J. Med. Chem. 1993,36, 3904-3909
3984
3,4-Dihydroxychalcones as Potent 5-Lipoxygenase and Cyclooxygenase Inhibitors Satoshi Sogawa, Yasunori Nihro, Hiroki Ueda, Akihiro Izumi,' Tokutaro Miki: Hitoshi Matsumoto, and Toshio Satoh' Faculty of Pharmaceutical Science, Tokushima Bunri University, Yamushiro-Cho, Tokushima, 770, Japan, and Drug Development Laboratories, Nippon Hypox Lab. Inc., 9420 Nanbu, Nanbu-Cho, Minumikoma-Gun, Yamanashi, 409-22,Japan Received April 22, 1993.
A novel series of 3,4-dihydroxychalcones was synthesized to evaluate their effects against 5-lipoxygenase and cyclooxygenase. Almost all compounds exhibited potent inhibitory effects on 5-lipoxygenase with antioxidative effects, and some also inhibited cyclooxygenase. The 2',5'disubstituted 3,4-dihydroxychalcones with hydroxy or alkoxy groups exhibited optimal inhibition of cyclooxygenase. We found that 2',5'-dimethoxy-3,4-dihydroxychalcone(37; HX-0836)inhibited cyclooxygenase to the same degree as flufenamic acid and 5-lipoxygenase, more than quercetin. Finally, these active inhibitors of 5-lipoxygenase inhibited arachidonic acid-induced mouse ear edema more than phenidone.
Introduction The leukotrienes are important mediators of smooth muscle constriction,' increased vascular permeability: and leukocyte ~hemotaxis.~ The enzyme 5-lipoxygenasecatalyzes the initial step in the metabolism of arachidonic acid leading to leukotrienes. Inhibition of the enzyme activity has provided a new therapeutic approach to treating a variety of inflammatory and allergic diseases. Especially, 5-lipoxygenaseinhibitors have emerged as an attractive approach to treatment of inflammatory skin diseases, such as psoriasis and contact dermatitis, since these inflammatory dermatoses are not significantly improved by nonsteroidal antiinflammatory drugs. Leukotrienea are elevated in the skin of patienta with psoriasis and atopic eczema, whereas prostaglandins are not significantly e l e ~ a t e d . ~ Four mechanisms can be considered for 5-lipoxygenase (1)antioxidant and/or free radical scavenging, inhibiti~n:~ (2) iron chelation, (3) the inhibition of 5-lipoxygenase translocation, and (4) substrate mimicking. In particular, several antioxidant 5-lipoxygenase inhibitors (e.g., DuP 654: lonapalene? and R-68,1518)have been shown to be clinicallyeffective againsttopical inflammatory conditions. Numerous studies have been undertaken on the 5 4 poxygenaseinhibitors. The current status of the research has been reviewed in Journal of Medicinal Chemistry and Progress in Medicinal C h e m i ~ t r y . ~Some . ~ reports have pointed out toxicological problems associated with nonspecific antioxidants, such as methemoglobinemia.'0 However, clinical trials of lonapalene have demonstrated no side effectsto preclude ita administration.' It is possible that the efficacyof the antioxidant 5-lipoxygenaseinhibitor is independent of its toxicity. For example,5-lipoxygenase inhibitors readily metabolizedby systemicadministration would be suitable topical antiinflammatory agents. Nakadate et al. have reported that known hydroxychalcones inhibit 124ipoxygenase and cyclooxygenase in the mouse epidermis.l' Some chalcone derivatives have been reported as antiinflammatoryor antiallergicagents.12 Futhermore, we found that chalcones with a 3,4-dihydroxycinnamoyl structure strongly inhibited lipid peroxidation in rat liver micros0mes.~3 The 3,4-dihydroxy+ Nippon Hypos Lab. Inc. e
Abstract published in Advance ACS Abstracts, October 15, 1993.
chalcones are rapidly and extensively metabolized after systemicadministration. These findingssuggestthat some chalcones may be promising nontoxic topical antiinflammatory agents. In this paper, we describe the biological activity of various hydroxychalcones against 5-lipoxygenase and cyclooxygenase using in vitro and in vivo topical inflammatory models and discusstheir structure-activity relationships.
Results Chemistry. We synthesized3,4-dihydroxychalcone(8) using the Claisen-Schmidt condensationof acetophenone with a unprotected 3,4-dihydroxybenzaldehyde. However, this procedure afforded compound 8 in a low yield (below 10%1. The 3,4-dihydroxybenzaldehydeprotected as the bis(tetrahydropyrany1) ether was condensed with acetophenone to give compound 8 in a good yield (45%). Previously, the benzyl and methoxymethyl ethers of the phenolic hydroxy group were used in the Claisen-Schmidt condensation,14althoughthese ethers required purification before the condensation step. However, the purification procedure was not necessary when using tetrahydropyranyl ethers to yield the desired hydroxychalcones. The chemical structures and yields of the entitled chalcones (1-63) are listed in Tables 1-111. Known chalcones (1-9, 11-13, 16,19, and 41) are presented with supplementaryreference numbers in Tables I and II.12bJ5 Biological Evaluation. We investigated the effects of hydroxychalcones (1-13) and related compounds (1419) on 5-lipoxygenase,cyclooxygenase, and lipid peroxidation. Those which had a 3,4-dihydroxycinnamoyl moiety inhibited 5-lipoxygenase and lipid peroxidation (Table I). We found that 3,4-dihydroxychalcone 8 and 2',3,4-trihydroxychalcone 9 also inhibited cyclooxygenase. However, these activities were less effective than those against 5-lipoxygenase(cf. ICs0 values of compound 8: 34 and 0.043rM against cyclooxygenaseand 5-lipoxygenase, respectively). We selected compound 8 for the lead compound as a dual inhibitor against cyclooxygenaseand 5-lipoxygenase. Therefore we synthesized monosubstituted 3,4-dihydroxychalcones (20-31) which bear a substitution group at the 2/-3'-, or 4'-position (Table11).Among these monosubstituted 3,4-dihydroxychalcones (2&51), 4'-chloro, 3/-methoxy,or 3'- or 4'-(dimethylamino) groups were effective against 5-lipoxygenase, whereas 2'- or 3'-
0022-262319311836-3904$04.00/0 0 1993 American Chemical Society
Journal of Medicinal Chembtry, 1993, Vol. 36, No.24 8901
Potent 5-Lipoxygenuse and Cyclooxygenase Inhibitors
Scheme I. Synthesis of 4',3,4-Trihydroxychalcone (11)" " q C H S
H f l Z 0
0
11.
llb
li q
C
H
+
,
0
0
lle
"(i) 3,CDihydro-W-pyran
ii
0 lle
lld
+ pyridinium ptoluenesulfonate;
11
(ii) Ba(0H)pHZO.
Table I. Inhibition of CLipoxygenase, Cyclooxygenase, and Lipid Peroxidation by Hydroxychalcones and Related Compounds R' ' g3
R 2
0 ~~~
RLM LPOXC yield ( % ) mp ("C) cryetn solvent % inhibn at 1pM 65 82-85 MeOH/H20 6 171-174 MeOH/HzO 86 5 145-146 MeOH/H20 74 5 178-180 MeOHIHZO 78 9 158-160 MeOH/H20 4-OH 75 6 4-OH 76 215-219 MeOH/HzO 9 40H 70 200-201 MeOH/H20 8 202-204 MeOH/HzO 81* 3,4OH 45 44 9'b 2'-OH 3,40H 66 178-180 MeOH/H20 45 191-192 MeOH/H20 10 3'-OH 3,4OH 48 24 lllsb 4'-OH 3,4OH 67 218-219 MeOH/H20 36 250-253 MeOH/HaO 12l" 2',4'-OH 3,4OH 64 40 15'' 249-252 MeOH/Hfl 2',4',6'-OH 3,40H 63 18 14 183-185 benzene 2-thienyl 3,4OH 24 56 242-250 EtOH/HzO 3-pyridyl 3,4OH 57 10 16 16l'j 2'-OH 126-127 MeOH/HpO 3-OCHa,4OH 37 14 94-101 MeOH/H20 17 4/41 3-OCH3,dOH 57 not tasted 18 156156 MeOH/H20 4'-OCHa 3-OCHa,4OH 49 2.3 19'" 2'-OH 153-156 MeOH 3-OH,4-OCHa 39 11 CLipoxygenaw from RBL-1 cella. Sheep seminal vesicle cyclooxygenasa. e Lipid peroxidation in rat liver microsomes. ICWbaaed on duplicate three-point titration. Values in parentheses are percent inhibition at the concentration shown (pM) where ICWvalues were not determined.
R' 2'-OH 4'-OH 2',4'-OH 2',4',6'-OH SI6. 2'-OH 61b 2',4'-OH 7IM 2',4',6'-OH
no. 11b 2'6b 3'" 41M
R
*
methoxy groups had better anti-cyclooxygenaseactivity. The anti-cyclooxygenase activity waa reduced by 4'substitution. We investigated disubstituted chalmnes (32-39) which bear hydroxy, methyl, and/or methoxy groups at the 2',4'-, 2',5'-, 2',6'-, or 3',4'-positions. Additionally, we assessed the biological evaluations of trisubstituted chalcones (40 and 41). We found Z',S'-dimethoxy-3,ddihyhydro.ychalcone (37) waa aa effective a dual inhibitor as DuP 654 (ICs0 values of 7.8 nM and 9.2 wM, against S-lipoxygenaseand
cyclooxygenase, respectively). Furthermore, we assayed the 3,4-dihydroxychalcones(42-53) bearing disubstitution groups at the 2',6'-position in vitro and in vivo. These chalcones (42-63), except for compound 47, exhibited potent inhibitory effect on S-lipoxygenase. Especially, compounds 44,46, 50, and 63 inhibited S-lipoxygenaaeto a greater degree than compound 37 (ICs0 values of compounds 44, 46, 60, and 63: 5.3, 4.0, 3.8, and 2.4 nM against 5-lipoxygenase,respectively). In the series of theae anti-cyclooxygenaeeactivities, compound 62 was the most
Sogawa et
3906 Journal of Medicinal Chemistry, 1993, Vol. 36,No. 24
41.
Table 11. Inhibition of 5-Lipoxygenaseand Cyclooxygenaea by 3,4-Dihydroxychalconee
0 RBL-1 &LO SSV CO crystm solvent formula mp (“C) no. ICW(nM)a ICs0 bM)a R yield (7%) 202-204 43 34 CldhO3 45 MeOH/H20 8 H 20 2’41 58 174-178 benzene ci~&oscl 92 71 58 228-230 8.5 1400 Ci~.hOaCl 21 4’- C1 acetone/HzO 16 268-269 CisHiiOsN benzene/EtOAc 23 5%b 22 4’-No2 58 230 cl~llo83 63 178-180 MeOH/H20 23 2’-CF3 27 71 C1&403 43 163-164 benzene 24 3‘-CH3 35 201-202 76 210 Cl&403 MeOH/H20 25 4’-CH3 33 146147 27 13 Cl&1404 benzene 26 2’-OCH3 52 152-153 6.5 15 C1&404 benzene/acetone 27 3’-OC& 43 172-178 20 490 Cl&404 MeOH/H20 28 4’-OCH3 49 145-149 9.8 41 Ci7Hi703N benzene 29 3’-N(CHs)z 4.7 810 Ci7Hi703N 39 211-213 acetone/H20 30 4’-N(CH3)2 41 ll%* 42 160-166 CleHleO4 31 4’-OCH(CH3)2 benzene/acetone 15 66 Cl&406 32 2’-OH,4’-OCHa 46 170-173 benzene 41 41 63 149-151 Cl6&4OS 33 2’-OH,5’-OCH3 MeOH/HaO 44 174-177 Cld&4OS 34 4’-OH, 3’-OCH3 9.0 14% benzene 17 41 33 131-134 C17Hle03 35 2’-CH3,4’-CH3 benzene/n-hexane 51 160-161 10 160 C17Hl6OS 36 2’-OCH3,4’-OCH3 EtOH/HaO 53 158-160 7.8 9.2 C17HreOs 37 2’-OCH3,5’-OCH3 benzene/acetone 22 192-194 370 15%b C17H1606 38 2’-OCH3,6’-OCH3 benzeneln-hexane 55 132-137 18 140 Cl7HleO5 39 3’-OCH3, 4’-OCH3 bewne/acetone 40 2’-CH3, 4’-CH3, 6’-CH3 27 176176 400 280 CleH1604 benzeneh-hexane 4112b 3’-OCH3, 4’-OCH3, 5’-OCH3 48 153-154 benzene Cl~l8O6 16 13%b a ICWbased on duplicate three-point titration. Values are percent inhibition versus control at 100 wM,average of two determinatiom. Table 111. Inhibition of 5-Lipoxygenaee, Cyclooxygenase, and Arachidonic Acid-Induced Mouse Ear Edema by 2’,5’-Dieubetituted 3,4-Dihydroxychalcones
Rp
0
RBL-1 &LO ssvco AAearedem8 yield (%) mp (“C) crystn solvent formula ICW(nM)a ICs0 bM)a % inhibition’ 7.8 (4.5-14) 53 158-160 benzene/acetone 9.2 (6.8-13) 77 9.6’. 64 (39-110) 170 (100-280) 61 204-205 benzene/acetone 39 f 3.42 39 (24-61) 62 7.6** 44 177-183 MeOH/H20 120 (69-230) 41 (21-79) 41 (34-51) 63 38 f 3.8* 149-151 MeOH/H20 5.3 (3.1-9.2) 47 42 6.8* 130 (100-160) 167-169 benzene/acetone 4.0 (0.90-17) 16 f 3.6 37 157-158 benzene 37 (31-46) 11 (6.2-21) 67 8.S** 177-178 benzene 140 (80-230) 33 4.2%“ 53 f 4.6 47 162-163 benzene/acetone lo00 (300-3600) 16 (7.1-37) 68 7.7. 154-155 benzene 52 44 (36-54) 24 (1149) 21 30 (24-37) 47 f 11.9* 150-151 benzene/EtOAc 3.8 (2.2-6.7) 116122 benzene 49 26 (21-32) 50 13 3.6 14 (8.7-21) 51 42-43 60 (48-76) 34 9.2* 29 not recrystallized 27 (13-63) 60 52 2.0 (1.4-2.9) 7.4 2.2 122-125 benzene/acetone 2.4 (1.24.6) 53 53 -18 19.4 24 (20-29) 153-155 benzene a ICWbased on duplicate three-point titration; 95% confidence liiite are in parenthesea. Mean SE value (n = 4-8) at 30 &ear. Significautlydifferentfromcontrol,( * ) p
