IRREVERSIBLE
Journal of Medicinal Chemistry, 1971, Vol. 14, No. 9 805
ENZYME INHIBITORS. 186
TABLE IV PHYSICAL PROPERTIES OF
0
Yield,
No.
R
Methoda
%
M P (dec). Cb
34 310 290 22 315 29 296 A 18 m-NHCOC~Hz-2,4-Mez-5-S0~F 7 300 A 20 m-NHCOC~Hz-2,4-Cl2-5-S0~F 8 320 A 26 10 m-NHCOC6H3-3-Cl-4-SOzF A 23 320 11 m-NHCOC6H3-3-Me-4-SO2F 320 m-NHCOCeH4-(21-0 12 A 32 A 21 320 m-NHCOCeH4-Cl-m 13 310 rn-NHC0C~H3-2,4-C12 A 28 14 258 m-NHCONHC6H3-3-Me-4-SO~F B 57 16 220 m-NHCONHC6H3-2-~~e-5-so~F B 68 18 320 m-NHCONHC~H3-4-Cl-3-SOzF B 67 19 300 m-NHCONHC6Hz-2,4-Mez-5-SOzF 50 B 20 258-261 A 27 p - 0 (CHz)z-NHCOCsH3-2-Me-5-SOzF 21 237 p - 0 (CHz)z-NHCOC6H3-2-MeO-5-SOzF A 32 22 265 p-0 (CH~)Z-NHCOC~H~-~-C~-~-SOZF A 53 23 273-275 A 40 24 p - 0 (CHz)~-NHCOC~H~-2,4-Me~-5-So~F 207 B 36 26 p - 0 (CHz)z-NHCONHC6H3-2-Me-j-SozF 238-242 42 27 p - 0 (CHZ)Z-NHCONHCBH~-~-C~-~-SOZF B 225 B 34 28 p - 0 (CHz)z-NHCONHC~H~-2,4-Me~-5-So~F 24 1 B 47 30 p - 0 (CHz)zNHCONHC6H3-3-~Ie-4-S~zF 57 229 p - 0 (CHZ)ZNHCONHC~H~-~-C~-~-SOZFB 31 p - 0 (CHz )zNHCON HCHzCsH4-SOzF-p B 48 286 32d a For method A see B. R. Baker and W. F. Wood, J. Med. Chem., 11,650 (1968); for method B see ref 6; yields of anal. pure material, * All compds were recrystd from CH~OCHZCHZOH-HZO. All compds were analyzed for C, H, and N. See B. R. Baker and M. Cory, J. Med. Chem., 14, 805 (1971), for intermediate carbamate. 4 5 6
m-NHCOCeH3-2-Me-5-SOzF m-NHCOCeH3-2-Me0-5-SOzF m-NHCOCeH3-4-Cl-3-SOzF
A
A A
Irreversible Enzyme Inhibitors. 186."' Irreversible Inhibitors of the C'la Component of Complement Derived from rn-(Phenoxypropoxy)benzamidine by Bridging to a Terminal Sulfonyl Fluoride3 B. R. BAKER* AND MICHAEL CORY Departmat of Chemistry, University of California at Santa Barbara, Santa Barbara, California
93106
Received March 9, 1971 A series of 2 1 derivatives of m-(phenoxypropoxy)-, m-(phenoxybutoxy)-, m-(phenoxyethoxy)-, and m-(phenylbuty1)benzamidine bridged from the ortho position of the P h moiety to a terminal SOZFwere synthesized, then investigated as irreversible inhibitors of the C ' l a component of complement. The 2 most effective compds were m-[o-(2-chloro-5-fluorosulfonylphenylureido)phenoxybutoxy] benzamidine (25) and the corresponding propoxy compd (17) which showed 50% irreversible inhibition of C ' l a a t about 5 and 8 p M , respectively; these 2 compounds were also potent inhibitors of whole complement when assayed by inhibition of lysis of sheep red blood cells by hemolysin and complement.
The possible medicinal utility of inhibitors of serum complement for organ transplantation4 and in treatment of some arthritic states4 has been discussed previously. 8+5 The serum complement system involves 11 distinct proteins for killing invading organisms or for lysis of foreign mammalian cells.4 The most powerful (1) This work was generously supported by Grant CA-08695 from t h e National Cancer Institute, U.S. Public Health Service. (2) For t h e previous paper in this series see B. R. Baker and H.-U.Siebeneick, J . Med. Chem.. 14,802 (1971). (3) For the previous paper on complement see B. R. Baker and M. Cory, i b i d . , 14, 119 (1971). (4) H . J. Muller-Eberhard, Aduan.Immunol., 8, 1 (1968). ( 5 ) B. R. Baker and E . H. Erickson, J . Med. Chem., 12,408 (1969).
inhibitor of serum complement known to date3 is the benzamidine meta bridged to SOnF (1); however, 1 is not an irreversible inhibitor of the C'la component of ~ o m p l e m e n t . ~In contrast, the ortho-bridged SOzF
1, R
m-NHCONHCGH4SOZF-p
2, R = 0-NHCOCsHISOzF-m
806 Journal of Medicinal Chemistry, 1971, Vol. 14, No. 9
BAKER
AND
CORY
TABLE I INHIBITION OF WHOLECOMPLEMENT A N D IRREVERSIBLE INHIBITIOP O F ITSC‘la COMPONENT BY
Y
H~NCINH
I
NHCOR C’la,6
NO.
Inhib, mJ4
Yo inactvn
2e
0.25 0.125 0.062 0.031 0.125 0,062 0.5 0.25 0.125 0.12Y 0,062 0.031 0.50 0.2.5 0 , 12Fj 0.062 0.031 0.0621 0,062’ 0.031 0.015 0,0077 0.123 0.062 0.031 0.015 0.2.51 0.125 0.062 0.031 0.125, 0,062 0.1251 0.062 0.125J 0.062 0.031 0.125, 0.062 0.2.7 0.12.5 0.062 0.031 0.1251 0.062 0.031 0.0lc5 0.12df 0,062 0.031 0.015 0.0077 0.062’ 0.031 0,062J 0.031 0.062f 0.25, 0.125 0.062 0.125’ 0.062
89 84 70 29 16
3e
4c
7 8
9
10
11
12 13
14 15.
16
17
18 19 20 21 22
- 13 -8
98 81 41 9-5 81 56 27 9 92 89 76 42 90 79 39 18 97 93 75 47 81 51
$0 j1
73 39
40 1.i
-Whole corndement-% inhibn‘ % lysisd
80 64
9
38 23 90 83 32 51 R
6
82 91 60 36 15 63 23
10
0
80 3
2 0
X5
59 24
72 69 45
11 :3
66 44 70 46 29 31 25 52 34 61 57
0 0
33 2
0
21 9
51
100 99 87 49 95 96 95 78 41 30 0
28 92 70 43 71 43
26 6,5 70 34
12 0 0
74 80 60 32
11
33 36 43 31 38 74 54
9
69 51
0
0
2 0 5
Journal of Medicinal Chemistry, 1971, Vo2. 14, No. 9 807
IRREVERSIBLE ENZYME INHIBITORS. 186
TABLE I (Continued) C’la,” NO.
B
23
O(CHz)zO
R
Inhib, mM
% inactvn
--Whole complement-% inhibnC % ’ lysisd
PU”CBH~-~-CI-~-SOZF
0.125f 59 61 0 0.062 39 28 24 O(CHz)aO C6H4-4-Me-3-So2F 0.125f 48 14 0.062 24f 56 3 0.031 25 25 O(CHz)aO NHC6H3-2-Cl-5-SOzF 0.062J 100 86 4 0.031 100 77 0.015 92 50 0.0077 61 0.0039 39 26 (CHZ)~ NHC6H4S02F-m 0.125f 71 55 12 0.062 46 58 0 30 0.031 27 (CHZ)4 NHCsHaSO?F-p 0.125f 42 26 0.062 101 61 0.031 0 33 The technical assistance of Pauline Minton is acknowledged. Inhibitor incubated 10 min at 37“ with C’la, then remainder of complement added as previously described.3 Lysis of sheep red blood cells by guinea pig complement as previously described.6 d Lysis by the compd in the absence of complement expressed as per cent of total lysis possible.5 e Data from ref 3. J Maximum solubility. Q
2 is a powerful irreversible inhibitor of the C’la component of complement, but is only 0.125 as effective as 1 on the whole complement ~ y s t e m . ~Therefore a study n-as undertaken to see if the potency of the orthobridged sulfonyl fluoride 2 against whole complement or C’la or both could be increased by molecular manipulation of 2; the results are the subject of this paper. Biological Results.-The compds in Table I were compared by the concn necessary to give 50% inactivation when incubated with C’la at 37” for 10 min. The reasons why inactivation can be incomplete have been discussed previously. The first series of compds were derived from o-benzanilide by bridging to benzamidine with O(CHz)30. Of the 2 parent compds, the m-SOLF derivative 2 was considerably more effective as an irreversible inhibitor of C’la than the corresponding p-SO2F derivative 3 . 3 Introduction of a 2-AIe (6) substituent on the benzamido moiety of 2 decreased the effectiveness about 2fold; the (7) reduced activity even more. Activity was enhanced less than 2-fold by introduction of a 2-C1 substituent ( 5 ) . However introduction of 2,4-C12 substituents (8) increased activity 4-fold, 8 being the most active of the benzamide series; in contrast, introduction of 2,4-l\Ie2 substituents (9) did not change the activity. Insertion of a CH, group (10) next to the p-benzenesulfonyl fluoride moiety of 3 increased activity a t least %fold; this activity x a s maintained when CHzO (ll), (CH,), (12), or (CH?), (13) groups were inserted, but activity was not increased. The second series of compds were derived from an ophenylureido substituent (14). The parent m-SOtF derivative 16 was again more effective in inactivating C’la than the p-S02F derivative (15) by a factor of 16fold. Introduction of a 2-C1 atom (17) on 16 enhanced activity about 2-fold. Introduction of 4-C1 (18), 4-Me (19), or 2,4-Me? (20) on 16 considerably reduced the activity. Insertion of a CHz (21) or (CHS), (22) group next to the p-CeHlSOzF moiety of 15 enhanced activity 4-fold. The most active compd of this phenylureido series was the 2-C1 derivative 17 which showed 50% inhibition at about 8 p M . Therefore this 2-chloro-5-fluorosulfon-
ylphenylureido moiety of 17 was held constant while the length of the O(CH&O bridge was varied. The O(CH2)20 bridge (23) gave an 8-fold less effective compd while the O(CH2)40 bridge (25) increased the effectiveness by a factor of 2. Replacement of the ~ decreased effecO(CH2)30 bridge of 16 by ( C H Z )(26) tiveness 4-fold; this result can be attributed to the more restricted ground state conformation of the bridge (B) in 26 than in 16 which apparently restricts 26 from assuming easily the required conformation for maximum binding to the C’la component. The greater efficiency of 16 than of 26 on whole complement can also be rationalized in the same way. The 2 most active compds in Table I for irreversible inhibition of C’la are the 2-chloro-5-fluorosulfonylphenylureido derivatives, 25 and 17, which gave 50% inactivation at about 5 and 8 p M , resp. These 2 compds are also the most effective in Table I on whole complement; however, 25 is still only 0.5 as effective as l 3on the whole complement system. Since C1 is present in large excess in whole complement it is possible that 25 is not active enough to make C’la rate limiting in the whole system assay. Chemistry.-The intermediate substituted m-(phenoxya1koxy)benzonitriles 28 and 29 were prepared by the previously described alkylation of m-cyanophenol (method A);6 these were converted to the amidines 30 and 31 through the imino ether hydrochlorides (method B).’ Catalytic reduction of the NO2 group with 570 Pd/C (method C)’ gave crystalline aminoamidine salts 32 and 33 which could be acylated to the desired amides or ureas (methods D and E). The butadiene 34 was prepared by a Wittig reaction of m-cyanobenzyltriphenylphosphonium bromide5 and o-nitrocinnamaldehyde. Catalytic reduction of 34 with Pd/C catalyst (method C) and conversion of the crude oil to the amidine (method B) gave 35. The SOzF-substituted acyl chlorides and 0-(p-nitrophenyl) carbamates*used in methods D and E were prepared in these laboratories by published proce(6) B. R.Baker and E. H. Erickson, J . M e d . Chem., 10,1123 (1967). (7) B.R.Baker and E. H. Erickson, ibid., 11,245 (1968). (8) B. R.Baker and N . M . J. Vermeulen, ibid., 12, 74 (1969).
808 Journal of Medicinal Chemistry, 1971, Vol. 14,S o . 9
TABLE I1 PHYSICAL CONSTAKTS OF
Y Yield
5; 11p " C 20d 163-16.7 C(KHU)=NH 'TsOH N H C O C ~ H ~ ~ C I ~ S O L F ) 2Hd 148-131 C(SH,)=NH .TsOII 6 NHCOC6H3-2-hle-5-SOLF 1 Y' 222-224 C(?rTH,)=NH .TsOH 7 NHCOC6H3-2-hleO-3-SO~F 17' 196-195 C(h'Ha)=NH .TaOI1 x XHCOCsH*-2,4-Cl&-SO,F 221 214-21.5 C(NH2)=?JH .picrate 9 NHCOC6H2-2,4-~le*-j-SO?F G:ij 202-204 C(NH?)=NH .p']Crate 10 NHCOCHzC6HaSOzF-p 62f 189-191 C(NH?)=NH. TsOIT 11 NHCOCH2OC6H4SO2F-p 40' 20.5-207 C (NH?)=?;H .TsOH 12 NHCO(CH2),C6HaSO,F-p ,Xk 163-16.5 C ( N H ~ ) = X I l.TsOII 13 NHCO( CH2)aCeHnSOiF-p 4X"z 1%-18.5 C(NH,)=NH 'picrate 14 NHCONHCsH, 3,;' 193-197 C ( S H ? ) = S H . TsOH 16 NHCONHC~HISOZF-?TL 491 183-18.5 C(NHa)=NH .TsOII 17 NHCONHC6H3-2-Cl-MOiF 47f 148-1.50 C(NH?)=NH TsOJr 18 NHCONHC6Ha-4-CI-3-SO2F 5S" 137-159 C(NH,)=S€I TsOll 19 NHCONHC6H3-4-5Ie-3-SO2F .il ' 206-~20X 20 NHCOSHC6H1-2,4-~~e2-;,-SO>FC(NH,)=SH 'TsOlT ,j6i 17$1-lSl 21 N H C O K H C H ~ C ~ H ~ S O I F - ~ C(NHI)=SH .TsOII 17.' 202-20.5 TsOII -~ 22 N H C O N H ( C H ~ ) Z C ~ H ~ ~ O , FC(PI",)==KH 75> 212-213 C (NH, )=S H .pier a t o 23 NHCONHCsH3-2-CI-3-SO:F 19.' 193-19.5 C(NH2)=NH 'picrate 24 NHCOC6Ha-4-?\le-3-S02F :;3' 164-167 C(NHz)=NH 'TsOII 2
