Pergamon Bioorganic & Medicinal Chemistry Letters, Vol. 5, No. 7, pp. 699-704, 1995

Copyright © 1995 Elsevier Science Ltd Printed in Great Britain. All rights reserved

0960-894X/95 $9.50+0.00 0960-894X(95)00097-6

SYNTHESES OF ¢x-, I3-MONOGLYCOSYLCERAMIDES AND FOUR DIASTEREOMERS OF AN cx-GALACTOSYLCERAMIDE

Masahiro Morita, Takenori Natori, Kohji Akimoto, Tatsushi Osawa, Hideaki Fukushima,

and Yasuhiko Koezuka*

Pharmaceutical Research Laboratory, Kirin Brewery Co., Ltd.

3 Miyahara-cho, Takasaki-stu', Gunma 370-12, Japan

A b s t r a c t : To examine antitumor activities of monoglycosylceramide, we synthesized ~-, ~- galactosylceramides and ct-, I~-glucosylceramides which have the same ceramide portion, and four kinds of diastereomers of the ceramide portion in an ct-galactosylceramide.

Agelasphines (AGLs) have been isolated from an extract of the marine sponge, Agelas mauritianus, as

active substances in the course of screening of antitumor agents. 1,2 Since sufficient amounts of the four kinds

of AGLs (AGL-7a, AGL-9b , AGL.11, and AGL-13, Fig. 1) having ct-galactosylceramide (ct-GalCer)

structures and AGL-10 which has a 13-glucosylceramide (l~-GluCer) structure were obtained, we examined the

biological activities of five compounds using several assay systems, and found that ct-GalCers showed stronger

antitumor activities against B 16-bearing mice and stirnulatory effects of lymphocyte proliferation on allogeneic

mixed lymphocyte reaction (MLR) than 13-GluCer.

HO - - O H OH OH 0 ~ 0

HO ~ (CH2)x-CH 3 / OH ~ (CH2h-CH 3

HO! ~ QH H ~ O ~ - ' ~ . , s ~ R

OH HO OH

X R X R

AGL-7a 21 -(CH2)I 1-CH3 AGL-10 21 , . , ~ . ~ j (CH2)6-CH 3 AGL-9b 21 -(CH2)I 1-CH(CH3)2 AGL-11 21 -(CH2)n-CH(CH3)-C2H5 CH 3 AGL-13 22 -(CH2)11-CH(CH3)-C2H5

Fig. 1. Structures of AGL-7a , AGL-9b , A G L - I I , AGL-13, and AGL-10.

699

700 M. MORITA et al.

These findings suggested that not only the manner of combination between sugar and ceramide but also

the type of sugar combining to ceramide greatly affects the biological activities of monoglycosylated ceramides

(MonoCers). To confirm this possibility, we synthesized four kinds of MonoCers (AGL-517, 3 AGL-562, 4

AGL-563, 5 and AGL-564, 6) which have the same ceramide and their structures are shown in Fig. 2.

R I • . . . . OH 0 0

R1 OH R 2 H O ~ ~ HN,¢~ (CH2) 12CH3 f ~ . - v ~ (CH2) 12CH3

HOo ~ (CH2)I3CH 3 R2 ~ O ~ ' ~ (CH2) 13CH3 OH HO OH

AGL-517; RI=OH, R2=H AGL-564; RI=OH, R2=H AGL-563; RI=H, R2=OH AGL-562; RI=H, R2=OH

O O H.O ~ . OH II

[ ~ 0 x HN I ' ~ (CH2)24CH3 HN " ~ (CH2)24CH3 - o . -

HO, " - o -7 -

O ¢ ~ " ~ (CH2) 13CH3 H O ~ O ~ (CH2) 13CH3 OH HO OH

KRN7000 AGL-583

Fig. 2 Structures of AGL-517, AGL-564, AGL-563, AGL°562, KRN7000,

and AGL-583.

As shown in Scheme 1, AGL-517 (a-GalCer) and AGL-563 (u-GluCer) were synthesized from a

ceramide (1) 7 under Mukaiyama's ct-glycosidation condition 8 and following hydrogenolysis.

S c h e m e 1

0

R 2 ~ O + .= BnO ~ F H O ¢ , , ~ (CH2) 13CH 3

BnO OH

2; RI=OBn, R2=H 1 3; RI=H, R2=OBn

RI .....- OBn

B n O ~ - ~ HN f ~. (CHz) 12CH3 BnO ! .---

O @ ~ ~'~ (Ctt2) 13CH 3 OH

4; RI=OBn, R2=H 5; RI=H, R2=OBn

RIj.- OH

H O ~ HN_ ~ (CH2)I2CH3 HO ! -

O , ~ (CH2) 13CH3 OH

AGL-517; RI=OH, R2=H AGL-563; R1--H, R2--OH

(a) SnC12, AgC104, MS4A / THF; (b) H 2, Pd-BaSO 4 / THF

0c-, [~-Monoglycosylceramides 701

Scheme 2

l • l ~ OAc 0

AcO x - - " ~ " " ~ "" OAc HO " (CH2) 13CH3 AcO

OH

a

O R I ~ R 3 ] \ ,-, HN- "(CH2)12CH3

R3 OH

6; R1--OAc, R2=H 1 7; RI=H, R2--OAc

["-" 8 ; RI=R3=OAc, R2=H h AGL-564; RI=R3=OH, R2=H

b ~ 9 ; RI=H, R2=R3--OAc AGL-562; R I=H, R2=R3--OH

(a) TMSOTf, MS4A / CH2C12 ; (b) NaOMe / MeOH-THF

AGL-564 (13-GalCer) and AGL-562 (13-GluCer) were synthesized as follows: the same ceramide (1)

was treated with penta-O-acetyl sugars (6, 7) in the presence of trimethylsilyltrifrate (TMSOTf) and

molecularsives 4A 9 to afford the desired 13-glycosides (8, 9). Deacetylation of 8 and 9 by treatment with

sodium methoxide to give AGL-564 and AGL-562 (Scheme 2).

Futhermore we synthesized KRN700010 (a-GalCer) and AGL-58311 (13-GalCer) which have the

different ceramide portion 7 from AGL-517 using the same glycosidation methods as described for AGL-517

and AGL-564, and their structures are shown in Fig. 2.

Scheme 3

NH2

H O ~ , ~ , , ~ (CH2)I 1CH3 OH

lOa; (2S, 3S) lOb; (2R, 3R) Ilk:; (2S, 3R) lOd; (2R, 3S)

HN L (CH2)12CH 3 --- H O ~ (

a CH2)11CH3 b OH

l la; (2S, 3S) l ib; (2R, 3R) lle; (2S, 3R) lld; (2R, 3S)

BnO ~ OBn o

BnO , ~ HN (CH2)12CH 3

O ~ J ~ "~ (CH2)I 1CH3 OH

12a; (2S, 3S) 12b; (2R, 3R) 12c; (2S, 3R) 12d; (2R, 3S)

HO ~ OH o

~, HO HN,~,,,~ (CH2) 12CH3

O ~ (CH2) 11CH3 OH

AGL-555; (2S, 3S ) AGL-556; (2R, 3R) AGL-558; (2S, 3R ) AGL-559; (2R, 3S)

(a) p-nitrophenyl tetradecanoate, DMAP / THF; (b) 2, SnC12, AgC104, MS4A / THF; (c) H 2, Pd-BaSO4 / THF

702 M. MORITA et al.

H~ ~ OH

e o . ~ HN.J~ ca cx ~ 0

~ (CH2)I 1CH3 OH

AGL-555; (2S, 3S)

HO ~ OH

o 0 -

~ " ' ~ (CH2)I 1CH3 OH

AGL-558; (2S, 3R)

- -~O [ 1 1 (C 2)12CH3

O , , , ~ " - (CH2hICH3 OH

AGL-556; (2R, 3R) HO OH

~0 I H~I" "(CH2)12CH3

O ~ , , , , " ~ (CH2)I 1CH3 OH

AGL-559; (2R, 3S)

Fig. 3. Structures of AGL-555, AGL-556, AGL558, and AGL-559.

In addition, we previously reported 7 that, the structure of the ceramide portion in a-GalCer greatly affects

their biological activities, i.e., (1) the longer fatty acid in ceramide moiety shows more marked antitumor

activity in the range of C14 - C26, (2) the optimal length of long chain base in ceramide is C18 in the range of

Cll - C20, and (3) 3-hydroxyl group plays an important role in biological activities. However, the biological

activities of the diastereomers of the ceramide portion have not been reported yet. To examine their biological

activities, we synthesized four kinds of diastereomers having ct-GalCer structures starting from

dihydrosphingosines (10a-d) 12 by previously reported method 2, 7 (Scheme 3). Fig. 3 shows the structures of

four diasteromers which are AGL-555 (2S, 3S) 13, AGL-556 (2R, 3R) 14, AGL-558 (2S, 3R) 15, and

AGL-559 (2R, 3S) 16.

Acknowledgement: We thank Dr. Hiroyuki Matsuda, Central Research Laboratory, Takasago International

Corporation, for generous gift of four kinds of dihydrosphingosines.

0~-, ~-Monoglycosylceramides 703

References and Notes

1. Natori, T.; Koezuka, Y.; I-Iiga, T. Tetrahedron Lett. 1993, 34, 5591-5592.

2. Natori, T.; Morita, M.; Akimoto, K.; Ko~zuka, Y. Tetrahedron, 1994, 50, 2771-2784.

3. AGL-517 physical data: [aiD24 = +57.8 ° (c 1.0, pyridine); m.p. 159.0-161.0 °C; FDMS m/z 674 M+; 1H NMR (500 MI-Iz, CsDsN) 8 (ppm) 8.52 (IH, d, J = 8.6 Hz), 5.45 (1H, d, J = 3.7 I-Iz), 4.69-4.76 (1H, m), 4.63-4.68 (lI-I, m), 4.40-4.58 (6I-I, m), 4.36 (1H, dd, J = 5.5, 10.0 Hz), 4.25-4.31 (1H, m), 2.48 (2H, t, J = 7.0 Hz), 1.80-1.95 (4H, m), 1.52-1.62 (1H, m), 1.18-1.43 (45H, m), 0.88 (6H, t, J = 6.7 Hz): Anal. C38H75NO 8 (C, H, N). calcd. 67.72, 11.22, 2.08, found. 67.41, 11.28, 2.03.

4. AGL-562 physical data: [a]D 23 = -3.1 ° (c 0.85, pyridinc); m.p. 159.0-162.5 °C; FDMS m/z 675 (M+I)+; 1H NMR (500 MHz, C5D5N) 8 (ppm) 8.40 (1H, d, J = 8.5 I-Iz), 4.97 (1H, d, J = 7.9 Hz), 4.78 (1H, dd, J --4.9, 10.4 Hz), 4.68-4.75 (1H, m), 4.55 (1H, d, J = 11.6 Hz), 4.36 (1H, dd, J = 5.5, 11.6 Hz), 4.18-4.30 (4H, m), 4.06 (1H, t, J =7.9 Hz), 3.94-3.98 (1H, m), 2.47 (2H, t, J = 7.3 Hz), 1.78- 1.99 (4H, m), 1.52-1.61 (lI-I, m), 1.06-1.44 (45H, m), 0.88 (6H, t, J = 7.0 Hz): Anal. C38H75NO8 (C, H, N). calcd. 67.72, 11.22, 2.08, found. 67.88, 11.39, 1.92.

5. AGL-563 physical data: let]D23 = +57.4 ° (c 0.46, pyridinc); m.p. 162.0-163.0 °C; FDMS m/z 675 (M+I)+; 1H NMR (500 MI-Iz, CsDsN) 6 (ppm) 8.43 (1H, d, J = 8.6 Hz), 5.40 (1H, d, J = 3.1 Hz), 4.65-4.72 (lI-I, m), 4.22-4.60 (TH, m), 4.10-4.21 (2H, m), 2.43 (2H, t, J = 7.3 Hz), 1.76-1.93 (4H, m), 1.48-1.56 (1H, m), 1.10-1.42 (45H, m), 0.88 (6H, t, J = 6.7 Hz): Anal. C38H75NO8 (C, H, N). calcd. 67.72, 11.22, 2.08, found. 67.84, 11.27, 1.96.

6. AGL-564 physical data: [Ct]D24 = +7.3 ° (c 0.93, pyridine); m.p. 175.0-177.0 °C; FDMS m/z 675 (M+I)+; 1H NMR (500 MHz, CsD5N) 8 (ppm) 8.31 (1H, d, J = 9.2 Hz), 4.88 (1H, d, J = 7.3 Hz), 4.80 (1H, dd, J = 4.9, 10.4 I-Iz), 4.64-4.70 (1H, m), 4.55 (1H, d, J = 3.1 Hz), 4.49 (1H, dd, J = 7.9, 9.2 Hz), 4.40-4.45(2H, m), 4.10-4.25 (3H, m), 4.06 (1H, t, J = 5.8 Hz), 2.44 (2H, t, J = 7.3 Hz), 1.76- 1.92 (4H, m), 1.48-1.57 (lI-I, m), 1.10-1.42 (45H, m), 0.88 (6H, t, J = 6.7 Hz): Anal. C38H75NO8 (C, H, N). calcd. 67.72, 11.22, 2.08, found. 67.94, 11.37, 1.90.

7. Morita, M.; Motoki, K.; Akimoto, K.; Natori, T.; Sakai, T.; Sawa, E.; Yamaji, K.; Kobayashi, E.; Fukushima, H.; Koezuka, Y. Manuscript submitted for publication.

8. Mukaiyama, T.; Mural, Y.; Shoda, S. Chem. Lett., 1981, 431-432.

9. Ogawa, T.; Beppu, K.; Nakabayashi, S. Carbohydr. Res. 1981, 93, C6-C9.

10. KRN7000 physical data: [CC]D 23 = +43.6 ° (c 1.0, pyridine); m.p. 189.5-190.5 °C; negative FABMS m/z 857 (M-H)- ; 1H NMR (500 MHz, C5D5N) 8(ppm) 8.47 (1H, d, J = 8.5 Hz), 5.58 (1H, d, J = 3.7 Hz), 5.27 (1H, m), 4.63-4.70 (2H, m), 4.56 (1H, m), 4.52 (1H, t, J = 6.1 Hz), 4.37-4.47 (4H, m), 4.33 (2H, m), 2.45 (2H, t, J =7.3 Hz), 2.25-2.34 (1H, m), 1.87-1.97 (2H, m), 1.78-1.85 (2H, m), 1.62-1.72 (1H, m), 1.26-1.45 (66H, m), 0.88 (6H, t, J = 6.7 I-Iz); Anal. C50H99NO9 (C, H, N). calcd. 69.97, 11.63, 1.63, found. 69.77, 11.65, 1.53.

11. AGL-583 physical data: [C~]D24 -- -3.5 ° (c 0.81, pyridinc); m.p. 152.0-154.0 °C; negative FABMS m/z 857 (M-H)- ; 1H NMR (500 MHz, C5D5N) 8(ppm) 8.44 (1H, d, J = 8.5 Hz), 5.11-5.17 (1H, m), 4.91 (1H, d, J = 7.3Hz), 4.81 (1H, dd, J = 5.5, 10.4 Hz), 4.54 (1H, d, J = 3.1 Hz), 4.35-4.50 (6H, m), 4.17-4.24 (1H, m), 4.14 (1H, dd, J = 3.7, 9.8 Hz), 4.04 (1H, t, J = 6.1 Hz), 2.42 (2H, t, J =7.3 Hz), 2.17-2.21 (1H, m), 1.63-1.92 (6H, m), 1.05-1.50 (64H, m) 0.86-0.89 [6H, (0.881, t, J = 6.7 Hz), (0.876, t, J = 6.7Hz)]; Anal. C50H99NO9 (C, H, N). calcd. 69.97, 11.63, 1.63, found. 70.12, 11.70, 1.51.

12. Matsuda, H.; Yamamoto, T.; Sato, T. Japanease Patent Laid-Open Publication No. 80617/1994.

704 M. MOR1TA et al.

13.

14.

15.

16.

AGL-555 physical data: [Ct]D23 +52.5 ° (c 0.75, pyridine); m.p. 148.5-152.5 °C; FDMS m/z 646 M+; IH NMR (500MHz, CsD5N) 8 (ppm) 8.10 (IH, d, J = 8.5 Hz), 5.46 (1H, d, J = 3.7 Hz), 4.75-4.82 (1H, m), 4.66 (1H, dd, J = 3.7, 9.8 Hz), 4.34-4.56 (7H, m), 4.12 (1H, t, J = 6.1 Hz), 4.07 (1H, dd, J = 6.1, 9.8 Hz), 2.49 (2H, t, J = 6.5 Hz), 1.63-1.92 (4H, m), 1.50-1.60 (1H, m), 1.20-1.42 (41H, m), 0.88 (6H, t, J = 6.7 Hz): Anal. C36HT1NO8 (C, H, N). calcd. 66.94, 11.08, 2.17, found. 67.01, 11.28, 1.93.

AGL-556 physical data: [Or]D24 +80.7 ° (c 0.27, pyridine); m.p. 149.0-150.5 °C; FDMS m/z 646 M+; 1H NMR (500MHz, CsDsN) 8 (ppm) 8.04 (1H, d, J = 8.6 Hz), 5.49 (1H, d, J = 3.7 Hz), 4.77-4.82 (1H, m), 4.68 (1H, dd, J = 3.7, 9.8 Hz), 4.65 (IH, bd, J = 2.4 Hz), 4.36-4.58 (6H, m), 4.16 (1H, dd, J = 6.7, 10.4 Hz), 2.50 (2H, t, J = 7.3 Hz), 1.64-1.92 (4H, m), 1.47-1.57 (1H, m), 1.20-1.42 (41H, m), 0.88 (6H, t, J -- 7.0 Hz): Anal. C36H71NO8 (C, H, N). calcd. 66.94, 11.08, 2.17, found. 67.12, 11.34, 2.09.

AGL-558 physical data: [a]D24 +74.3 ° (c 0.35, pyridine); m.p. 153.5-156.0 °C; FDMS m/z 646 M+; 1H NMR (500MHz, C5D5N ) 8 (ppm) 8.52 (1H, d, J = 8.6 Hz), 5.47 (1H, d, J -- 3.7 Hz), 4.71-4.78 (1H, m), 4.67 (IH, dd, J = 3.7, 9.8 Hz), 4.34-4.60 (7H, m), 4.26-4.32 (1H, m), 2.48 (2H, dt, J = 1.2, 7.3 Hz), 1.80-1.95 (4H, m), 1.52-1.62 (1H, m), 1.20-1.42 (41H, m), 0.87 (6H, t, J = 6.8 Hz): Anal. C36HT1NO8 (C, H, N). calcd. 66.94, 11.08, 2.17, found. 66.99, 11.24, 2.01.

AGL-559 physical data: [~t]D24 +62.0 ° (c 0.50, pyridine); m.p. 145.0-147.0 °C; FDMS m/z 646 M+; 1H NMR (500MHz, CsDsN) 8 (ppm) 8.40 (1H, d, J = 8.5 Hz), 5.47 (1H, d, J = 3.7 Hz), 4.66-4.76 (3H, m), 4.10-4.62 (7H, m), 2.48 (2H, dt, J = 1.8, 7.3 Hz), 1.73-2.00 (4H, m), 1.52-1.62 (1H, m), 1.20- 1.42 (41H, m), 0.88 (6H, t, J = 6.7 Hz): Anal. C36H71NO8 (C, H, N). calcd. 66.94, 11.08, 2.17, found. 67.01, 11.30, 2.13.

(Received in Japan 19 January 1995; accepted 16 February 1995)