WATER-SOLUBLE
STEROIDAL
ANAESTHETICS
G. H. PHILLIPPS, B. E. AYRES.E. J. BAILEY,G. B. EWAN, B. E. LOCIKER and P. J. MAY Organic Chemistry Department. Glaxo-Allenburys Research (Greenford) Ltd., Greenford Road. Greenford. Middlesex. England
SUMMARY With the aim of finding a water-soluble intravenous general anaesthetic we have examined a range of steroids carrying a basic substituent at the I l-position. Compounds with 1Ij?-diaIkyIaminoacyIoxyor I lo-dialkyiamin~subs~ituen~s formed water-soluble salts that showed the desired activity in mice. The f lp-esters can be prepared via the corresponding t Ifi-haloacetoxy compounds. The I tz-dialkylamines can be prepared by reduction of I I -oximes and subsequent aIkyIation. Structure-activity studies have involved modification of the solubilising group, variation of substituents on ring A and at the 17-position and introduction of double bonds. 1~~-Dimethy~mino-2~~thoxy-3z-hydroxy-Sz-pregnan-20-one (minaxolone) forms salts that are readily soluble in water at physiologically acceptabIe pH to give stable solutions of high anaesthetic potency.
INTRODUCTION In a previous paper [I] we reviewed the history of steroid anaesthesia which led to the clinically useful formulation of a mixture of ?I%-hydroxyJr-pregnane- I I .20-dione (If (alphaxalone) and 2 1-acetoxy-3rhydroxy-So-pre~~e-I I,Z@dione (alphadolone acetate) in aqueous Cremophor EL, known as Althesin*, and described structural modifications which led to improved activity in mice or water-soiubility. Althesin occasionally causes a histaminoid response in man [2]: we have had reports of adverse effects of varying severity, about 1 per 10,000 doses sold. This occasional response in man may be akin to the consistently adverse effects shown in dogs given aqueous Cremophor EL alone, although one cannot rule out the possibility that man may sometimes react adversefy to the steroidal components of Althesin. We have therefore continued to search for water-soluble steroidal anaesthetics so that Cremophor EL is no longer necessary. Such compounds should also retain the advantages of Althesin, which shows high potency, instantaneous induction, minimal cardiovascular and respiratory side effects, and high therapeutic ratio, causes neither thrombophlebitis nor irritation nor paraesthesia and is compatible with common preand post-anaesthetic agents as well as inhalational anaesthetics. Another desirable feature is that the pharmacokinetics should be such that the anaesthetic can be considered not only for short procedures but also, when given by repeated administration or infusion, for prolonged operations. This target has been described as remarkably elusive [3]. Our earlier studies [I], which concentrated on anaIogues of 3r-hydroxy-Sr-ptegnane-11.20-dione (1) established the importance of a 3r-hydroxyl group and demonstrated the effect of a variety of ring Asubstituents on activity. Improvements were noted in activity, particularly with 2/l-alkoxy- and 2/?-alkyl-
substituents, and in therapeutic index, on introduction of double bonds at the 1.2 or 45positions. These effects were discussed in relation to a desirable flexibility of ring A, allowing it to take a conformation similar to that of the “r-boat”. 3r.l la-Dihydroxy-5~-pregnan-2~one and (2) 32.1 lo-dihydroxy-S~-pre~an-2~one (7) were both reported [ 1J to be much less active than alphaxalone (1) but in the present study we have found that a variety of 11-substituted analogues can show useful activity together with water-solubility. METHODS The compounds have been examined by the intravenous route in male mice of the CD1 strain, usually in groups of five. The steroids were given as sofutions in water, unless they were insoluble when they were given as solutions or suspensions in 20%, Cremophor EL, the doses being successively halved from 100 or 200 m&kg down to an ineffective level. immediately after injection, the mice were placed in a cabinet maintained at 35°C. The time between injection and the loss of “righting reflex” was recorded as the induc-tion time and duration of loss of “righting reflex” was recorded as the sleep-time. Deaths and signs of thrombophlebitis were recorded after 24 h. In the tables, the activities are shown in simplified form. Thus each column represents one dose level of a group of five mice. If all mice lost their righting reflexes without an induction period and a11 of them survived, the “sleeptime” in minutes is recorded. Where such a dose was not found, a diagonal line indicates that some of the mice siept and some died and a cross indicates that all of the mice died.
RESULTS AND DISCUSSION The 1lr-acetate (3) was much more active than the 1I a-alcohol (2) but activity was lost on introduction
* Trade Mark of Glaxo Laboratories Ltd. PB. I l/IA--c
79
19
E-OCOCH~N(C,H& a-OCOCH2N_-0 B-OH
I
I
5
6
7
X
3
a-0COCH2N(CH,),
I
/?-OCOCH2N(C,Hs)2 j?-OCOCH,N(C,Hs),
I
I
I
I
I
12
13
14
I5
16
b
&OCOCH ,/N
18
a3
BOCOCH3(
17
,OCO,,,,(
CH3
CH3
&OCOCH,N(C,H,),
/%OCOC.+Hg /?-OCOCH,NH,
I
I
11
fi-OCOC,H,
9
10
@-OCOCHJ
I
I
8
(r-OCOCHJ
Citric
Citric
Citric
2
6
8
5
6
1.5
I2
14
8
Citric
Citric
12
3
Citric
20
25
40
23
8
18
7 8
37
19
I
I8
27
I
2
17
3
6
3
4
50
a-OH
25
I
12.5
2
6.3
3.2
=0
1.6
I
II-
Dose (m&W
I
Other
Acid for salt
Formula
Compound No.
Substituent
Table 1. Sleeptime (mins) in mice of substituted 3z-hydroxypregnan-20-ones
\
\
41
100
64
200
Ez
3
2/?-0C2H5
2/?-OC2H5
2/?-OCsH,
2@-OCaHg
33
34
35
36
30
2/?-Br
2fi-OCHB
32
37
3/S-CH,
6/!f-CH,
31
2/Z?-CH,
29
30
Za-CH
28
/I-0COCH2N \
Ccf3
C4H9
/
@-OCOCH,NI=>-
@-OCOCH,Nc>-
/?-OCOCH,N(C,Hs),
fl-0COCH2N(C,H,)s
&OCOCH2N(CzHs)l
fl-OCOCH,N(C,Hs)s
8-OCOCH,NK,H,),
/3-OCOCH,rcI)
@-OCOCHZN(C,H,)z
BOC~H,N(C,Hsh
j%OCOCH(CH ,)o
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
4
6
4
II
2
3
11
13
4
17
9
21
8
7
3
I
3
11
IO
4
Citric
26
27
10
2
2
Citric
fl-OCOCH,m
24
Citric
25
fi-OCOCH ta
23
Iodide
6
24
S
3
12.5
6.3
S
8-OCOCH&(C,Hs),
22
3.2
HCI
1.6
so
27
2s
38
17
18
15
20
I7
15
20
18
13
X 8
14
12
2s
Dose tmg/W
Citric
Citric
Acid for salt
_
2
&OCOCH,NH
Confinued
Citric
&OCOCH,NH+
20
ll-
21
Other /I-0COCH2NHCH[CH(CH,),],
Formula
19
Compound No.
Substituent
Table 1.
100
200
m
A’
I
I
I
II
II
II
I
I
I
I
I
I
I
I
I
1
I
I
I
I
I
40
41
42
43
44
45
46
41
48
49
So
51
52
53
54
55
56
57
58
59
60
\
Citric Citric
Citric Acetic Citric
11a-N(CH,)r 11s+N(CH,), 1 lz-N(CH,), I Ix-N(CH,), I Ix-N(CH,)~ I Iz-N(CH,), 11 z-N(CH,), I Ir-N(CH,), 1 Iz-N(CH,), I Iz-N(CHs), I Iz-N(CH~)~ I Iz-N(CH3)r
Zz-CH,
2/I-CH,
6/I-CH,
2/I-F
2pJ
2/I-Br
2/I-SCN
2/I-OCH,
2b-0C,H5
2fl-OC,H,-i
A’
A4
Citric
Citric
Citric
Citric
Citric
Citric
Citric
HCI
Citric
Citric
Citric
Citric
Citric
CH,
ll-
Acid for salt
I IwNH,
/%OCOCH,N~
&OCOCH,N(C,H,),
B-OH
/I-OCOCHIN
/
/.I-0COCH,N(C,H5),
WCOCH,N(C,W,
Substituent
1. Continued
I la-N(CH,),
2B-OCzHs
I ‘lab-D-Homo
19-Nor
2jLSCN
Other
I
Formula
39
Compound No.
Table
1.6
5
7
2
3
3.2
14
17 9
14 3 II
19
12
7 9
30
27 16
9
26
12 28
3
21
18
13
14
13
14
25
8
19
7
5
4
14
6
5
12.5
14
9
2
18
5
6.3
Dose (mg/kS)
5
14
50
24
loo
50
15
200
Steroidai anaesthetics
83
84
G.
H.
PHILLIPPS
et d.
Fig. f. Structures of key steroids used in Tables 1 and 2 and Minaxolone. of basic groups (4, 5, 6) into the ester moiety. The 1I@-acyloxy-compounds (8. 9. 10) were more active than the I l/?-alcohol (7) and, encouragingly, activity improved with increasing size of group. The glycinate (11) was inactive but the diethylaminoacetate (12) showed useful activity when administered in aqueous Cremophor EL. Its slightly improved potency when presented as an aqueous solution of its citrate salt (13) may be a reflection of greater bio-availability. Many other physiologically acceptable salts show similar activity but the citrates have generaliy been used because they are conveniently prepared. Further dialkylamin~sters (1418) behaved similarly, as did monoalkylaminoesters in which the alkyl group branched at the r-carbon (1921). The quaternary ammonium derivative (22) was toxic. Activity was retained when the basic nitrogen formed part of a ring (23-27). Methyl substituents on the steroid at the 2x-, 2/I-, 3/j’- or 6/I-positions (2831) were compatible with .activity, as were 2/?-alkoxy-(32-36) halogeno-(37,38) and thiocyanato-(39) substituents. The 1.2~unsaturated (40) and t9-nor (41) analogues were effective, as was the 2~-ethoxy-~homo analogue (42) in which the acetyl side chain was carried at the 17a &position (Table 1). The 5&H analogues (4345) were only weakly active and this can perhaps be attributed to the flex-
ibie ester grouping taking up a different conformation in this series. The lI/?-aminoacetates can be made as shown in Fig. 2, for example, by first forming the 3.11-bis-chloroacetate using chloroacetic anhydride with 4-dimethylaminopyridine as catalyst in order to acylate the hindered 1I/&hydroxyL Selective hydrolysis of the 3-chloroacetoxy-group and subsequent nucleophihc displacement with an amine gives the desired ester: alternatively the reaction with an amine can be carried out prior to the selective hydrolysis. The 1I/&ester function is sterically hindered and therefore is resistant to chemical hydrolysis. Consequently, the aqueous solutions of the salts show good stability at physiologically acceptable pH. The esters are considerably more active than the parent alcohols and this indicates that they are probably active per se and also resistant to esterases. The possibility of instability of solutions due to hydrolysis was completely ruled out in a series of compounds in which an amino group was directly attached to the steroid nucleus. 3%-Hydroxy-%-pregnan-20-one (46) showed activity but the corresponding 1lJr-amine hydr~hloride (47) was toxic. However. N-dimethylation gave the tertiary amine which formed a very water-soluble citrate salt (48) with encouraging activity. The effect of introducing methyl
Fig. 2. Routes for the synthesis of 1lpaminoacetoxy-steroids.
Fig. 3. Routes for the synthesis of 1I*-amino-steroids.
85
Steroidal anaesthetics groups at the 2x-. 2fl- or 6/3-positions (49!!I), various atoms or groups at the 2/3-position (SZ-58), or 1,2or 4,5-unsaturation (59,60) was examined. The highest potency was found for the citrate salt of 1la-dimethylamino-2fl-ethoxy-3c+hydroxy-5~-pregnan-20-one (57) (minaxolone). Routes for the preparation of such compounds are (e.g. shown in Fig. 3. Thus the 2/?-substituent X = OC,H,) can be present before formation of the I I-oxime, which is conveniently carried out with hydroxylamine at >pH 11, reduction with for example sodium in propanol to the 1 lcr-amine and subsequent alkylation on nitrogen and deketalisation. N-Dimethylation is very conveniently carried out by the Eschweiler-Clarke procedure using formic acid (45 equivs) in 40”/, formalin at 100°C for about 15 min. Alternatively, the 11r-dimethylamino-group may be first introduced in the presence of a 2,3-double bond. Epoxidation of the double bond with peracid can be accomplished so long as the dimethylamino-group is protected by protonation, for example as the tosylate salt. If the dimethylamine is not protonated it reacts with ai-chloroperbenzoic acid to give the corresponding N-nt-chlorobenzoyloxymethyl-N-methyl-N-oxide, Minaxolone (57) formed a very water-soluble citrate which could be formulated as stable solutions at a concentration of 0.5% or more at pH 4 or above. Heating of the solutions at 100°C resulted in slow formation of traces of the corresponding inactive 17/?-H isomer and, in case this proved to be troublesome in pharmaceutical presentations, we therefore examined the effect on activity of changing the 17-side chain. Introduction of a 21-acetoxy group (66) was unlikely to improve stability and also did not improve activity. The 17jI-vinyl (67) and 17/%propionyl (68) derivatives were more active than the corresponding I7/?-acetyl compound (48). The D-home-analogue (69) carrying the acetyl side chain at the 17a/?-position was not quite as potent as minaxolone (57). The methylcarboxylate (70) and nitrile (71) were also active. Of these compounds the 17/!Lvinyl androstane (67) might be expected to be the most stable (Table 2). Turning to the SP-series, the 17&acetyl compound (72) showed greater activity -than the corresponding 21-acetoxy-derivative (73) and the nitrile (74) and, inTable 2. Sleeptime
(mins) in mice of substituted
deed, caused sleep at a lower dose than minaxolone (57) although the sleep times at the higher doses were similar. Further tests in other species indicated little difference between these two derivatives, but we eventually chose to progress minaxolone (57) because there was a possibility that a 5/LH compound might show pyrogenic activity apparent only in man, as has been reported for example to exclude the use of 3n-hydroxy-5/I-pregnan-20-one as a hypnotic [4]. Detailed studies of the properties of minaxolone (57) in animals have been described in a communication [5] and these have confirmed that it is not only more active than Alphaxalone (1) in a variety of species but also possesses properties that made it close to the ideal intravenous anaesthetic. One can speculate that this combination of watersolubility and high activity may be associated with the unusual steric situation in the 1la-dimethylaminosteroids. Examination of Dreiding models indicates that the nitrogen atom is very close to one of the protons attached to C (l&the l/&proton if ring A is a chair and the la-proton if ring A is an ‘*a-boat” [I]. Further, although even more speculatively, the nitrogen of an 1 lp-aminoacetate can occupy an almost identical position, although it is perhaps unlikely to prefer such a conformation. The unusual steric hindrance was demonstrable in the ‘H n.m.r. spectrum of an 1 la-dimethylamino-steroid in CDCl,. Whereas at high temperature (+75”C) the two N-methyl groups appeared as a six-proton singlet, indicating that rotation about the C (11)-N bond was occurring normally, as the temperature was reduced the signals became broader and then resolved until at -35°C two distinct 3-proton singlets were observed. Acknowledgements-We would like to thank our colleagues in Pharmacology Department [S] for the provision of the activity data given in this paper. REFERENCES
I. Phillipps G. H.: Structure-activity relationships in steroidal anaesthetics. J. sreroid Biochem. 6 (1975) 607-613. 2. Clarke R. S. J., Dundee J. W., Garrett R. T., McArdle G. K. and Sutton J. A.: Adverse reactions to intravenous anaesthetics. Er. J. Anausrh. 47 (1975) 575-585.
I lr-dimethylamino-3whydroxyandrostane
Substituent Compound No.
Formula
66
III
67
III
68 69 70 71 72 73 74
III III III III IV IV IV
2t3OC,H,
OCzHs OC2H5
OC,Hs
17/ICOCH,OCOCH, CH=CH, COC2Hs D-Homo; a-COCHs C02CHo CN COCH3 COCH20COCH3 CN
citrate
salts
Dose (mg/kg) 1.6
3
3.2
6.3
12.5
25
2
10 10 11
26 25
3
11
8 I
8 12 4 4
5 I6 22 8 15
18
86
G. H.
PHILLIPPS et ul.
3. Spinks A.: Chemistry and anaesthesia. Cherq~. Ind. (1977)475485. 4. Gyermek L. and Soyka L. F.: Steroid anaesthetics. Anrdwsio/og_v 42 (1975) 331-344.
5. Davis B., Dodds M. G.. Twissell D. J. and Valiance D. K.: Minaxolone. a new anaesthetic. J. stwoid Bio&WI. 9 (1978) 841.
STEROIDAL
ANAESTHETICS
G. H. PHILLIPPS, B. E. AYRES.E. J. BAILEY,G. B. EWAN, B. E. LOCIKER and P. J. MAY Organic Chemistry Department. Glaxo-Allenburys Research (Greenford) Ltd., Greenford Road. Greenford. Middlesex. England
SUMMARY With the aim of finding a water-soluble intravenous general anaesthetic we have examined a range of steroids carrying a basic substituent at the I l-position. Compounds with 1Ij?-diaIkyIaminoacyIoxyor I lo-dialkyiamin~subs~ituen~s formed water-soluble salts that showed the desired activity in mice. The f lp-esters can be prepared via the corresponding t Ifi-haloacetoxy compounds. The I tz-dialkylamines can be prepared by reduction of I I -oximes and subsequent aIkyIation. Structure-activity studies have involved modification of the solubilising group, variation of substituents on ring A and at the 17-position and introduction of double bonds. 1~~-Dimethy~mino-2~~thoxy-3z-hydroxy-Sz-pregnan-20-one (minaxolone) forms salts that are readily soluble in water at physiologically acceptabIe pH to give stable solutions of high anaesthetic potency.
INTRODUCTION In a previous paper [I] we reviewed the history of steroid anaesthesia which led to the clinically useful formulation of a mixture of ?I%-hydroxyJr-pregnane- I I .20-dione (If (alphaxalone) and 2 1-acetoxy-3rhydroxy-So-pre~~e-I I,Z@dione (alphadolone acetate) in aqueous Cremophor EL, known as Althesin*, and described structural modifications which led to improved activity in mice or water-soiubility. Althesin occasionally causes a histaminoid response in man [2]: we have had reports of adverse effects of varying severity, about 1 per 10,000 doses sold. This occasional response in man may be akin to the consistently adverse effects shown in dogs given aqueous Cremophor EL alone, although one cannot rule out the possibility that man may sometimes react adversefy to the steroidal components of Althesin. We have therefore continued to search for water-soluble steroidal anaesthetics so that Cremophor EL is no longer necessary. Such compounds should also retain the advantages of Althesin, which shows high potency, instantaneous induction, minimal cardiovascular and respiratory side effects, and high therapeutic ratio, causes neither thrombophlebitis nor irritation nor paraesthesia and is compatible with common preand post-anaesthetic agents as well as inhalational anaesthetics. Another desirable feature is that the pharmacokinetics should be such that the anaesthetic can be considered not only for short procedures but also, when given by repeated administration or infusion, for prolonged operations. This target has been described as remarkably elusive [3]. Our earlier studies [I], which concentrated on anaIogues of 3r-hydroxy-Sr-ptegnane-11.20-dione (1) established the importance of a 3r-hydroxyl group and demonstrated the effect of a variety of ring Asubstituents on activity. Improvements were noted in activity, particularly with 2/l-alkoxy- and 2/?-alkyl-
substituents, and in therapeutic index, on introduction of double bonds at the 1.2 or 45positions. These effects were discussed in relation to a desirable flexibility of ring A, allowing it to take a conformation similar to that of the “r-boat”. 3r.l la-Dihydroxy-5~-pregnan-2~one and (2) 32.1 lo-dihydroxy-S~-pre~an-2~one (7) were both reported [ 1J to be much less active than alphaxalone (1) but in the present study we have found that a variety of 11-substituted analogues can show useful activity together with water-solubility. METHODS The compounds have been examined by the intravenous route in male mice of the CD1 strain, usually in groups of five. The steroids were given as sofutions in water, unless they were insoluble when they were given as solutions or suspensions in 20%, Cremophor EL, the doses being successively halved from 100 or 200 m&kg down to an ineffective level. immediately after injection, the mice were placed in a cabinet maintained at 35°C. The time between injection and the loss of “righting reflex” was recorded as the induc-tion time and duration of loss of “righting reflex” was recorded as the sleep-time. Deaths and signs of thrombophlebitis were recorded after 24 h. In the tables, the activities are shown in simplified form. Thus each column represents one dose level of a group of five mice. If all mice lost their righting reflexes without an induction period and a11 of them survived, the “sleeptime” in minutes is recorded. Where such a dose was not found, a diagonal line indicates that some of the mice siept and some died and a cross indicates that all of the mice died.
RESULTS AND DISCUSSION The 1lr-acetate (3) was much more active than the 1I a-alcohol (2) but activity was lost on introduction
* Trade Mark of Glaxo Laboratories Ltd. PB. I l/IA--c
79
19
E-OCOCH~N(C,H& a-OCOCH2N_-0 B-OH
I
I
5
6
7
X
3
a-0COCH2N(CH,),
I
/?-OCOCH2N(C,Hs)2 j?-OCOCH,N(C,Hs),
I
I
I
I
I
12
13
14
I5
16
b
&OCOCH ,/N
18
a3
BOCOCH3(
17
,OCO,,,,(
CH3
CH3
&OCOCH,N(C,H,),
/%OCOC.+Hg /?-OCOCH,NH,
I
I
11
fi-OCOC,H,
9
10
@-OCOCHJ
I
I
8
(r-OCOCHJ
Citric
Citric
Citric
2
6
8
5
6
1.5
I2
14
8
Citric
Citric
12
3
Citric
20
25
40
23
8
18
7 8
37
19
I
I8
27
I
2
17
3
6
3
4
50
a-OH
25
I
12.5
2
6.3
3.2
=0
1.6
I
II-
Dose (m&W
I
Other
Acid for salt
Formula
Compound No.
Substituent
Table 1. Sleeptime (mins) in mice of substituted 3z-hydroxypregnan-20-ones
\
\
41
100
64
200
Ez
3
2/?-0C2H5
2/?-OC2H5
2/?-OCsH,
2@-OCaHg
33
34
35
36
30
2/?-Br
2fi-OCHB
32
37
3/S-CH,
6/!f-CH,
31
2/Z?-CH,
29
30
Za-CH
28
/I-0COCH2N \
Ccf3
C4H9
/
@-OCOCH,NI=>-
@-OCOCH,Nc>-
/?-OCOCH,N(C,Hs),
fl-0COCH2N(C,H,)s
&OCOCH2N(CzHs)l
fl-OCOCH,N(C,Hs)s
8-OCOCH,NK,H,),
/3-OCOCH,rcI)
@-OCOCHZN(C,H,)z
BOC~H,N(C,Hsh
j%OCOCH(CH ,)o
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
Citric
4
6
4
II
2
3
11
13
4
17
9
21
8
7
3
I
3
11
IO
4
Citric
26
27
10
2
2
Citric
fl-OCOCH,m
24
Citric
25
fi-OCOCH ta
23
Iodide
6
24
S
3
12.5
6.3
S
8-OCOCH&(C,Hs),
22
3.2
HCI
1.6
so
27
2s
38
17
18
15
20
I7
15
20
18
13
X 8
14
12
2s
Dose tmg/W
Citric
Citric
Acid for salt
_
2
&OCOCH,NH
Confinued
Citric
&OCOCH,NH+
20
ll-
21
Other /I-0COCH2NHCH[CH(CH,),],
Formula
19
Compound No.
Substituent
Table 1.
100
200
m
A’
I
I
I
II
II
II
I
I
I
I
I
I
I
I
I
1
I
I
I
I
I
40
41
42
43
44
45
46
41
48
49
So
51
52
53
54
55
56
57
58
59
60
\
Citric Citric
Citric Acetic Citric
11a-N(CH,)r 11s+N(CH,), 1 lz-N(CH,), I Ix-N(CH,), I Ix-N(CH,)~ I Iz-N(CH,), 11 z-N(CH,), I Ir-N(CH,), 1 Iz-N(CH,), I Iz-N(CHs), I Iz-N(CH~)~ I Iz-N(CH3)r
Zz-CH,
2/I-CH,
6/I-CH,
2/I-F
2pJ
2/I-Br
2/I-SCN
2/I-OCH,
2b-0C,H5
2fl-OC,H,-i
A’
A4
Citric
Citric
Citric
Citric
Citric
Citric
Citric
HCI
Citric
Citric
Citric
Citric
Citric
CH,
ll-
Acid for salt
I IwNH,
/%OCOCH,N~
&OCOCH,N(C,H,),
B-OH
/I-OCOCHIN
/
/.I-0COCH,N(C,H5),
WCOCH,N(C,W,
Substituent
1. Continued
I la-N(CH,),
2B-OCzHs
I ‘lab-D-Homo
19-Nor
2jLSCN
Other
I
Formula
39
Compound No.
Table
1.6
5
7
2
3
3.2
14
17 9
14 3 II
19
12
7 9
30
27 16
9
26
12 28
3
21
18
13
14
13
14
25
8
19
7
5
4
14
6
5
12.5
14
9
2
18
5
6.3
Dose (mg/kS)
5
14
50
24
loo
50
15
200
Steroidai anaesthetics
83
84
G.
H.
PHILLIPPS
et d.
Fig. f. Structures of key steroids used in Tables 1 and 2 and Minaxolone. of basic groups (4, 5, 6) into the ester moiety. The 1I@-acyloxy-compounds (8. 9. 10) were more active than the I l/?-alcohol (7) and, encouragingly, activity improved with increasing size of group. The glycinate (11) was inactive but the diethylaminoacetate (12) showed useful activity when administered in aqueous Cremophor EL. Its slightly improved potency when presented as an aqueous solution of its citrate salt (13) may be a reflection of greater bio-availability. Many other physiologically acceptable salts show similar activity but the citrates have generaliy been used because they are conveniently prepared. Further dialkylamin~sters (1418) behaved similarly, as did monoalkylaminoesters in which the alkyl group branched at the r-carbon (1921). The quaternary ammonium derivative (22) was toxic. Activity was retained when the basic nitrogen formed part of a ring (23-27). Methyl substituents on the steroid at the 2x-, 2/I-, 3/j’- or 6/I-positions (2831) were compatible with .activity, as were 2/?-alkoxy-(32-36) halogeno-(37,38) and thiocyanato-(39) substituents. The 1.2~unsaturated (40) and t9-nor (41) analogues were effective, as was the 2~-ethoxy-~homo analogue (42) in which the acetyl side chain was carried at the 17a &position (Table 1). The 5&H analogues (4345) were only weakly active and this can perhaps be attributed to the flex-
ibie ester grouping taking up a different conformation in this series. The lI/?-aminoacetates can be made as shown in Fig. 2, for example, by first forming the 3.11-bis-chloroacetate using chloroacetic anhydride with 4-dimethylaminopyridine as catalyst in order to acylate the hindered 1I/&hydroxyL Selective hydrolysis of the 3-chloroacetoxy-group and subsequent nucleophihc displacement with an amine gives the desired ester: alternatively the reaction with an amine can be carried out prior to the selective hydrolysis. The 1I/&ester function is sterically hindered and therefore is resistant to chemical hydrolysis. Consequently, the aqueous solutions of the salts show good stability at physiologically acceptable pH. The esters are considerably more active than the parent alcohols and this indicates that they are probably active per se and also resistant to esterases. The possibility of instability of solutions due to hydrolysis was completely ruled out in a series of compounds in which an amino group was directly attached to the steroid nucleus. 3%-Hydroxy-%-pregnan-20-one (46) showed activity but the corresponding 1lJr-amine hydr~hloride (47) was toxic. However. N-dimethylation gave the tertiary amine which formed a very water-soluble citrate salt (48) with encouraging activity. The effect of introducing methyl
Fig. 2. Routes for the synthesis of 1lpaminoacetoxy-steroids.
Fig. 3. Routes for the synthesis of 1I*-amino-steroids.
85
Steroidal anaesthetics groups at the 2x-. 2fl- or 6/3-positions (49!!I), various atoms or groups at the 2/3-position (SZ-58), or 1,2or 4,5-unsaturation (59,60) was examined. The highest potency was found for the citrate salt of 1la-dimethylamino-2fl-ethoxy-3c+hydroxy-5~-pregnan-20-one (57) (minaxolone). Routes for the preparation of such compounds are (e.g. shown in Fig. 3. Thus the 2/?-substituent X = OC,H,) can be present before formation of the I I-oxime, which is conveniently carried out with hydroxylamine at >pH 11, reduction with for example sodium in propanol to the 1 lcr-amine and subsequent alkylation on nitrogen and deketalisation. N-Dimethylation is very conveniently carried out by the Eschweiler-Clarke procedure using formic acid (45 equivs) in 40”/, formalin at 100°C for about 15 min. Alternatively, the 11r-dimethylamino-group may be first introduced in the presence of a 2,3-double bond. Epoxidation of the double bond with peracid can be accomplished so long as the dimethylamino-group is protected by protonation, for example as the tosylate salt. If the dimethylamine is not protonated it reacts with ai-chloroperbenzoic acid to give the corresponding N-nt-chlorobenzoyloxymethyl-N-methyl-N-oxide, Minaxolone (57) formed a very water-soluble citrate which could be formulated as stable solutions at a concentration of 0.5% or more at pH 4 or above. Heating of the solutions at 100°C resulted in slow formation of traces of the corresponding inactive 17/?-H isomer and, in case this proved to be troublesome in pharmaceutical presentations, we therefore examined the effect on activity of changing the 17-side chain. Introduction of a 21-acetoxy group (66) was unlikely to improve stability and also did not improve activity. The 17jI-vinyl (67) and 17/%propionyl (68) derivatives were more active than the corresponding I7/?-acetyl compound (48). The D-home-analogue (69) carrying the acetyl side chain at the 17a/?-position was not quite as potent as minaxolone (57). The methylcarboxylate (70) and nitrile (71) were also active. Of these compounds the 17/!Lvinyl androstane (67) might be expected to be the most stable (Table 2). Turning to the SP-series, the 17&acetyl compound (72) showed greater activity -than the corresponding 21-acetoxy-derivative (73) and the nitrile (74) and, inTable 2. Sleeptime
(mins) in mice of substituted
deed, caused sleep at a lower dose than minaxolone (57) although the sleep times at the higher doses were similar. Further tests in other species indicated little difference between these two derivatives, but we eventually chose to progress minaxolone (57) because there was a possibility that a 5/LH compound might show pyrogenic activity apparent only in man, as has been reported for example to exclude the use of 3n-hydroxy-5/I-pregnan-20-one as a hypnotic [4]. Detailed studies of the properties of minaxolone (57) in animals have been described in a communication [5] and these have confirmed that it is not only more active than Alphaxalone (1) in a variety of species but also possesses properties that made it close to the ideal intravenous anaesthetic. One can speculate that this combination of watersolubility and high activity may be associated with the unusual steric situation in the 1la-dimethylaminosteroids. Examination of Dreiding models indicates that the nitrogen atom is very close to one of the protons attached to C (l&the l/&proton if ring A is a chair and the la-proton if ring A is an ‘*a-boat” [I]. Further, although even more speculatively, the nitrogen of an 1 lp-aminoacetate can occupy an almost identical position, although it is perhaps unlikely to prefer such a conformation. The unusual steric hindrance was demonstrable in the ‘H n.m.r. spectrum of an 1 la-dimethylamino-steroid in CDCl,. Whereas at high temperature (+75”C) the two N-methyl groups appeared as a six-proton singlet, indicating that rotation about the C (11)-N bond was occurring normally, as the temperature was reduced the signals became broader and then resolved until at -35°C two distinct 3-proton singlets were observed. Acknowledgements-We would like to thank our colleagues in Pharmacology Department [S] for the provision of the activity data given in this paper. REFERENCES
I. Phillipps G. H.: Structure-activity relationships in steroidal anaesthetics. J. sreroid Biochem. 6 (1975) 607-613. 2. Clarke R. S. J., Dundee J. W., Garrett R. T., McArdle G. K. and Sutton J. A.: Adverse reactions to intravenous anaesthetics. Er. J. Anausrh. 47 (1975) 575-585.
I lr-dimethylamino-3whydroxyandrostane
Substituent Compound No.
Formula
66
III
67
III
68 69 70 71 72 73 74
III III III III IV IV IV
2t3OC,H,
OCzHs OC2H5
OC,Hs
17/ICOCH,OCOCH, CH=CH, COC2Hs D-Homo; a-COCHs C02CHo CN COCH3 COCH20COCH3 CN
citrate
salts
Dose (mg/kg) 1.6
3
3.2
6.3
12.5
25
2
10 10 11
26 25
3
11
8 I
8 12 4 4
5 I6 22 8 15
18
86
G. H.
PHILLIPPS et ul.
3. Spinks A.: Chemistry and anaesthesia. Cherq~. Ind. (1977)475485. 4. Gyermek L. and Soyka L. F.: Steroid anaesthetics. Anrdwsio/og_v 42 (1975) 331-344.
5. Davis B., Dodds M. G.. Twissell D. J. and Valiance D. K.: Minaxolone. a new anaesthetic. J. stwoid Bio&WI. 9 (1978) 841.
