Applications of quantitative structure-activity relationships to drug design of piperazine derivatives

QSAR and Drug Design - New Developments and Applications T. Fujita, editor 9 1995 Elsevier Science B.V. All rights reserved

413

A P P L I C A T I O N S OF QUANTITATIVE S T R U C T U R E - A C T I V I T Y R E L A T I O N S H I P S TO D R U G D E S I G N OF PIPERAZINE DERIVATIVES Hiroshi OHTAKA

New Drug Research Laboratories, Kanebo Ltd., 1-5-90 Tomobuchi-cho, Miyakojima-ku, Osaka 534, J a p a n ABSTRACT: Applications of quantitative structure-activity relationship (QSAR) procedures to our own practical drug research are reviewed. A benzylpiperazine cerebral vasodilator (KB-2796) and a piperazine-acetate antiulcer agent (KB-5492) were successfully optimized by use of QSAR information. In these cases, appropriate strategies of the synthetic research were devised and QSAR analyses were performed repeatedly in each step during the research. In the last example, the selection of a 2-homopiperazinylbenzimidazole (KG-2413) as a candidate for antihistaminics was confirmed to be valid by QSAR analysis after the synthetic research project was over. The antiulcer and cerebral vasodilative agents are now under extensive clinical trials, while the antihistaminic agent has been used clinically. 1.

INTRODUCTION

A considerable number of 1,4-disubstituted piperazines have been found to possess interesting pharmacological properties and some of them have been used clinically. Syntheses of novel 1,4-disubstituted piperazine derivatives have been performed with the aim of finding new drugs. Piperazine is still a very important starting material in the pharmaceutical industry. The drug research and development project involves a number of steps and requires exhaustive effort and considerable expenditure. Medicinal chemists contribute mainly to the earliest stages of the project wherein new drug candidates are synthesized and selected. Such chemists must not only design novel chemical structures and perform syntheses, but also arrange the huge amount of structureactivity data in a meaningful order. However, since the majority of

414 newly synthesized compounds are abandoned in the earlier stages of the project, chemists are eager to discover or apply more efficient drug design methods than ever which reduce trial and error to a minimum. Quantitative structure-activity relationship (QSAR) analysis introduced by Hansch and Fujita (1) about 30 years ago, is still used widely as an effective and rational drug design approach. In this approach, the potency variations in a series of bioactive compounds are considered to be determined by several physicochemical properties. The q u a n t i t a t i v e relationship is analyzed, for example, by use of the following equation, log 1/C = kl~ + k2a + k3E s + .... + const.

[1]

where C is the equieffective dose or concentration, which should be properly expressed on a molar basis, and ~, ~ and E s represent hydrophobic, electronic and steric factors, respectively. When the activity of a compound is represented by potency rating (++, +, -, etc.), one of the modifications of the Hansch method, an adaptive least squares (ALS) method developed by Moriguchi and co-workers (2), is t h o u g h t to be appropriate. There have been m a n y successful applications of these QSAR methods to practical problems (3-7). In this article, our own examples in designing three types of drugs derivatized from piperazine are reviewed. 2. BENZYLPIPERAZINE CEREBRAL VASODH ATOR Cerebrovascular disorders were the most common cause of death in J a p a n up to 1981. Although the mortality attributable to these disorders has been decreasing recently, the number of patients showing after-effects has been increasing. Cerebrovascular disorders can roughly be classified into i n t r a c r a n i a l hemorrhage and cerebral infarction. In these disorders, infarction of the brain parenchyma occurs by h e m o r r h a g e , t h r o m b u s or embolus, leading to an insufficiency of glucose and oxygen, which supply the energy necessary for n e u r o n a l activity. This results in functional and organic disturbances in the ischemic area. Accordingly, drugs which facilitate

415

the supply of glucose or oxygen to the ischemic area by increasing the cerebral blood flow are effective for the t r e a t m e n t and prevention of these disorders. The potency, duration of action and cerebrovascular specificity are considered to be i m p o r t a n t properties for cerebral vasodilators (8). From these points of view, however, such c u r r e n t drugs as papaverine hydrochloride (9) and cinnarizine (10), are not necessarily satisfactory. 2 . 1 Design of the Possible ~ d S t r u c ~ In the course of our search for novel cerebral vasodilators, we have noticed publications showing t h a t 1-(2,3,4-trimethoxybenzyl)piperazine (I: trimetazidine) dihydrochloride, a coronary vasodilator, is not only distributed to the brain and to the heart of mice (11) but also relaxes dog basilar arteries more effectively t h a n coronary arteries after contraction with prostaglandin F 2 a (12). Trimetazidine is one of a few monosubstituted piperazines used clinically and seems to be much more hydrophilic t h a n cinnarizine (II). Therefore, we t h o u g h t it appropriate to modify the structure of trimetazidine to more lipophilic 1,4-disubstituted piperazine derivatives with the aim of finding a new cerebral vasodilator, and designed c i n n a m y l - ( I I I a ) and diphenylmethyl-trimetazidines (1Va).

OMe

I: Trimetazidine (CoronaryVasodilator)

I1: Cinnarizine Q (CerebralVasodilator)

I

MeO

OMe

Ilia

e ~

416

Cinnamyl-trimetazidine and Related Compounds A series of 1-benzyl-4-cinnamylpiperazine derivatives (III) was prepared and tested for cerebral vasodilative potency, which was evaluated from the response in dogs anesthetized with pentobarbital. The response was measured in terms of the ratio of the maximum change in blood flow in vertebral a r t e r i e s after i n t r a v e n o u s administration of the test compound (1 m g / k g ) t o that produced by papaverine (1 mg/kg). Although compounds III showed negligible activity, the synthetic intermediates (V) exhibited considerable activity (13). Qualitative analysis according to the Topliss scheme (14) suggested that the activity was positively dependent on the lipophilicity of the substituent X. Further analogs of V were therefore synthesized and this tendency was confirmed. Then, in order to make the molecule more lipophilic and to study the effects of the number and location of methoxy groups, the derivatives of V represented by the general formula (VI)were prepared and tested (15). 2.2

~ . . N ~ ~x OMe

OH V

OH Vl

For the QSAR analysis, the potency of compounds should be appropriately expressed on the molar basis. In this research, the response was observed to increase almost linearly with the log(dose) value within a certain range of concentrations where the response was from about 1/2 to 3/2 of t h a t of papaverine (1 mg/kg) for four derivatives. The log(dose)-response relationships for the four derivatives were almost parallel, the slope being estimated as 1.25 + 0.01 (n = 4), taking the log(dose) as the independent variable. For the rest of compounds, the response was evaluated at a single dose (1 mg/kg). Therefore, the log(dose)-response relationships for these compounds were assumed to have identical slopes. This assumption can be expressed by Eq. 2.

417 response = 1.25 log(dose) + C

[2]

Equation 2 can be rewritten as Eq. 2' by converting the log(dose) into the dependent variable. log(dose) = 0.8 response + C'

[2']

In these equations, C and C' are intercepts with the respective ordinates. The biological activity index of interest for structure-activity analysis is the negative l o g a r i t h m of the dose, D, required to produce a given response, and so the "response" in Eq. 2' is set as a certain value. At the given response, Alog D - AC' among derivatives. Since AC' - Alog(dose) 0.8 Aresponse, where the "response" is not set as a c o n s t a n t but variable and t h a t induced by the dose applied, the biological activity in terms of log(l/D) of a derivative A is represented as Eq. 3 where S stands -

for a reference compound. log(1/D)A - log(1/D)s + log(dose) s - log(dose)A 0.8 (response s - response A) -

[3]

C o m p o u n d Vie was t a k e n as a reference, since the dose (mol/kg) r e q u i r e d for 100% increase in blood flow in v e r t e b r a l a r t e r i e s was m e a s u r e d so t h a t log(l/D) S = 6.30. By converting the log(dose) value (dose = 1 mg/kg) into the log value on a molar basis for each compound and introducing the "response" values in terms of the ratio to t h a t of p a p a v e r i n e (1 mg/kg) into Eq. 3, the log(l/D) v a l u e s for the 35 compounds listed in Table 1 were estimated. The D (mol/kg) value is t h a t required for 100% increase in blood flow for each compound and is a p p r o x i m a t e l y equivalent with t h a t inducing 2/3 of the effect of the s t a n d a r d compound, papaverine (1 mg/kg). QSAR analysis for these derivatives was performed to yield Eq. 4 (16), log(l/D) - - 0.686(+0.129)~R 2 + 1.361(+0.274)~ R + 0.342(+0.128)~ x [4] + 0.288(+0.091 )Im + 0.207(+0.171 )Ip + 4.292 n-35, r-0.934, s-0.132, F=39.70

418 TABLE 1. S t r u c t u r a l F e a t u r e s a n d Cerebral Vasodilative Activities of 1-Benzyl-4(3-hydroxy-3-phenylpropyl)piperazine Dihydrochlorides (V a n d VI)

Compd.

log(l/D)

No.

Y

R

X

Ip

Im

~R

XX

Obs.

Calcd. (Eq.4)

Va

2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3

H H H

H 4-Me 4-C1

1 1 1

3 3 3

0.00 0.00 0.00

0.00 0.56 0.71

5.37 5.51 5.66

H H H Me Me Me Me Me

3,4-C12 3,4-Me 2 3,4-(CH) 4 H 4-Me 4-OMe 4-C1 3,4-C12

1 1 1 1 1 1 1 1

3 3 3 3 3 3 3 3

0.00 0.00 0.00 0.50 0.50 0.50 0.50 0.50

1.25 0.99 1.32 0.00 0.56 0.56 0.71 1.25

5.84 5.67 0.59 5.87 6.03 6.03 6.06 6.30

5.3C~ 5.56 5.61 5.79 5.70 5.82

VIk VI1 Vim VIn VIo VIp VIq

2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(0Me) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3 3,4,5-(OMe) 3 2,4,6-(OMe) 3 2,4-(OMe) 2 3,4-(OMe) 2 3,5-(OMe) 2

Me Me Et Et Pr Bu Bz Me Me Me Me Me

3,4-(CH) 4 2,4-C12 3,4-C12 2,4-C12 3,4-C12 3,4-C12 3,4-C12 3,4-C12 3,4-C12 3,4-C12 3,4-C12 3,4-C12

1 1 1 1 1 1 1 1 1 1 1 0

3 3 3 3 3 3 3 3 3 2 2 2

0.50 0.50 1.00 1.00 1.50 2.00 2.01 0.50 0.50 0.50 0.50 0.50

1.32 1.42 1.25 1.42 1.25 1.25 1.25 1.25 1.25 1.25 1.25 1.25

6.15 6.38 6.64 6.54 6.33 5.91 5.74 6.47 6.22 6.26 6.06 5.51

6.32 6.36 6.47 6.53 6.29 5.77 5.76 6.30 6.30 6.01 6.01

VIr

2,3-(OMe) 2

Me

3,4-C12

0

2

0.50

1.25

5.75

5.81

VIs

2-OMe

Me

3,4-C12

0

1

0.50

1.25

5.55

5.52

VIt

4-OMe

Me

3,4-C12

1

1

0.50

1.25

5.61

5.72

VIu

3,4,5-(OMe) 3

Me

2,4-C12

1

3

0.50

1.42

6.34

6.36

VIv

3,4,5-(OMe) 3

Et

3,4-C12

1

3

1.00

1.25

6.50

6.47

VIw VIx

3,4,5(OMe) 3 3,4,5-(OMe) 3

Et Pr

2,4-C12 3,4-C12

1 1

3 3

1.00 1.50

1.42 1.25

6.30 6.37

6.53 6.29

Vb Vc Vd Ve Vf Via VIb a

2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3 VIc a 2,3,4-(OMe) 3 V I d a 2,3,4-(OMe) 3 Vie 2,3,4-(OMe) 3 VIf 2,3,4-(OMe) 3 VIg VIh Vii

vij

5.87 6.07 6.07 6.12 6.30

5.81

419 TABLE 1. C o n t i n u e d

Compd. No.

log(l/D) Y

VIy VIz

3,4,5-(OMe)3 3,4,5-(OMe)3 V I a a 3,4,5-(OMe)3 Vlbb 3,5-(OMe)2 Vice 4-OMe

R

X

Ip

Im

XR

XX

Pr Bu Bu Et Et

2,4-C12 3,4-C12 2,4-C12 3,4-C12 3,4-C12

1 1 1 0 1

3 3 3 2 1

1.50 2.00 2.00 1.00 1.00

1.42 1.25 1.42 1.25 1.25

Obs. Calcd. (Eq.4) 6.20 5.64 5.81 6.29 5.84

6.35 5.77 5.83 5.97 5.89

a) Dimaleate.

where the figures in parentheses are the 95% confidence intervals, n is the n u m b e r of data points used in deriving the equation, r is the correlation coefficient, s is the s t a n d a r d deviation, and F is the ratio between regression and residual variances. In Eq. 4, ~X is the lipophilicity of substituent X of the phenyl moiety, ~R is the lipophilicity of substituent R at the benzylic position, I m represents the number of methoxy groups on the benzyl moiety and Ip is an indicator variable for the presence (Ip = 1) or absence (Ip - 0) of the para-methoxy group. F r o m Eq. 4, the optimum ~R was calculated to be 0.993 and an ethyl group was confirmed as being the best R substituent. Many kinds of drug activity have been found to depend upon lipophilicity, which is one of the most f u n d a m e n t a l c h a r a c t e r i s t i c s of d r u g s t r u c t u r e s determining biological activity. In the present case, Eq. 4 indicates that the local lipophilicity around the a s y m m e t r i c carbon atom is of importance. This may be the case, because intravenous administration of drugs does not involve the absorption process or first-pass effect, and so the interaction of drugs with the active site is the most critical step. The most favorable s u b s t i t u t i o n p a t t e r n of the benzyl moiety is suggested to be 2,3,4,5,6-pentamethoxy, but this is not practical. E q u a t i o n 4 also suggests t h a t the introduction of more lipophilic s u b s t i t u e n t s for the group X would make the compound more active. The most active of those prepared, however, exhibited no cerebral vasodilative activity when administered intraduodenally, although they

420

were more potent t h a n cinnarizine when applied intravenously. Therefore, our search for new cerebral vasodilators in this series was terminated. Diphenylmethyl-trimetazidine a n d Related C o m p o u n d s Unlike the series of compounds described in the preceding section, d i p h e n y l m e t h y l - t r i m e t a z i d i n e (IVa) showed considerable cerebral vasodilative activity even on intraduodenal application. This compound has been described in the patent literature but nothing about its medical utility has been disclosed (17). We considered this compound to be a possible lead for new cerebral vasodilators and therefore a series of 1benzyl-4-diphenylmethylpiperazines ( I V ) w e r e p r e p a r e d and tested (18). 2.3

z

Most compounds in this series were also administered at a certain single dose, but the log(dose)-response plots of potent derivatives were examined and found to be parallel. Accordingly, the potency in terms of the log(l/D) value was estimated in a manner similar to that used in the preceding section. QSAR analyses were performed for each subseries where Y = Z = H ( 1 V a - 1Vm), Y - Z - F (Wp, l V x - lVff) and X - 2,3,4-(OMe) 3 (lVa, l V p - lVw)(19). The two compounds (lVn and 1Vo) showed much lower activity t h a n expected. They also caused acute toxicity in addition to showing activity. Since data on these compounds were thought to distort the correlation, they were not included in the analyses. Then, 35 compounds in Table 2 were subjected to analysis leading to Eq. 5, log(l/D) = - 0.839(+0.258)Z~ - 0.075(+0.038)MR + 5.582 n = 3 5 , r=0.788, s=0.231, F=26.18

[5]

421

TABLE 2. S t r u c t u r a l F e a t u r e s a n d C e r e b r a l Vasodilative Activities of 1-Benzyl-4d i p h e n y l m e t h y l p i p e r a z i n e D i h y d r o c h l o r i d e s (IV)

Compd. No.

log(l/D) X

Y

Z

Z~

MR

Obs.

Calcd. (Eq.5)

Duration Obs.Calcd. (Eq.6)

IVa

2,3,4-(OMe) 3

H

H

-0.42

1.03

6.23

5.86

0

1

IVb a

4-OAc

H

H

0.31

1.03

5.45

5.25

1

0

IVc

4-C1

H

H

0.23

1.03

5.34

5.31

0

0

IVd a

4-F

H

H

0.06

1.03

5.40

5.46

0

0

IVe ivf b

H

H

H

0.00

1.03

5.39

5.51

0

0

4-NHAc

H

H

0.00

1.03

5.44

5.51

0

0

IVg

3,4,5-(OMe) 3

H

H

-0.03

1.03

5.48

5.53

0

0

IVh

3,4-(OMe) 2

H

H

-0.15

1.03

5.43

5.63

0

0

IVi

4-Me

H

H

-0.17

1.03

5.70

5.65

1

0

ivj

3,4-OCH20-

H

H

-0.32

1.03

5.58

5.77

0

0

IVk

4-OH

H

H

-0.37

1.03

5.92

5.82

0

0

IVl a

2,4-(OMe) 2

H

H

-0.54

1.03

5.97

5.96

1

1

IVm a

2,4,6-(OMe) 3

H

H

-0.81

1.03

6.58

6.19

0

d

IVn a

4-NH 2

H

H

-0.66

1.03

5.70

c

0

1

IVo a

4-NMe 2

H

H

-0.83

1.03

5.55

c

1

1

IVp

2,3,4-(OMe) 3

F

F

-0.42

0.92

6.32

5.87

0

0

IVq

2,3,4-(OMe) 3

Me

Me

-0.42

5.65

6.08

5.51

0

0

IVr

2,3,4-(OMe) 3

C1

C1

-0.42

6.03

5.57

5.48

0

0

IVs a

2,3,4-(OMe) 3

OMe OMe

-0.42

7.87

5.22

5.35

0

0

IVt a

2,3,4-(OMe) 3

F

H

-0.42

1.03

6.08

5.86

1

1

IVu a

2,3,4-(OMe) 3

Me

H

-0.42

5.65

5.32

5.51

0

0

IVv a

2,3,4-(OMe) 3

C1

H

-0.42

6.03

5.64

5.48

1

0

IVw a

2,3,4-(OMe) 3

OMe H

-0.42

7.87

5.04

5.33

0

0

IVx

3,4,5-(OMe) 3

F

-0.03

0.92

5.47

5.54

0

0

IVy

2,4,6-(OMe) 3

F

F

-0.81

0.92

6.09

6.19

0

d

IVz

2,4-(OMe) 2

F

F

-0.32

0.92

5.57

5.78

0

0

IVaa

3,4-OCH20-

F

F

-0.32

0.92

5.57

5.78

0

0

lVbb

4-OH

F

F

-0.37

0.92

5.70

5.82

1

1

IVcc

H

F

F

0.00

0.92

5.37

5.51

0

0

IVdd

4-Me

F

F

-0.15

0.92

5.41

5.66

1

0

IVee a

4-NMe 2

F

F

-0.83

0.92

6.22

6.21

1

1

lVff a

4-OAc

F

F

0.31

0.92

5.51

5.25

0

0

IVgg a

2,4,6-(OMe) 3

F

H

-0.81

1.03

5.91

6.19

1

d

F

422

TABLE 2. C o n t i n u e d

Compd. No.

IVhh a IVii IVjj IVkk a

X 2,4-(OMe)2 3,4,5-(OMe) 3 3,4-OCH204-NMe 2

Y

Z

Zo

F F F F

H H H H

-0.54 -0.03 -0.32 -0.83

log(l/D) Duration MR Obs. Calcd. Obs.Calcd. (Eq.5) (Eq.6) 1.03 1.03 1.03 1.03

5.95 5.52 5.38 6.04

5.96 5.53 5.77 6.20

1 0 0 1

1 0 0 1

a) Fumarate. b) Maleate. c) Not included in the correlation, d) Not included in the analysis.

where Z(~ represents the electronic effect of substituent X on the benzyl moiety and MR r e p r e s e n t s the molar refractivity value for the larger s u b s t i t u e n t of Y and Z. The correlation coefficient of Eq. 5 is not so high as one would like. However, the correlation is highly significant and is not inconsistent with those for the above mentioned subseries where much b e t t e r correlations were observed. Equation 5 suggests t h a t the e l e c t r o n - d o n a t i n g effect of the s u b s t i t u e n t on the benzyl moiety is i m p o r t a n t for the p o t e n t cerebral vasodilative activity. Since the coefficients of Zc are close to unity, protonation at the benzylic nitrogen atom seems to play a significant role. Although lipophilicity itself is a very i m p o r t a n t factor for these c o m p o u n d s to e x e r t a c t i v i t y as e x p e c t e d for t h e h y d r o p h o b i c trimetazidines, the term for the lipophilicity was not significant in Eq. 5. This may be t a k e n to indicate t h a t the molecular lipophilicity of these compounds is sufficiently high and t h a t r a t h e r small differences in lipophilicity are not critical in governing the variations in activity. The electronic and steric interactions of the drugs with the active site is more critical. B u l k y s u b s t i t u e n t s at t h e para-position of the diphenylmethyl moiety are not favorable for potency. Aromatic fluorine is the s m a l l e s t group in t e r m s of MR, and t h u s fluorine is the best substituent for both Y and Z. Next, the r e l a t i o n s h i p b e t w e e n the s t r u c t u r e and d u r a t i o n of action was examined. Compounds were judged to be long-acting when their durations of action were greater t h a n 20 times t h a t of papaverine.

423

From the pharmacological data, compounds of higher potency also seemed to be long-acting. ALS analysis for the duration of action with Z(~ and MR appearing in Eq. 6, gave a significant result omitting three 2,4,6-trimethoxybenzyl derivatives (IVm, IVy and lVgg). Y = - 1.774 Za - 0.128 MR - 0.302 n = 3 4 , Rs=0.620, nmi s - 6 , t - 4 . 4 7 ,

[6] p < 0.001

In Eq. 6, Y is the rating score taking either unity or zero depending upon the judgment for long-acting or not. The Y value ultimately calculated by Eq. 6 was categorized into integers corresponding to the rating score according to certain rules (2). In Eq. 6, n represents the number of compounds used to derive the equation, nmi s is the number of compounds where observed and calculated rating scores do not match, Rs is the Spearman rank correlation coefficient, t is Student's t value calculated by t - Rs[(n - 2)/(1 - Rs2)] 1/2, and p is the level of significance. One possible justification for the exclusion of the above three compounds is that their conformation may be different from the others because of their di-ortho-substitution on the benzyl moiety. The duration of action of drugs depends upon various factors such as its elimination, distribution, metabolic transformation, and, in some cases, the biological activity of the metabolites. However, Eqs. 5 and 6 suggest t h a t introduction of electron-donating substituents at the benzyl-benzene ring and sterically small substituents at the paraposition on the diphenylmethyl moiety bring about strong interaction of the compounds with the active site, resulting in high potency as well as prolonged action. Compounds with high cerebral vasodilative activity would have high affinity for the active site and may bind too tightly to allow easy washing out by the blood flow, giving a long-lasting action, whereas the opposite situation would apply to compounds with ow activity. Generally, substituents containing such hetero atoms as O and N are electron-donating. However, compounds bearing 4-NH 2 (lVn) or 4 - N M e 2 (lVo and 1Vkk) are acutely toxic. Therefore, the substitution on the benzyl moiety has to be a combination of alkoxy groups. From the results obtained with compounds having various numbers and

424 locations of methoxy groups, the 2,3,4-trimethoxy seemed to be the best substitution pattern. Therefore, 1-[bis(4-fluorophenyl)methyl]-4-(2,3,4trimethoxybenzyl)piperazine was thought to be the best compound with respect to potency and duration. The dihydrochloride of this compound (IVp : KB-2796) was selected as a candidate for the development of a cerebral vasodilator, and is now under extensive clinical evaluation.

2HCI

KB-2796 F

2.4

A p p l i c a t i o n to ~

Evolution

From the slope and sign of the Z~ term in Eq. 5, it was suggested t h a t an increase of electron density on the benzylic nitrogen atom is much more important t h a n t h a t on the other nitrogen for enhancement of the potency. Since factors for the t r a n s p o r t process were assumed not to be critical, and considering the steric effect of p a r a substituents of the diphenylmethyl moiety, a model for the interaction of these compounds with the active site is proposed, as shown in Fig. 1.

~

.,y

Anionic

Site

iuClsion

Fig. 1 Model of the Active Site (Reproduced from ref. 19 by permission of the

Pharmaceutical Society of Japan).

425

The putative active site may consist of a hydrophobic pocket, bearing an anionic site which interacts electrostatically with the positively charged benzylic nitrogen atom. The hydrophobic effect of the diphenylmethyl moiety does not appear explicitly in the equations, but its introduction resulted in much higher activity than that of trimetazidine. Thus, there may be a strong hydrophobic interaction between the diphenylmethyl moiety and the wall of the pocket. The depth of the pocket from the anionic site is limited so the steric repulsion of bulky Y and Z substituents lowers the binding interaction of the molecule. The above results prompted us to a t t e m p t further structural modifications for new leads. Since the electron density on one of the two nitrogen atoms of the piperazine ring is important, we first designed the vinylog, the structure with a double bond between the phenyl moiety and the methylene bridge, to t r a n s m i t the electronic effect of substituents on the benzyl-benzene ring through the double bond to the nitrogen atom.

x

~, x

IV ~ z

VII

Y

Namely, 1-(substituted cinnamyl)-4-diphenylmethylpiperazines (VII), i.e., substituted analogs of cinnarizine (II), were synthesized and their activities were tested (20). As expected, compounds bearing electron-donating substituents on the cinnamyl moiety showed potent activity (Table 3). The 4-NMe 2 derivative (VIIe) was one of the most potent compounds, but its acute toxicity was high, as in the case of the benzyl analog. Among the compounds p r e p a r e d , l-[bis(4-fluorophenyl)methyl]-4-(2,3,4-trimethoxycinnamyl)piperazine dihydrochloride (VIIa, KB-3512) was selected for further studies. Many substituted cinnarizine derivatives have been reported (21, 22), but there seems to be no compound so far which is more potent than c i n n a r i z i n e (VII :X - Y = H) and f l u n a r i z i n e (VII :X - H , Y - F),

426

TABLE 3. Structural Features and Cerebral Vasodilative Activities of 1Cinnamyl-4-diphenylmethylpiperazines (VH)

Compd. No.

X

2,3,4-(OMe) 3 2,3,4-(OMe)~ VIIc d 2,4-(OMe) 2 VIId d 2,4-(OMe) 2 VIIe d 4-NMe 2 Cinnarizine H Flunarizine" 2HC1 H VIIa b

VIIb b

e.j

Y

Activitya

F H F H F H F

1.25c 0.98 1.25 0.99 1.65 0.71 0.79

Duration 1 1 1 1 1 0 1

a) Activity is expressed in terms of increase of maximun blood flow relative to that of papaverine ata dose of 1 mg/kg, i.v. b) Dihydrochloride. c) The dose of this compound was 0.3 mg/kg, i.v. d) Fumarate.

b e c a u s e s o m e h a v e a n e l e c t r o n - w i t h d r a w i n g g r o u p on t h e c i n n a m y l moiety a n d some h a v e a b u l k y s u b s t i t u e n t on the d i p h e n y l m e t h y l group. T h u s , KB-3512 (VIIa) is t h o u g h t to be a m o n g the most p o t e n t analogs of cinnarizine. Recently, a series of vinylogs of s u b s t i t u t e d c i n n a r i z i n e s (VIII) h a s b e e n r e p o r t e d to h a v e p o t e n t v a s o d i l a t i v e action (23). A m o n g t h e m , a vinylog of KB-3512 ( V I I I a 9 X = 2,3,4-(OMe)3, Y = F) w a s one of the most potent compounds.

It is of i n t e r e s t t h a t t h e s t r u c t u r e - a c t i v i t y

r e l a t i o n s h i p of this series of c o m p o u n d s s e e m s to be s i m i l a r to t h o s e of t h e i r benzyl (IV) a n d c i n n a m y l (VII) analogs.

•

Q.~N VIII Y

The

benzylic

nitrogen

atom

p i p e r a z i n e d e r i v a t i v e s (lV) p l a y s v a s o d i l a t i v e activity.

of 1 - b e n z y l - 4 - d i p h e n y l m e t h y l a significant

role in c e r e b r a l

T h e r e f o r e , t h r e e t y p e s of p i p e r i d i n e d e r i v a t i v e s

427 (IX - X I) w e r e p r e p a r e d in o r d e r to i n v e s t i g a t e t h e role of t h e n i t r o g e n a t o m a t t a c h e d to t h e d i p h e n y l m e t h y l m o i e t y (24). The compounds

substituted

with electron-donating

X groups

e x h i b i t e d p o t e n t a c t i v i t y as well as a long d u r a t i o n of t h e action, w h e r e a s u n s u b s t i t u t e d d e r i v a t i v e s w e r e less a c t i v e a n d did n o t s h o w a n y longl a s t i n g a c t i o n (Table 4).

A l t h o u g h t h e p o t e n c y w a s v a r i e d d e p e n d i n g on

Y

X

x OH

TABLE 4. Structural Features and Cerebral Vasodilative Activities of 1-Benzylpiperidines (IX, X and XI) Compd. No.

X

Zo

Activity a

Duration

lXa

H

0.00

N.T. b

N.T.

IXb IXc Xa c

2,3,4-(OMe) 3 2,4-(OMe) 2 H

-0.42 -0.54 0.00

1.00 1.37 0.78

0 1 0

Xb c Xc c XIa d XIb c XIc c XId e

2,3,4-(0Me) 3 2,4-(OMe) 2 H 2,3,4-(OMe) 3 2,4-(0Me)2 4-NMe 2

-0.42 -0.54 0.00 -0.42 -0.54 -0.83

1.05 0.96 0.67 1.00 1.10 1.08

1 1 0 1 1 1

a) See Footnote of Table 3. b) N.T. 9not tested, c) Hydrochloride. d) Fumarate. e) Dihydrochloride.

428 the type of X substituent in a manner similar to that observed in the original series, the mode of connection between the piperidine ring and the diphenylmethyl moiety showed little effect in these modified series. When the substituent on the benzyl moiety was equal, the piperidine analog (X I) was almost equipotent to the corresponding piperazine (IV). These results suggest that the nitrogen atom attached to the d i p h e n y l m e t h y l moiety in 1-benzyl-4-diphenylmethylpiperazine derivatives (IV) plays no special role in manifestation of the activity and is exchangeable for the carbon atom, and that the piperidine derivatives interact with the active site in a manner very similar to piperazines. Unfortunately, these compounds were observed to lack cerebrovascular specificity. F u r t h e r evaluations of these derivatives as cerebral vasodilators were terminated. 3.

ANTIULCERATlVE PIPERAZINEACETATES

Peptic ulcers are classified into duodenal and gastric types based on the region affected. These ulcers are considered to be due to imbalances between aggressive factors such as acid and pepsin and the resistance of gastrointestinal mucosa against them. Acid secretion is critical for production of duodenal ulcers, whereas gastric ulcers are mainly induced by weakening of the defensive factors. Thus, antiulcer agents are generally classified into two categories, antisecretory agents, which suppress the aggressive factors, and cytoprotective agents, which strengthen the defensive mechanisms of the gastrointestinal mucosa. For the purpose of treatment and prophylaxis of peptic ulcers, however, the cytoprotective effect is thought to be more important. 3 . 1 Strategy for ~ Identification and Optimization Various antiulcer agents exert cytoprotective activity. Examples are cetraxate hydrochloride (XII) (25), sucralfate (XIII) (26) and teprenone (XIV) (27). The structures of these known agents are quite diverse. Since these compounds do not have sufficient activity, we thought it unlikely to be fruitful to derivatize them to develop novel antiulcer agents with higher cytoprotective activity.

429

NH2CH 2 9

,, ICO0

CH2CH2COOH

H 0

HCI

Ik~R Xll 9 Cetraxate Hydrochloride

R ( ~ H

H

H/ ~0 OR

-J ~ LOR OR H

R = S03[AI2(OH)5] XlV"

Teprenone

XlII 9 Sucralfate

Therefore, we used a random screening procedure to find a novel lead structure. After the lead compound was found, a series of congeners was synthesized and tested by the indomethacin-induced ulcer model using rats. Compounds which caused a statistically significant decrease in the ulcer index defined by the size of the ulcer from the control at a dose of 200 mg/kg were judged to be active. Oral toxicity (LD50) was examined in mice. Then, QSAR analyses (ALS method) were performed for activity ratings (1 for active and 0 for inactive compounds) to obtain a p r i m a r y clue to s t r u c t u r a l requirements for the activity. For active compounds with low toxicity, the antiulcer activity was measured in terms of ED50 and the QSAR was performed (Hansch-Fujita method) for more accurate analysis of factors enhancing the activity. The combined results of two QSAR analyses were used to predict the optimal structure. 1 - B e n z y l - 4 - p i p e r a z i n e a c e ~ d e Analogs During the course of general screening of benzylpiperazine derivatives, which were originally synthesized as possible cerebral vasodilators but found to have only low activity, 1-(pyrrolidinocarbonylmethyl)-4-(2,3,4-trimethoxybenzyl)piperazine dimaleate (XVa) was found to possess potent antiulcer activity without any antisecretory activity. This prompted us to synthesize and test various analogs of this compound (28). In the first attempt, compounds with various N-substituents in place of the entire 2,3,4-trimethoxybenzyl moiety of XVa were synthesized and tested. We found that the 2,3,4-trimethoxybenzyl moiety of XVa was not replaceable by a simple alkyl or acyl moiety without significant loss of the activity. 3.2

430

MeO

"v~

N

"

~COOH

OMe XV

XVa

Compounds X V a - X V i shown in Table 5, in which the substitution p a t t e r n X in the s t r u c t u r e X V was fixed to 2,3,4-trimethoxy, were synthesized and the effects of modifications in the amide moiety on antiulcer activity were analyzed by the ALS method to obtain Eq. 7 (29), Y = - 2.474 Vw + 2.293 n-9,

Rs=0.800,

[7]

nmi s - l ,

t = 3.52,

p < 0.01

where Y is the activity rating and Vw is the van der Waal's volume in ~3 scaled by 1/100 (30) of the NRR' moiety. Equation 7 suggests t h a t compounds with the less bulky amide group were favorable for activity. Considering the data together with toxicity data, the pyrrolidino moiety seemed to be the most suitable. Next, the effects of s u b s t i t u e n t X on the benzyl moiety were i n v e s t i g a t e d fixing the amide moiety as pyrrolidinocarbonyl with compounds XVj - X V u . In preliminary experiments, we observed that substitution at both the 3 and 4 positions was necessary for the activity. Thus, we examined various physicochemical p a r a m e t e r s of 3- and 4substituents. Among compounds with various substitution patterns of methoxy groups, only two were active: the 2,3,4- and 3,4,5-trimethoxy derivatives. The unhindered methoxy group is thought to be coplanar

~ f-~

a

CH

b

Fig. 2 Copl~n~r conformation (a; conjugated) and the out.of-plane conformation (b ; non-conjugated) (Reproduced from res 29 by p e ~ i . ~ i o n of the Pharmaceutical Society of Japan).

431 TABLE

5.

S t r u c t u r a l F e a t u r e s a n d Antiulcer Activities of 1-(Aminocarbonylalkyl)~-benzylpipernzlne Dimaleates (XV)

Compd. No.

Activity X

R

R'

n

Vw

H

-(CH2) 4H

1 1

0.705 0.177

1.35 1.35

1.90 1.90

XVc

2,3,4-(OMe) 3 2,3,4-(OMe) 3 2,3,4-(OMe) 3

Et

Et

1

0.809

1.35

1.90

1

1

XVd

2,3,4-(0Me) 3

Pr

Pr

1

1.117

1.35

1.90

0

0

XVe

2,3,4-(OMe) 3

1

0.859

1.35

1.90

0

1

XVf

1

1.005

1.35

1.90

0

0

Ph

1

0.879

1.35

1.90

1

1

CH2Ph Ph

1

1.033

1.35

1.90

0

0

1

1.041

1.35

1.90

0

0

-(CH2) 4-

1

0.705

1.00

1.00

0

0

1

0.705

1.00

1.52

0

0

1 1

0.705 0.705

1.00 1.00

1.80 1.35

0 0

0 0

XVa

XVb

2,3,4-(OMe) 3

-(CH2) 5H c-Hex

XVg

2,3,4-(0Me) 3

H

XVh

2,3,4-(0Me) 3

XVi

2,3,4-(0Me) 3 H

H Me

XVj XVk

T3

T4

Obs. Calcd. (Eq.8) 1 1

1 1

XVI

4-Me 4-C1

XVm

4-OMe

-(CH2) 4-(CH2) 4-(CH2) 4-

XVn

3,4-C12

-(CH2) 4-

1

0.705

1.80

1.80

1

1

XVo

2,4-C12 2-OMe

-(CH2) 4-

1

0.705

1.00

1.80

0

0

-(CH2) 4-

1

0.705

1.00

1.00

0

0

0.705

1.35

1.00

0

0

XVp XVq

3-OMe

-(CH2) 4-

1

XVr

2,4-(OMe) 2 3,4-(OMe) 2

-(CH2) 4-(CH2) 4-

1

0.705

1.00

1.35

0

0

XVs

1

0.705

1.35

1.35

0

0

XVt XVu

3,4,5-(OMe) 3 2,4,6-(OMe) 3

-(CH2) 4-(CH2) 4-

1 1

0.705 0.705

1.90 1.00

1.35 1.35

1 0

1 0

XVv

2,3,4-(OMe) 3

-(CH2) 4-

2

0.705

1.35

1.90

1

1

XVw

2,3,4-(0Me) 3

-(CH2) 4-

3

0.705

1.35

1.90

0

0

XVx

XVy

2,3,4-(OMe) 3 3,4,5-(OMe) 3

-(CH2) 4-(CH2) 4-

4 2

0.705 0.705

1.35 1.90

1.90 1.35

0 1

0 1

XVz

3,4,5-(0Me) 3

-(CH2) 4-

3

0.705

1.90

1.35

0

0

XVaa

3,4,5-(0Me) 3

-(CH2) 4-

4

0.705

1.90

1.35

0

0

432 with the benzene ring owing to the conjugation of the lone pair electrons with the aromatic ring. On the other hand, the central methoxy group of the 2,3,4- and 3,4,5-trimethoxyphenyl moiety may be forced out of and nearly perpendicular to the plane of the aromatic ring (31). The rotational barrier of Me around the Ar-O bond of substituted anisoles is reported to be about 3-6 kcal/mol (32), so the methoxy group could be relatively easy to rotate. For the hindered perpendicular methoxy group, the smaller value of the two thickness values for directions above and below the ring plane is represented by the Verloop's STERIMOL p a r a m e t e r B 1 which is attributed mostly to the radius of the oxygen atom (33). For the unhindered coplanar methoxy group, the smaller thickness is expressible by the STERIMOL B 2. These situations are depicted in Fig. 2. With these conformational features in mind, all analogs shown in Table 5 were subjected to analysis using an additional indicator variable, n, which is the number of methylene groups between the carbonyl group and piperazine, to obtain Eq. 8, Y = - 2.631 Vw + 1.960 T 3 + 1.449 T 4 - 0.482 n - 2.166 n - 2 7 , Rs-0.918, nmi s - l , t-11.57, p<0.001

[8]

where T is the smaller of the two thickness values for each substituent above and below the ring plane. The subscript represents the substituent position. The T value of each substituent is equivalent with the STERIMOL B 1 except for the unhindered methoxy group where it is taken as the B 2 parameter. Equation 8 shows that (a) a bulky amide moiety is disadvantageous, (b) the number of methylene units should be low, and (c) the "minimum" thickness of 3- and 4-substituents on the benzyl-benzene ring should be large for high antiulcer activity. Several compounds exhibited an antiulcer activity superior to that of the reference compounds, but, because of their subacute toxicity, they were not acceptable. 3.3

Esters of 1-Benzyl-4-piperazinealkanoic acids If just small size is among the most important conditions for the potent antiulcer activity, then the NRR' moiety could be replaceable

433 with other small groups. Thus, we expected their ester analogs to be active and found t h a t some of t h e m were indeed active as shown in Table 6 (34).

x~N/"~

0 L~,~/N- (CH2)n,,~OR XMI

P r e l i m i n a r y examinations showed t h a t the van der Waal's volume, Vw, of the OR group is a significant factor governing the potency rating, as expected. Among derivatives judged as active, the phenyl ester (XVIi) was moderately potent and least toxic as summarized in Table 7. Therefore, the s u b s t i t u t e d phenyl ester derivatives were synthesized and tested. The ALS analysis of these derivatives (Table 6, XVI1 - XVIq) gave Eq. 9, [9]

Y = - 4.591~ - 0.459 n-6,

Rs-l.000,

nmi s = 0

w h e r e ~ is the H a m m e t t constant and the ~p value was used for the ortho substituent. In Eq. 9, Vw was not significant, because variations in the Vwvalue were not so large within these substituted phenyl s u b s t i t u e n t s . The negative o t e r m of Eq. 9 s u g g e s t s t h a t electrond o n a t i n g s u b s t i t u e n t s m a k e the e s t e r s stable a g a i n s t hydrolytic transformation. H y d r o l y s i s prior to r e a c h i n g the action site(s) decreases the antiulcer activity, because the corresponding carboxylic acids are inactive. Next, all of the ester compounds listed in Table 6 (XVIa - X V I a a ) were analyzed together and Eq. 10 was obtained. Y - - 2.624 Vw + 1.779 T 3 + 1.649 T 4 - 4.109 u - 1.621 n - 1.562 n-27,

Rs-l.000,

[10]

nmi s - 0

E q u a t i o n 10 shows s t r u c t u r a l r e q u i r e m e n t s for activity of esters essentially the same as Eq. 8 for those of amides, in addition to the requirement for the phenyl moiety to possess electron-donating groups.

434 TABLE 6. S t r u c t u r a l F e a t u r e s a n d A n t i u l c e r Activities of 1-Benzyl~l-piper~_ zinea l k a n o i c Acid E s t e r D i m a l e a t e s (XVI) Compd.

Activity

No.

X

R

n

Vw

~

T3

T4

Obs. Calcd. (Eq.10)

XVIa

2,3,4-(OMe) 3

Me

1

0.304

0.00

1.35

1.90

1

1

XVIb

2,3,4-(OMe) 3

Et

1

0.458

0.00

1.35

1.90

1

1

XVIc

2,3,4-(OMe) 3

Pr

1

0.612

0.00

1.35

1.90

1

1

XVId

2,3,4-(0Me) 3

Bu

1

0.766

0.00

1.35

1.90

1

1

XVIe

2,3,4-(OMe) 3

Am

1

0.920

0.00

1.35

1.90

0

0

XVIf

2,3,4-(0Me) 3

Hex

1

1.074

0.00

1.35

1.90

0

0

XVIg

2,3,4-(0Me) 3

Hep

1

1.228

0.00

1.35

1.90

0

0

XVIh

2,3,4-(0Me) 3

iso-Pr

1

0.607

0.00

1.35

1.90

1

1

XVIi a

2,3,4-(OMe) 3

Ph

1

0.844

0.00

1.35

1.90

1

1

XVIj a

2,3,4-(0Me) 3

CH2Ph

1

0.998

0.00

1.35

1.90

0

0

XVIk

2,3,4-(OMe) 3

(CH2)2Ph

1

1.152

0.00

1.35

1.90

0

0

XVI1 a

2,3,4-(0Me) 3

4-Me-Ph

1

0.998

-0.17

1.35

1.90

1

1

X V I m a 2,3,4-(0Me) 3

4-C1-Ph

1

1.009

0.23

1.35

1.90

0

0

XVIn a

2,3,4-(OMe) 3

4-MeO-Ph

1

1.079

-0.27

1.35

1.90

1

1

XVIo a

2,3,4-(OMe) 3

3-MeO-Ph

1

1.079

0.12

1.35

1.90

0

0

XVIp a

2,3,4-(0Me) 3

2-MeO-Ph

1

1.079

-0.27

1.35

1.90

1

1

XVIq a

2,3,4-(OMe) 3

4-EtO-Ph

1

1.233

-0.24

1.35

1.90

1

1

XVIr a

H

4-MeO-Ph

1

1.079

-0.27

1.00

1.00

0

0

XVIs a

4-Me

4-MeO-Ph

1

1.079

-0.27

1.00

1.52

0

0

XVIt a

4-C1

4-MeO-Ph

1

1.079

-0.27

1.00

1.80

0

0

XVIu a

4-OMe

4-MeO-Ph

1

1.079

-0.27

1.00

1.35

0

0

XVIv a

3,4-C12

4-MeO-Ph

1

1.079

-0.27

1.80

1.80

1

1

XVIw a

3,4-(OMe) 2

4-MeO-Ph

1

1.079

-0.27

1.35

1.35

0

0

XVIx a

3,4-0CH20-

4-MeO-Ph

1

1.079

-0.27

1.90

1.90

1

1

XVIy b

3,4,5-(0Me) 3

4-MeO-Ph

1

1.079

-0.27

1.90

1.35

1

1

XVIz a

2,3,4-(0Me) 3

4-MeO-Ph

2

1.079

-0.27

1.35

1.90

0

0

X V I a a a 3,4,5-(0Me) 3

4-MeO-Ph

2

1.079

-0.27

1.90

1.35

0

0

a) Dihydrochloride. b) Difumarate.

435 Although s t r u c t u r a l modifications conforming to the above r e q u i r e m e n t s could be made, Eq. 10 only predicts w h e t h e r the compound is active or not. The prediction of compounds more potent than other potent compounds is beyond the ability of Eq. 10, when every compound belongs to the same category. At this point, we used the Hansch-Fujita analysis for eleven compounds listed in Table 7, five amides and six esters, selected in terms of the low acute toxicity so that the LD50 is higher than 2 g/kg, p.o. Compound XVIn is the most active with an ED50 against the indomethacin-induced ulcers of 10 mg/kg, p.o., whereas compound XVa is the least active with an ED50 of 164 mg/kg, p.o. For 11 compounds, Eq. 11 was derived, log(l/C) = 1.253(+0.392) Vw + 3.349(+0.293) n = 1 1 , r=0.923, s=0.15, F=52.13

[11]

where C is the ED 50 (mol/kg p.o.)value. Equation 11 indicates that the larger the Vw value of the OR and NRR' group, the more potent is the antiulcer activity. The number of methylene units in the bridging moiety is not critical. This result is contrary in terms of the steric effect to the ALS results represented in Eqs. 8 and 10. The discrepancy could be attributed to the fact t h a t the ALS analyses are just to distinguish "active" compounds from a number of "inactive" compounds. The overall trend that the lower steric Vw value is more favorable may be the case. For the active compounds included in Eq. 11, which were given the rating score of one in Eqs. 8 and 10 irrespective of the potency variations in terms of log(I/C), the steric effect of NRR' and OR on the activity may well be different from that suggested by the ALS analyses. The above results suggested that even if a compound of this series were predicted to be active according to Eqs. 8 and 10, there is no need to synthesize and test it when the OR group is smaller t h a n t h a t in compounds XVIn and XVIy. Therefore, we thought it advisable to terminate any further analog synthesis from the standpoints of availability of raw materials and ease of synthesis. For the selected six ester derivatives (Table 7), antiulcer activities against other ulcer models were examined and two compounds (XVIn, XVIy) were found to possess an antiulcer activity superior to those of

436 TABLE 7.

S t r u c t u r a l Features, Antiulcer Activities a n d Toxicities of Selected 1Benzylpiperazinealkanoic acid Derivatives

Compd. No.

log (1/C) X

OR or NRR'

n

Vw

NIZ2, Pyrr D

1 1

0.177 0.705

LD50 a (mg/kg)

Obs.

Calcd. (Eq.ll)

~ 2500

3.53 4.31

3.57 4.23

XVb XVa

2,3,4-(0Me) 3 2,3,4-(OMe) 3

XVt

3,4,5-(0Me) 3

Pyrr

1

0.705

4200

4.29

4.23

XVv

2,3,4-(0Me) 3

Pyrr

2

0.705

3400

4.37

4.23

XVy

3,4,5-(0Me) 3

Pyrr

2

0.705

3600

4.34

4.23

XVIa

2,3,4-(0Me) 3

OMe

1

0.304

3120

3.87

3.73

XVIc

2,3,4-(0Me) 3

OPr

1

0.612

3930

3.80

4.11

XVId

2,3,4-(0Me) 3

OBu

1

0.776

4000

4.21

4.31

XVIi

2,3,4-(0Me) 3

OPh

1

0.844

5660

4.24

4.41

XVIn

2,3,4-(0Me) 3

OPh(4-OMe)

1

1.079

4200

4.70

4.70

XVIy

3,4,5-(0Me) 3

OPh(4-OMe)

1

1.079

4800

4.79

4.70

a) The LD50 values for the active compounds listed in Tables 5 and 6 except for those listed here are below 3000 mg/kg, b) Pyrr : pyrrolidino.

such reference compounds as XII, XIII and XIV. Since its toxicity was low and acceptable, we selected 1-(3,4,5-trimethoxybenzyl)-4-(4methoxyphenyloxycarbonylmethyl)piperazine (XVIy) as a candidate for the antiulcer drug development (KB-5492 as monofumarate).

OMe

KB-5492

The antiulcer mechanism of this novel series of compounds seems to be their cytoprotective activity, because no suppressive effect was observed against secretion of acid or pepsin. Several 1-piperazineacetamides, including esaprazole (XVII) (35), pirenzepine (XVIII) (36) and fenoverine (X IX) (37), have been used clinically as antiulcer agents.

437

XVII: Esaprazole

XVlll : Pirenzepine

X l X : Fenoverine

These compounds show antisecretory activity. It is impossible to deny that experienced medicinal chemists would easily be able to design compounds with the structure X V as possible antiulcer agents from structures of such existing drugs as shown above, but it would not be readily predictable that not only amides (XV) but also esters (XVI) of 1benzyl-4-piperazineacetic acid show antiulcer activity based on cytoprotection. Structural resemblance does not necessarily imply similarity in the mechanisms of antiulcer activity. Extrapolative application of the QSAR results of the amides series brought forth a novel lead structure (alkyl esters) and the antiulcer activity was optimized efficiently by substituent modifications aided by the QSAR. 4. ANTIHISTAMINIC 2-PIPERAZINYLBENZIMIDAZOLES In the course of search for antiinflammatory benzimidazole derivatives, 1-alkyl-2-(4-methyl-l-piperazinyl)benzimidazoles (38) were prepared. Pharmacological profiles suggested t h a t these compounds could be possible leads for Hl-antihistaminics without significant side effects. Antihistaminics are useful for treating the symptoms of allergic reactions including seasonal hay-fever, allergic rhinitis and conjunctivitis. Conventionally used antihistaminics have, however, certain drawbacks in that they frequently induce side effects such as dry mouth resulting from their anticholinergic activity, and exert central nervous system (CNS)-depressive effects such as sedation and hypnosis. Therefore, Iemura and co-workers started a project to develop compounds with not only higher antihistaminic activity but also lower anticholinergic and CNS-depressive activities. Analogs were synthesized and their antihistaminic activities (IC50 in M) were measured using isolated ileum from guinea pigs. Among these analogs, 1- (2-ethoxyethyl)-2-(4-m ethyl- 1-hom opiper az inyl) ben zimid az ole

438

(COOH KG-2413

N " ~ HOOC/ I CH2CH2OCH2CH3

,Me ~N N-R2 I i (CH2)m R1 XX

CI~

N. Me XXI: Chlorpheniramine

difumarate (XXnn : KG-2413) was selected for further study (39). Its activity in vivo was 39 times more potent than t h a t of chlorpheniramine (XX I) (40), which is one of the most potent H 1-antihistaminic agents known. QSAR analyses were performed in order to confirm the validity of the selection (41). 4 . 1 Antihistaminic Activity Because hydrophobic (n) and steric (MR) p a r a m e t e r s are linearly related to the n u m b e r of methylenes (NM) in s t r a i g h t chain alkyl groups, NM was used as a temporary makeshift. For the compounds (XX a - X X g) in Table 8 with straight alkyl chains at the 1-position of the 2-(4-methyl-l-piperazinyl)benzimidazole, a plot of antihistaminic activity (log 1/IC50) against NM of the alkyl chain as R 1 suggested a parabolic relationship. Then, compounds with various substituents at the 1-position of the 2-(4-methyl-l-piperazinyl)benzimidazole ( X X a X X d d ) were subjected to analysis (Table 8). Analysis using another variable NA for all but seven compounds ( X X h - X X k , X X p , X X s and X X c c) gave Eq. 12. logl/IC50 = - 0.079(+_0.033) NA 2 + 0.875(+0.397) NA + 5.155(+1.165) n = 23, r = 0.754, s = 0.480, F= 13.16 [12] NA is defined as the n u m b e r of atoms other t h a n h y d r o g e n in substituents at the 1-position. The addition of other physicochemical s u b s t i t u e n t p a r a m e t e r t e r m s was not significant. Although the correlation was not very good, NA seemed to rationalize the effects of R 1

439

best as a single parameter. Since the NA p a r a m e t e r was not depend on the type of atom or bond, the physicochemical meaning of NA seemed to be steric bulk. As described above, the activities of seven compounds deviated markedly from those calculated using Eq. 12. For compounds X X h X X k , the large width of the s u b s t i t u e n t s may be unfavorable for activity. The low activities of X X p and X X s might be ascribed to the folding of the R 1 substituent, as observed in 1-(3-phenoxypropyl)uracil (42). The higher activity t h a n predicted for compound XXcc seemed to be due to the fact t h a t the phenoxyethyl group fits the cavity of the receptor better t h a n expected. If these assumptions were reasonable, the variations in the activity could be analyzed by steric p a r a m e t e r s representing the width as well as the length of substituents. Thus, an analysis including these seven compounds ( X X a - X X d d ) was performed using Verloop's STERIMOL parameters (33). In e s t i m a t i n g STERIMOL p a r a m e t e r s , the R 1 chain was assumed to take the fully staggered conformation extending toward the direction p e r p e n d i c u l a r to the benzimidazole ring except in the following cases: the 1-phenyl group (XXk) is coplanar with the benzimidazole ring, the 3-(ethylthio)propyl (XXp) and 3-(methoxy)propyl (XXs) groups fold onto the benzimidazole ring, and the benzene ring not directly attached to the benzimidazole ring in compounds XXl, X X m , X X c c and X X d d is perpendicular to the benzimidazole plane. With the two types of STERIMOL parameters, a good correlation was obtained as shown in Eq. 13. logl/IC50= - 0.096(+0.025) L 2 + 1.413(+0.369) L - 1.173(+0.321) B 3 + 4.686(_+1.401) [13] n = 30, r = 0.891, s = 0.397, F = 33.32 In Eq. 13, L represents the length p a r a m e t e r for R 1 substituents along the axis connecting the 1-N atom of the benzimidazole with the a-atom of R 1 substituents considering the folding factor for compounds X X p and X X s . B 3 is the p a r a m e t e r for the second largest width of substituents. Here, it represents the larger one of the two width value

440 TABLE 8.

Structural Features azoles (XX)

a n d A n t i h i s t a m i n i c Activities of Benzimid-

Compd. No.

log 1/IC50 R1

R2

m

B3

L

I

Obs.

Calcd. (Eq.14)

XXa a XXb b

Me Pr

Me

2

1.90

3.00

0

6.50

5.75

Me

2

1.90

5.05

0

6.42

7.13

XXc a XXd c

Bu Am

Me

2

1.90

6.17

0

7.30

7.55

Me

2

1.90

7.11

0

7.72

7.71

XXe c

Hex

Me

2

1.90

8.22

0

7.59

7.67

XXf c

Hep

Me

2

1.90

9.16

0

6.82

7.46

XXg b

Dec

Me

2

1.90

12.33

0

5.38

5.47

XXh XXi b

(CH2)2CHMe 2 CHMePr

Me

2

2.76

6.17

0

5.85

6.51

Me

2

3.66

6.17

0

5.85

5.43

X~ b XXk c

CH2CHMePr Ph

Me

2

3.18

7.11

0

5.82

6.17

Me

2

3.11

6.28

0

6.12

6.12

XXl a

CH2Ph

Me

2

1.90

5.91

0

7.77

7.47

XXm c

(CH2)2Ph

Me

2

1.90

8.41

0

7.62

7.64

XXn

CH2SPr

Me

2

1.90

7.59

0

7.17

7.72

XXo

(CH2)2SEt

Me

2

1.90

7.29

0

7.72

7.72

XXp XXq c

(CH2)3SEt CH2OPr

Me

2

1.90

3.62

0

6.28

6.25

Me

2

1.90

6.95

0

7.75

7.69

XXr c

(CH2)2OEt

Me

2

1.90

6.97

0

8.00

7.69

XXs b

(CH2)3OMe

Me

2

1.90

3.62

0

6.06

6.25

XXt a

(CH2)2NHEt

Me

2

3.03

6.68

0

6.54

6.30

XXu a XXv a

(CH2)2OH (CH2)2OMe

Me

2

1.90

4.79

0

6.39

7.00

Me

2

1.90

6.03

0

7.89

7.51

XXw

(CH2)2OCH=CH 2

Me

2

1.90

7.09

0

8.00

7.70

XXx d XXy c

(CH2)20(CH2)2OH (CH2)2OPr

Me

2

1.90

7.95

0

7.37

7.70

Me

2

1.90

8.10

0

7.70

7.69

XXz c

(CH2)2OCH2CH=CH 2

Me

2

1.90

8.32

0

7.77

7.66

XXaa c

(CH2)2OCH2C-CH

Me

2

1.90

8.73

0

7.92

7.58

XXbb c

(CH2)2OBu

Me

2

1.90

9.04

0

7.42

7.49

XXcc c

(CH2)2OPh

Me

2

1.90

7.85

0

8.16

7.71

XXdd d

(CH2)2OCH2Ph

Me

2

1.90

10.33

0

6.37

6.95

XXee c

(CH2)2OEt

H

2

1.90

6.97

0

7.96

7.69

XXff e

(CH2)2OEt

Et

2

1.90

6.97

0

7.75

7.69

441 TABLE 8. Continued Compd. No.

R1

XXgga XXhh a XXii a XXjj a XXkk a XXll a XXmm c XXnn b XXoob XXpp a XXqqa XXrr a XXss a XXttb XXuu b XXvvb XXwwb XXxx b

R2

(CH2)2OEt Pr (CH2)2OEt Bu (CH2)2OEt Am (CH2)2OEt Hex (CH2)2OEt CH2Ph (CH2)2OEt (CH2)2Ph (CH2)2OEt H (CH2)2OEt Me (CH2)2OEt Et (CH2)2OEt Pr (CH2)2OEt Bu (CH2)2OEt Am (CH2)2OEt CH2Ph (CH2)2OPr Me (CH2)2OCH2CH=CH 2 Me (CH2)2OCH2C-CH Me (CH2)2OBu Me (CH2)2OPh Me

m

B3

2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 3

1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90 1.90

6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 6.97 8.10 8.32 8.73 9.04 7.85

I

log 1/IC50 Obs. Calcd. (Eq.14)

0 0 0 0 0 0 1 1 1 1 1 1 1 1 1 1 1 1

7.80 7.82 7.52 8.06 7.57 7.51 7.60 8.21 7.80 8.08 8.08 8.13 7.82 7.80 8.00 8.00 8.00 8.04

7.69 7.69 7.69 7.69 7.69 7.69 7.99 7.99 7.99 7.99 7.99 7.99 7.99 7.99 7.96 7.88 7.79 8.01

a) Dimaleate. b) Difumarate. c) 1.5 Fumarate. d) Fumarate.

for each s u b s t i t u e n t from the L-axis in two opposite di rect i ons p a r a l l e l w i t h th e b e n z i m i d a z o l e r i ng plane. The l a r g e s t w i d t h p a r a m e t e r B 4 did not w o r k b e t t e r t h a n B 3, because of t he difference in directions defining the

B parameters

of s u b s t i t u e n t s .

The

assumptions

for

the

c o n f o r m a t i o n of t h e R 1 s u b s t i t u e n t s o t h e r t h a n those t a k e n here did not w o r k well either. Thus, Eq. 13 would indicate t h a t the lower t he l a r g e r width,

i.e.,

t h e m or e s y m m e t r i c t he w i d t h s of s u b s t i t u e n t s from t h e L-

axis in t h e direction p a r a l l e l w i t h the b e n z i m i d a z o l e ring, t h e h i g h e r is the activity.

The o p t i m u m l engt h is r e q ui red for the R 1 substituents.

N e x t , t h e c o m p o u n d s (XXee

- XXxx)

in w h i c h R 2 is e i t h e r

h y d r o g e n or a n a l k y l g r o u p o t h e r t h a n m e t h y l a n d m is t h r e e w ere

442 considered together, leading to Eq. 14, l o g l / I C 5 0 - - 0.098(+0.019) L 2 + 1.440(+0.287) L - 1.194(+_0.252) B 3 + 0.338(+0.231) I + 4.643(+_1.135) [14] n = 5 0 , r=0.912, s=0.329, F=55.63 where I is the indicator variable for homopiperazine (I = 1) derivatives. Equation 14 indicates that the substituent R 2 at the 4-N-position of piperazine and homopiperazine has essentially no effect on the activity, and t h a t homopiperazines (I = 1) are almost uniformly more active t h a n the corresponding piperazines. The positively charged homopiperazine nitrogen may be situated closer to the anionic site of the receptor than the piperazine nitrogen. Otherwise, Eq. 14 is practically equivalent with Eq. 13. The length (L - 7.3) of the R 1 substituent is optimal for activity. From the above results, a model for the receptor binding features of this type of compounds is proposed as shown in Fig. 3. In the hypothetical receptor, an anionic site is present, which interacts electronically with the positively charged piperazine or homopiperazine nitrogen atom. There is also a slit-shaped cavity perpendicular to the region where the benzimidazole ring moiety p e r h a p s binds hydrophobically, as proposed by Rekker et. al. for antihistaminic diphenhydramine derivatives (43)

t

,

~~NHR (~

AnionicSite

:t,J"....":Cavy

Fig. 3 Model of the Binding Sites of 2-Piper~zinylbenzimidazoles (Reproduced from res 41 by permission of the Pharmaceutical Society of Japan).

443

Substructural units possessing tertiary amino groups such as piperidine, ethylenediamine, piperazine and homopiperazine moieties in conventional antihistaminics are thought to be bioisosteric. Thus, the benzimidazole derivatives with possible bioisosteric substituents at the 2position ( X X I I - XXV) shown in Table 9 were synthesized and tested (44). ~~"/',N > - -~"NNH

--~N-

I

R2

,~"',,,,~.. N R2 ~!~ 2>-- NH(CH2'm'--N:R2 I

R1 XXll

R1

XXIII

N

/--k

N I

R1 XlV

I

R1 XXV

(CH2)m

The a n t i h i s t a m i n i c activities of these compounds were in good a g r e e m e n t with those expected from Eq. 14. Therefore, all 82 compounds were subjected to analysis, and Eq. 15 was obtained as the best equation. log 1/IC50- - 0.097(+0.018) L 2 + 1.458(+0.260) L - 1.202(+0.234) B 3 + 0.299(+0.202) I + 4.528(+1.081) [15] n = 8 2 , r=0.875, s=0.321, F=62.71 Equations 14 and 15 are essentially equivalent. The antihistaminic activity (in vitro) of the additionally prepared compounds was correctly predicted by Eq. 14. In the in vivo test, however, only compounds X X V showed considerably potent activity comparable to t h a t of KG-2413. These results indicate that the pharmacokinetic characteristics such as absorption, distribution and metabolism of compounds X X V and X X, are more favorable to the activity than those of other derivatives. 4 . 2 Anticholinergic Activity As described above, classical antihistaminics commonly have unfavorable side effects due to anticholinergic activities. Therefore, the anticholinergic activities of twelve compounds (X X, Table 1 0 ) t h a t

444 TABLE

9.

S t r u c t u r a l F e a t u r e s a n d A n t i h i s t a m i n i c Activities of Various Types of Benzimidazoles

Compd.

log 1/IC50

No.

R1

R2

m

B3

L

I

Obs. Calcd. (Eq.15)

XXIIa b

(CH2)2OEt

Me

-

1.90

6.97

0

7.75

7.67

XXIlb c

(CH2)2OCH2CH=CH 2

Me

-

1.90

8.32

0

7.85

7.63

XXIIc b

(CH2)2OPh

Me

-

1.90

7.85

0

6.89

7.68

XXIId b

(CH2)2OEt

CH2Ph

-

1.90

6.97

0

7.18

7.67

XXIIe b

(CH2)2OEt

H

-

1.90

6.97

0

7.70

7.67

XXIIIa a

(CH2)2OEt

Me

2

1.90

6.97

0

7.89

7.67

XXIIIb a

(CH2)2OEt

Et

2

1.90

6.97

0

7.36

7.67

XXIIIc a

(CH2)2OEt

(CH2)2 -e

2

1.90

6.97

0

8.06

7.67

XXIIId a

(CH2)2OEt

Me

3

1.90

6.97

0

7.59

7.67

XXIIIe a

(CH2)2OEt

Et

3

1.90

6.97

0

7.23

7.67

XXIVa d

(CH2)2OEt

Me

-

1.90

6.97

0

7.77

7.67

XXIVb c

(CH2)2OCH2CH=CH 2

Me

-

1.90

8.32

0

7.96

7.63

XXIVc c

(CH2)2OCH2C-CH

Me

-

1.90

8.73

0

7.92

7.55

XXIVd c

(CH2)2OPh

Me

-

1.90

7.85

0

7.72

7.68

XXIVe c

(CH2)2OEt

H

-

1.90

6.97

0

7.92

7.67

XXIVf c

(CH2)2OEt

Et

-

1.90

6.97

0

7.82

7.67

XXVa d

(CH2)2OEt

Me

2

1.90

6.97

0

7.68

7.67

XXVb

(CH2)2OCH=CH 2

Me

2

1.90

7.09

0

7.70

7.68

XXVc b

(CH2)2OPr

Me

2

1.90

8.10

0

7.80

7.66

XXVd d

(CH2)2OCH2CH=CH 2

Me

2

1.90

8.32

0

8.02

7.63

XXVe d

(CH2)2OCH2C-CH

Me

2

1.90

8.73

0

7.92

7.55

XXVf d

(CH2)2OEt

H

2

1.90

6.97

0

7.62

7.67

XXVg c

(CH2)2OEt

Et

2

1.90

6.97

0

7.68

7.67

XXVh c

(CH2)2OEt

Pr

2

1.90

6.97

0

7.47

7.67

XXVi c

(CH2)2OEt

2

1.90

6.97

0

7.08

7.67

XXVj c

(CH2)2OEt

Me

3

1.90

6.97

1

7.85

7.96

XXVk c

(CH2)2OPr

Me

3

1.90

8.10

1

7.75

7.95

XXV1 c

(CH2)2OCH2CH=CH2

Me

3

1.90

8.32

1

8.04

7.92

XXVm c

(CH2)2OCH2C-CH

Me

3

1.90

8.73

1

8.04

7.83

XXVn c

(CH2)2OPh

Me

3

1.90

7.85

1

7.70

7.97

XXVo c

(CH2)2OEt

H

3

1.90

6.97

1

7.70

7.96

XXVp c

(CH2)2OEt

Et

3

1.90

6.97

1

8.00

7.96

(CH2)2OH

a-d) See footnot of Table 8. e) pyrrolidino.

445 s h o w e d p o t e n t a n t i h i s t a m i n i c activity in vitro as well as in vivo, w e r e m e a s u r e d . The IC 50 (M) values were e v a l u a t e d u s i n g isolated ileum from g u i n e a pigs by t h e u s u a l m e t h o d . Since t h e a n t i c h o l i n e r g i c potency is about four orders of m a g n i t u d e lower t h a n the a n t i h i s t a m i n i c potency in t e r m s of 1/IC 50' the anticholinergic side effects of this series of compounds were not serious. To examine the factors p a r t i c i p a t i n g in the anticholinergic potency, analysis was performed to give Eq. 16. log 1 / I C 5 0 - 0.287(_+0.198) B 4 + 0.725(+0.405) I + 2.411(+1.103) n-12,

r-0.879,

s-0.304,

[16]

F-15.37

In Eq. 16, B 4 r e p r e s e n t s the S T E R I M O L m a x i m u m w i d t h p a r a m e t e r of t h e R 1 s u b s t i t u e n t a n d I is the i n d i c a t o r v a r i a b l e for t h e homopiperazines. E q u a t i o n 16 indicates t h a t the more s y m m e t r i c the widths of R 1 s u b s t i t u e n t s in compounds carrying the piperazine ring, the lower is the anticholinergic activity.

TABLE 10. Anticholinergic and CN~Depressive Activities of Selected Benzimidazoles

Compd. No.

B4

MR/10

I

XXr XXy XXz

4.82 5.75 5.92 4.38 7.42 4.82 4.82 4.82 5.75 5.92 4.38 7.42

2.136 2.602 2.555 2.526 3.684 2.136 2.136 2.136 2.602 2.555 2.526 3.684

0 0 0 0 0 0 0 1 1 1 1 1

XXaa

XXcc XXee

XXff XXnn XXtt XXuu

XXvv XXxx

Anticholinergic log 1/IC50 Obs. Calcd. (Eq.16) 3.59 4.76 3.99 3.82 4.35 3.67 3.60 4.51 5.12 4.60 4.32 5.26

3.80 4.06 4.11 3.67 4.54 3.80 3.80 4.52 4.79 4.84 4.40 5.27

CNS-Depressive Effect Obs. Recog. Pred. (Eq.17) 1 1 1 1 1 0 0 0 1 0 0 1

0 1 1 1 1 0 0 0 1 0 0 1

0 1 1 1 1 0 0 0 0 0 0 1

446

4.3

CNS-Depressive Effect The other type of the common side effects of antihistaminics is hypnotic-sedative ( C N S - d e p r e s s i v e ) a c t i v i t y , r e s u l t i n g in d a y t i m e drowsiness, lack of concentration, diminished mental acuity and impaired handling of machinery or driving of vehicles. We evaluated the CNS-depressive effects of twelve benzimidazole derivatives (Table 10) in terms of their potentiation of hexobarbital-induced sleep in mice. Compounds that caused statistically significant increase in the period of hexobarbital-induced sleep at a dose of 200 mg/kg p.o. were classified as "active", and others as "inactive". QSAR was analyzed by the ALS method to obtain Eq. 17, where MR is the value of the R 1 substituent. Y = 1.061 (MR/10) - 0.431 1 - 2.458 n=12,

Rs=0.845,

nmi s = l ,

[17]

t=5.00,

p < 0.001

For confirmation of the validity of the ALS result, the leave-one-out test was performed. The predictive results showed t h a t 83% of the compounds were classified correctly. Equation 17 suggests t h a t a sterically small substituent at the 1-position and the homopiperazine moiety at the 2-position decrease the extent of CNS side effects. Astemizole, 1-(4-fluorobenzyl)-2-[[1-(4-methoxyphenethyl)-4piperidyl]amino]benzimidazole (XXVI), has been used clinically as a long-acting a n t i h i s t a m i n i c agent with few CNS side effects (45). Recently, the 1-(2-ethoxyethyl) analog of XXVI was publicized in the patent form (46). The structure of XXVI is similar to that of X X I I , and if the two compounds interact with the same active site(s) in a similar m a n n e r , Eqs. 15 and 17 suggest that the 2-ethoxyethyl moiety (L = 6.97, B 3 = 1.90 and MR/10 = 2.136) would be preferable to the 4-fluorobenzyl moiety (L - 5.91, B 3 - 1.90 and MR/10 - 2.990) in terms of both antihistaminic and CNS activity.

~_

NNk'~--NH" - ~ N - CH2CH2- - ~ ~

F

OMe

XXVl " Astemizole

447

In conclusion, compounds which have a 1-(2-ethoxyethyl) and 2(1-homopiperazinyl) substitution on the benzimidazole nucleus were confirmed to have not only potent antihistaminic activity but also low anticholinergic and CNS-depressive activities from the results of QSAR. Therefore, we selected KG-2413 as a candidate compound for d e v e l o p m e n t a l trials. KG-2413 also showed antiallergic and antiasthmatic effects and has been used clinically since August 1993. 5. C O N C L U S I O N Our application of the QSAR technique has been pragmatic to disclose the optimized structure in the shortest and most efficient way. Therefore, we have restricted ourselves to the application of established methods and the use of well defined substituent constants as far as possible. As shown in the above examples, the methods and the tabulated substituent constants are thought to be sufficient for QSAR analysis in most cases. Although the precision of biological data is of primary importance for QSAR analysis, it is costly and time-consuming to establish precise dose-response relationships for every congener. Therefore, a strategy is needed to reduce the amount of biological work as well as to accelerate the project research. In the first example, we converted the fixed dose activity data so t h a t they are appropriately utilizable in regression analysis after confirming that the log(dose)-response curves of some congeners are parallel in certain concentration ranges. In the second example, compounds were classified into two groups according to the fixed dose data and a rough analysis using the ALS method was performed first to examine structural requirements for exhibition of activity. For some potent compounds, more precise dose-response data were measured and rendered to the Hansch-Fujita analysis to establish factors enhancing the potency. The combined results of these two QSAR procedures were used to predict the optimal structure. In the first two examples, we examined QSAR analyses repeatedly at each step using biological activity data for a smaller number of compounds to design additional compounds for subsequent syntheses. Such repetitions of the cycle of synthesis-biological evaluation-analysis gradually clarified the structural requirements for exhibiting potent

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activity, and finally, structure-activity data were summarized in one equation. Moreover, insights into structure-activity relationships gained quantitatively were extrapolated and transposed successfully to determine new lead structures (lead evolution). Thus, we found two new drug candidates more quickly and efficiently than before. The compounds, KB-2796 and KB-5492, are now undergoing extensive clinical trials. In the third example, QSAR analyses were performed after the project was over. The results confirmed that the candidate selection was valid. If the QSAR procedure had been used in the course of the project research, a much smaller number of compounds would have been needed to obtain the same information. It is by no means an exaggeration to say that QSAR analyses helped us to reduce the time required as well as the cost of the new drug research by facilitating rational and speedy decision-making. ACKNOWLEDGEMENT

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