Bioorganic & Medicinal Chemisny Letters, Vol. 4, No. 21. pp. X21-2626, 1994 Copyright CO 1994 Elsevier Science Ltd

Printed in Great Britain. All rights mmv.d 0960-894X/94 S7.OOKI.00

0960-894X(94)00388-2

SYNTHESIS AND BIOLOGICAL ACTIVITY OF A POTENT

ANTIPLATELET 7-AMINOFUROCHROMONE

Joel Morris*, Donn G. Wishka, William R. Humphrey, Alice H. Lin, Ann L. Wiltse,

Christopher W. Benjamin, Robert R. German, and Ronald J. Shebuski

Upjohn Laboratories, The Upjohn Company, Kalamazoo, Michigan 49001

Abstract: The synthesis of a series of rigid analogs designed to explore the conformational requirements associated with the related 2-aminochromone 1 led to the identification of class of novel antiplatelet 7- aminofumchromes 2. A potent derivative 13 was further evaluated with regard to mechanism of action as well as in a canine model of platelet dependent thrombus formation.

The search for pharmacological inhibitors of blood platelet aggregation has led to the identification of

the 2aminochromones as a new class of antiplatelet agents.‘. * We have recently described the synthesis and

biological evaluation of a series of 2morpholinochromones as inhibitors of ADP-induced platelet aggregatiou3

Moreover, significant efficacy in a canine model of coronary, platelet-dependent thrombus formation was

demonstrated for the 7-(4methyl-l-piperazinyl)ethyloxy derivative, 1. At a dose of 300 pg/kg (plus a constant

infusion of 30 @kg/min), 1 was completely effective in eliminating the platelet dependent cyclic flow

reductions (CFR’s) in this model. As part of an effort to examine the conformational requirements for activity

associated with thii class of compounds, a series of rigid analogs were targeted for synthesis and biological

evaluation. In this letter we describe the preparation of these 7aminofurochromones and their dihydro

derivatives 2 in which the aminoethyloxy side chain of 1 has been appended to the 6-position of the chromone

ring. This effort has led to the identification of the furochromone derivative 13 as a potent antiplatelet agent.

The 7aminofurochromones 2a were prepared starting with the commercially available

hydroxyacetophenone 3 according to the outline in Scheme I. Treatment of the corresponding 5iodo-4-

2621

2622 J. MORRIS et al.

acetyloxy derivative of 3 with boron nifluoride ethereate afforded the BFr complex 4. Reaction of 4 with 4-

morpholine dichloromethyleniminium chloride followed by hydrolysis of the resulting intermediate led to 2-

aminochrome 5 (53%).’ Elaboration of the iodophenol of 5 to benzofuran 6 with propargyl alcohol and Cu10

proceeded in 68% yiekLs Chlorination of 6 followed by displacement with a series of cyclic amines provided

the 7-furochromones 2a (44-6796).

Scheme I

0 1. Hl4, 1s 2. AcCl

HO 3. BF30Et2

CH3 56% f-J-‘3 3. LiOH

3

&N+& -’

0

/

HO;

5

HC=C-CH,OH cup0

0 68%

1. SOCl, / ’

2. NR:, O\‘O’N?

CHa 0 C’43 LO

2a 6

The 2-atninochromone 7 (available from 3 using related phosgeniminium salt chemistry)’ was the

source for production of the corresponding dihydro 7-furochromones 2b (Scheme II). Alkylation of 7 with

ally1 bromide (+ 877%) followed by Claisen rearrangement gave the phenol 9 (70%). Iodocyclization of 9 in

the presence of MeSCrH produced the dihydrofuran derivative 10 in 66% yield. The use of acid in this

reaction served to prevent over iodination of the product. Reaction of 10 with a group of cyclic amines as

above afforded 2b (40-49%).

Scheme II

HO

/ 12. MeS4H

MeCN, rt

R2N&Q A

CH3 0

,dN;

CH3 0 2b 10

Antiplatelet 7-aminofurochromone 2623

The 7-aminofuroch.romones 2 prepared accordingly were tested for their ability to inhibit ADP-induced

human platelet aggregation using 2-aminochmmone 1 as a control (Table D3 Rigidiflcation of the

aminoethyloxy side chain of 1 in the form of 7aminofurochromone 11 resulted in preservation of the

a&platelet activity (Table I). The corresponding dihydro derivative 12 proved to be approximately half as

potent as 1 was did the pyrrolidine and piperidine analogs 15 and 16. Interestingly, both of the thiomorpholine

substrates showed a sign&ant increase in potency relative to 1, particularly the benxofuran derivative 13 (I&

0.35 pM).

Table L Structures and Inhibitory Activity against ADP-Induced Human Platelet Aggregation.’

compd NR, A2.3 IC, (PM)

11 4-methylpiperaxine a 9.1 f 4.7

12 Cmethylpiperaxine b 5.6 + 1.7

13 thiomorpholine a 0.35 * 0.13

14 thiomorpholine b 3.6 + 2.4

15 pyrrolidine a 15.4 + 1.6

16 piperidine b 6.2 + 3.5

All values are mean & SD. b ICsa’s obtained for compd 1 ranged from 1.6-8.6 pM.

Relative Potencyb

0.9

0.5

4.5

2.4

0.5

0.5

Compound 13 was further examined in a canine model of coronary, platelet-dependent thrombus

formation.6 In this model, cyclical declines in blood flow (CFRs) occur spontaneously and periodically after

placement of an obstructive cylinder on the coronary artery. This 7aminofurochromone was extremely

effective in this model, completely blocking the in vivo platelet thrombus formation (CPP rating of 3) at a dose

of 100 pg/kg (plus a constant infusion of 10 pgkg/min) (Table II). A lower dose of 30 @kg (Plus 3

lq&/min) was somewhat effective. In addition, the effect of 13 on blood pressure and heart rate was also

assessed (Table II). Antithrombotic doses of 13 induced transient hypotension in the dogs and was

accompanied by reflex tachycardia. In an attempt to achieve antiplatelet activity with minimal hemodynamic

effects, low doses (3-10 pg/kg/min) of 13 were delivered via a prolonged, constant infusion. Although

2624 J. MORRIS et al.

minimized, hypotension and tachycardia induced by antithrombotic doses of 13 were not eliminated by the

change in drug delivery. These results with 7-aminofurochromone 13 were found in contrast to results

obtained with 2-aminochromone 1 where the compound was free of hemodynamic effects at the effective

antithrombotic dose.’

Table II. Effect of 13 on Cyclical Flow Reductions Induced by Intracoronary Platelet Aggregation in

Stenosed Canine Coronary Arteries.’

I.V. Dose CFR Rating Onset Timeb

(O-3) (min)

10 lwg

+ 1 pg/kghin~ 1 10

30 @kg

+ 3 p&JJmin=~’ 2 1

I I

10 @kg/ming 3 8

’ See experimental section of ref. 3 for a description c

Offset Time’

(min)

Mean Arterial

Pressure’

(% of

Control)

Heart Rated

(% of

Contrd)

3 99 106

34 88 112

>60 74 116

27 93 104

36 86 116

the CFR rating system. n = 1 unless otherwise

indicated. b Time interval between the start of drug treatment and the first change in the CFR pattern of blood

flow. ’ Time interval between the end of drug treatment and the restoration of the CFR pattern to control

levels. d Value at 25 min infusion time point. ’ n = 2. ’ 30 min. 8 60 min.

Compounds of in the 2-aminochromone class have previously been shown to inhibit platelet

aggregation through the elevation of cAMP.*~* ’ The 7-aminofurochromone 13 was also evaluated in this

regard. In a dose dependent manner, 13 was found to inhibit thrombin-stimulated aggregation as well as

increase CAMP levels in washed platelets (Table III). As a potent activator of platelet adenylate cyclase,

prostacyclin (PGI9 produces a marked increase in CAMP levels in washed platelets (Table IV).’ The

combination of 0.3 PM 13 and 1 q/ml PGI2 was found to synergystically increase CAMP levels to 163.1 _+ 2.7

pmol CAMP / 5 x lo* platelets. These data suggest the 13 is stimulating CAMP levels by another mechanism

other than the activation of adenylate cyclase, possibly by the inhibition of CAMP-phosphodiesterase (CAMP-

Antiplatelet 7aminofurochromone 2625

PDE) activity. To test this hypothesis, the effect of 13 on CAMP-PDE was assessed in platelet lysates. In a

dose dependent manner, 13 was found to inhibit platelet low km CAMP-PDE activity with an ICsO of 1.5 uM

(Pigure I). Thus the increase in CAMP stimulated by 13 can be attributed to the inhibition of platelet CAMP-

PDE activity.

Table III. Effect of 13 on CAMP Levels in and Aggregation of Thrombin-Stimulated Washed Human

Platelets.

Conditions CAMP (pmol I5 x 10’ platelets) % Aggregation

DMSO vehicle 10.3 -+ 0.7 0

0.25 u/ml thrombm control 10.0 * 0.3 100

0.1 l.tM 13 13.2 _+ 1.6 14* 10

0.3 pM 13 25.7 it: 1.6 0

1 w 13 36.1 k 0.4 0

Table IV. Effect of 13 on PGJ-Stimulated CAMP in Washed Human Platelets.

Conditions CAMP (pmol / 5 x 10’ platelets)

Basal 10.3 * 0.7

1 rig/ml PGI, 85.0 + 5.5

0.3 l.tIvI 13 25.7 + 1.6

0.3 pM 13 + 1 q/ml PGI, 163.1 + 2.7

In summary, the synthesis and biological evaluation of a series of antiplatelet 7aminofurochmmones

has been described. The thiomorpholine derivative, 13, proved extremely potent in a canine model of platelet

dependent thrombus formation. The hemodynamic effects associated with this series of compounds appear to

be due to their ability to inhibit platelet CAMP dependent phosphodiesterase leading to elevated levels of

CAMP.

2626 J. MORRIS et al.

Figure 1. Inhibition of Human Platelet Cytosolic CAMP-PDE Activity by 13.

120 r

E :: 1 , , ,,,,,,, , , ,,( ,,,, , ( ,,,,,,, , , ,,,,(,, , , ,,,,,,, , ( ,,,, ul

0.01 1 .o 10 100 0.1

References 13 [PM1

1. Gammill, R. B.; Judge, T. M.; Morris, J. WO 90/0692 1.

2. (a) Mazzei, M.; Balbi. A.; Roma, G.; Di Braccio, M.; Leoncini, G.; Buzzi, E.; Maresca, M. Eur. J. Med

Chem. 1988,23, 237-242. (b) Mazzei, M.; Sottofattori, E, Di Braccio, M.; Balbi, A.; Leoncini, G.; Buzzi, E.;

Maresca, M. Eur. J. Med. Chem. 1990, 25, 617-622. (c) Leoncini, G.; Maresca, M.; Colao, C.; Buzzi, E.;

Mazzei, M. Cell Biochem. Function 1991, 9, 79-85. (d) Leoncini, G.; Maresca, M.; Colao, C.; Buzzi, E.;

Mazzei, M.; Balbi, A. Pharm. Res. 1991,23,139-148.

3. Morris, J; Wishka, D. G.; Lin, A. H.; Humphrey, W. R.; Wit&e, A. L.; Gammill, R. B.; Judge, T. M.; Bkaha, S.

N.; Olds, N. L.; Jacob, C. S.; Bergh, C. L.; Cudahy, M. M.; Williams, D. J.; Nishizawa, E. E.; Thomas, E. W.;

German, R. R.; Benjamin, C. W.; Shebuski, R. J. J. Med. Chem. 1993,36,2026.

4. Morris, J.; Wishka, D. G.; Fang, Y. J. Org. Chem. 1992,57,6502-6508.

5. Doad, G. J. S.; Barhrop, J. A.; Petty, C. M.; Owen, T. C. TenuhedronLen, 1989, 1597-1600.

6. Bush, L. R ; Shebuski, R. J. FASEB, 1990,4,3087-3098.

7. Benjamin, C. B.; Lin, A. H.; Morrk, J.; Wishka, D. G.; German, R. R. JPET, 1993,265,457-462.

8. German, R. R.; Bunting, S.; Miller, 0. V. Prostuglundins, 1977,13,377-388.

(Received in USA 22 August 1994; accepted 3 October 1994)