depolarising blocking agents such as decamethonium and succinylcholine is their flexibility which contrasts with the rigidity of curariform drugs. The N + - N ÷ separation in decamethonium is 1370 pm in the fully extended antiplanar conformation as observed in the crystal structure 3z but it is known that polymethylene chains curl up in the gas phase 33 and it is probable that this occurs to some extent in solution. GILL34" has suggested probable N + - N ÷ separations for polymethylene bisonium chains in solution from tetra- to octamethonium, and extrapolation of these results gives a value of between 950 and 1100 pm for decamethonium. The structure of the rapidly hydrolysed depolarizing muscle blocking agent succinylcholine has been analysed in crystals of three different salts. In the perchlorate 3~ and the picrate 36 the molecule is crystallographically centrosymmetric, but in the iodide 37 the observed conformation is far from centrosymmetric. The three observed conformations are each different from the others 36, showing that the molecule is highly flexible. The observed N + - N ÷ distances are 1188 pm in the perchlorate, 1064 pm in the picrate and 784 pm in the iodide. Since the observed range of N + - N + distances of the depolarizing muscle blocking agents include the fixed N ÷ - N ÷ distance of the non-depolarising curare agents and their pharmacological actions are related, it is likely that the two types of compounds interact with the same pair of receptor anionic sites. The main requirement for depolarizing blocking activity is a flexible link between the two charged ammonium groups which allows the N ÷ - N + distance to change. While hydrophilic groups appear to be important in curariform agents, they do not seem to be essential to depolarizing blocking agents and a relatively hydrophobic region between the two ammonium groups is acceptable and possibly desirable. THE RECEPTOR
It is likely that the curares and the depolarizing decamethonium-like blocking agents interact with the same pair of anionic sites because of the similar N ÷ - N + separations and related actions. The difference in mode of action has been explained. Decamethonium produces blockade accompanied by depolarisation of the membrane which is identical to the effect of excess ACh, while the curares produce blockade with no change in the membrane resting potential. It is possible that depolarisation is brought about by some conformational change in the receptor 38'39. Decamethonium, succinylcholine and similar molecules are flexible and do not hinder conformational change in the receptor upon interaction of the - N ÷ groups with anionic sites. The curares, however, are rigid and prevent conformational change in the receptor. They thus provide information about the resting separation of anionic sites in the receptor (1080 i 30 pm) and the distribution of hydrophobic and hydrophilic groups in the immediately surrounding area. The complement of Fig. 9 would show the relationships. We propose that with depolarization, the anionic sites of the
NEUROMUSCULARBLOCKINGAGENTS: STRUCTUREAND ACTIVITY
363
receptor move closer together, to about 800-900 pm apart. It is likely that one or both anionic sites of the blocking receptor are the sites of action of ACh. If so, this correlation of the structural features of neuromuscular blocking agents provides information about the local environment and spatial distribution of ACh receptors. APPENDIX Possible new neuromuscular blocking agents. Type 1. Substituted 1,2,5,6,6a,7,8,11,12,12a-decahydrochrysenes. y, x
~
X'
Y X,X' is any of-N+(CH3)s, Me
Me
Y,Y' is any of-OH, -OCHa, -O'CO'CHa substituted at any of positions 6, 6a, 7, 8 combined with any of positions 12, 12a, 1, 2. The oxygen-containing substituents should both be above the plane of the ring system as drawn if possible.
Type 2. Substituted 1,2,3,5,6,6a,7,7a,8,9-decahydro benz[a]anthracenes. yI
X ~ Y
X~
Substitution as in type 1 ; --Y substitution in any combination of positions 7, 7a, 8, 9 with 1, 2, 3 and the same conditions pertaining.
Type 3. Steriods, 5~ series: Gonane, oestrane, androstane.
X'
X Y
364
P. PAULING, T. J. PETCHER X : 2fl,16fl-di( N - m e t h y l p i p e r i d i n o ) - o r 2 f l , 1 6 f l - d i ( N - m e t h y l p y r r o l i d i n o ) - . Y , Y ' : as i n t y p e s 1 a n d 2 in a n y c o m b i n a t i o n o f p o s i t i o n s 3, 4, 5, 6 w i t h l l , 12,
(13), 17. L a r g e r s u b s t i t u t i o n t h a n M e a t p o s i t i o n s l 0 a n d 13 w h e r e a p p r o p r i a t e m i g h t b e u s e f u l i n m a i n t a i n i n g c o n f o r m a t i o n a l inflexibility. Pancuronium
is 3 t 3 , 1 7 / 3 - d i a c e t o x y - 2 f l , 1 6 f l - d i - ( N - m e t h y l
piperidino)-5ct-andro-
stane. ACKNOWLEDGEMENTS W e t h a n k t h e M e d i c a l R e s e a r c h C o u n c i l f o r s u p p o r t , M r . JOHN CRESSWELL f o r t h e figures, M i s s M . DELLOW a n d M i s s P. BRENNAN f o r a s s i s t a n c e a n d D r . BIRTHE JENSEN f o r c o m m u n i c a t i o n
of her results in advance of publication.
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