Chiral dihydroxylation of acronycine: Absolute configuration of natural cis-1,2-dihydroxy-1,2-dihydroacronycine and cytotoxicity of (1R,2R)- and (1S,2S)-1,2-diacetoxy-1,2-dihydroacronycine

Article abstract of DOI:10.1021/np980420q

Sharpless asymmetric dihydroxylation of acronycine (1) gave (1R,2R)- 1,2-dihydroxy-1,2-dihydroacronycine (2) and (1S,2S)-1,2-dihydroxy-1,2- dihydroacronycine (3), which allowed determination of the absolute configuration of natural cis-1,2-dihydroxy-1,2-dihydroacronycine as 1R,2R. The cis isomer had been previously isolated from various Sarcomelicope species. Benzylic reduction of isomers 2 and 3 gave the alcohols 4 (2R) and 5 (2S), respectively. Acetylation of 2 and 3 afforded the corresponding esters 6 and 7. No significant difference of cytotoxicity was observed between these (1R,2R)- and (1S,2S)-enantiomers and the recently described, highly active racemic cis-1,2-diacetoxy-1,2-dihydroacronycine, when tested against L-1210 cells in vitro.

Full text of DOI:10.1021/np980420q

490
J . Nat. Prod. 1999, 62, 490-492
Ch ir a l Dih yd r oxyla tion of Acr on ycin e: Absolu te Con figu r a tion of Na tu r a l
cis-1,2-Dih yd r oxy-1,2-d ih yd r oa cr on ycin e a n d Cytotoxicity of (1R,2R)- a n d
(1S,2S)-1,2-Dia cetoxy-1,2-d ih yd r oa cr on ycin e
Nadine Costes,† Sylvie Michel,† Franc¸ois Tillequin,*,† Michel Koch,† Alain Pierre´,‡ and Ghanem Atassi‡
Laboratoire de Pharmacognosie de l’Universite´ Rene´ Descartes-UMR 8638, Faculte´ des Sciences Pharmaceutiques et
Biologiques, 4 avenue de l’Observatoire, 75006 Paris, France, Institut de Recherches Servier, 11 rue des Moulineaux,
92150 Suresnes, France
Received September 28, 1998
Sharpless asymmetric dihydroxylation of acronycine (1) gave (1R,2R)-1,2-dihydroxy-1,2-dihydroacronycine
(2) and (1S,2S)-1,2-dihydroxy-1,2-dihydroacronycine (3), which allowed determination of the absolute
configuration of natural cis-1,2-dihydroxy-1,2-dihydroacronycine as 1R,2R. The cis isomer had been
previously isolated from various Sarcomelicope species. Benzylic reduction of isomers 2 and 3 gave the
alcohols 4 (2R) and 5 (2S), respectively. Acetylation of 2 and 3 afforded the corresponding esters 6 and
7. No significant difference of cytotoxicity was observed between these (1R,2R)- and (1S,2S)-enantiomers
and the recently described, highly active racemic cis-1,2-diacetoxy-1,2-dihydroacronycine, when tested
against L-1210 cells in vitro.
The acridone alkaloid acronycine (1), first isolated from
Acronychia baueri Schott (Rutaceae),1 was found to be
active against numerous solid tumors.2-5 Nevertheless,
clinical trials gave only poor results, due to both the low
water solubility and the moderate potency of acronycine.6
We have recently reported the synthesis of some (()cis-
1,2-dihydroxy-1,2-dihydroacronycine esters that were more
active in vivo than acronycine as antitumor drugs.7 Indeed,
these new acronycine derivatives have shown promising
antitumor activity, with a broadened spectrum and an
increased potency when compared with acronycine itself
on several tumor strains both in vitro and in vivo. Among
these esters, racemic cis-1,2-diacetoxy-1,2-dihydroacrony-
cine seems of particular interest because of its high activity
in vivo against P-388 leukemia and against the highly
resistant tumor C-38 colon carcinoma. Therefore, it was of
great interest, in a continuation of our work on the
acronycine series, to study the influence of the absolute
configuration on the biological activity of this compound.
In addition, the chiral synthesis of the two enantiomers of
cis-1,2-dihydroxy-1,2-dihydroacronycine should allow us to
determine the absolute configuration of this alkaloid, which
had been previously isolated in its optically active (-)-form
from several Sarcomelicope species.8,9 In this paper, we
report the enantioselective preparation of (1R,2R)-1,2-
dihydroxy-1,2-dihydroacronycine and (1S,2S)-1,2-dihydroxy-
1,2-dihydroacronycine, according to the Sharpless meth-
odology,10 which gives access to the corresponding diacetates.
The biological activity of these latter derivatives is evalu-
ated.
give an excess of the corresponding enantiomer (1S,2S)-
1,2-dihydroxy-1,2-dihydroacronycine (3).
The enantiomeric excess of the chiral diols 2 and 3,
determined by chiral HPLC, was 40% and 70%, respec-
tively. This technique was also used for the purification of
the enantiomers on a preparative scale.
To ensure that the absolute configuration of each diol
matched that predicted by Sharpless, we carried out a
benzylic reduction of the chiral diols 2 and 3 to the
corresponding homobenzylic alcohols, respectively, 4 and
5. This was achieved by the use of NaBH3CN in the
presence of ZnI2.11 The absolute configuration at C-2 of each
2-hydroxy-1,2-dihydroacronycine enantiomer was previ-
1
ously assigned in our laboratory by comparison of their H
and13C NMR spectra with published data for the related
diastereoisomeric glycosides of the angular hydroxydihy-
dropyranocoumarins (+)- and (-)-lomatin.12-14
Furthermore, the enantioselective synthesis of acrony-
cine diols enabled us to determine the absolute configura-
tion of the natural (-)cis-1,2-dihydroxy-1,2-dihydroacro-
nycine, previously isolated from the bark of Sarcomelicope
glauca Hartley15 and from the bark of Sarcomelicope
dogniensis Hartley,16 as 1R,2R.
Finally, the desired (1R,2R)-1,2-diacetoxy-1,2-dihydroa-
cronycine (6) and (1S,2S)-1,2-diacetoxy-1,2-dihydroacrony-
cine (7) were obtained in a good yield by treatment of cis-
diols 2 and 3 with excess acetic anhydride in pyridine.
The study of the biological properties of the enatiomeri-
cally pure acronycine derivatives 6 and 7 was carried out
in vitro on L-1210 leukemia. The results (IC50), reported
in Table 1, indicate there is no significant difference in
cytotoxicity between the two enantiomers (1R,2R)- and
(1S,2S)-1,2-diacetoxy-1,2-dihydroacronycine and the race-
mic cis-1,2-diacetoxy-1,2-dihydroacronycine.
Following the Sharpless methodology, osmium-catalyzed
asymmetric dihydroxylation (AD) of acronycine (1) was
performed using diphenylpyrimidine ligands involving
either dihydroquinine [(DHQ)2-PYR] or dihydroquinidine
[(DHQD)2-PYR] (Scheme 1).10 From the stereoselectivity
rules established by Sharpless for the AD reaction, it was
concluded that the use of the (DHQ)2-PYR ligand should
give an enantiomeric excess of (1R,2R)-1,2-dihydroxy-1,2-
dihydroacronycine (2) and that (DHQD)2-PYR ligand should
Exp er im en ta l Section
Gen er a l Exp er im en ta l P r oced u r es. Optical rotations
1
were obtained on a Perkin-Elmer 241 polarimeter. H NMR
[chemical shifts δ (ppm), J (Hz)] and 13C NMR spectra were
recorded at 300 and 75 MHz, respectively, using a Bruker AC-
300 spectrometer. Chemical ionization mass spectra (DICMS;
NH3, positive ion mode) and electronic impact mass spectra
(EIMS) were recorded on a Nermag R 10-10C spectrometer.
* To whom correspondence should be addressed. Tel.: 0033153739803.
Fax: 0033140469658. E-mail: tillequi@pharmacie.univ-paris5.fr.
† Laboratoire de Pharmacognosie de l’Universite´ Rene´ Descartes.
‡ Institut de Recherches Servier.
10.1021/np980420q CCC: $18.00
© 1999 American Chemical Society and American Society of Pharmacognosy
Published on Web 03/04/1999

Notes
J ournal of Natural Products, 1999, Vol. 62, No. 3 491
Sch em e 1a
for AD-mix-R or (DHQD)2-PYR for AD-mix-â (13.65 mg, 1 mol
%) were ground together to give a fine powder, then blended
into the bulk powdered ingredients (99.4% by wt) K3Fe(CN)6
(1.5 g, 3 equiv/mole of olefin) and K2CO3 (640 mg, 3 equiv),
producing a fine yellow powder. This powder was added rapidly
to a stirred mixture of 1:1 tert-butyl alcohol-H2O (18 mL) at
room temperature. After 15 min, methanesulfonamide (CH3-
SONH2) (295 mg, 2 equiv) and acronycine (500 mg, 1.55 mmol)
were added. The reaction mixture was maintained under
vigorous agitation over 48 h. Then, solid Na2S2O5 (932 mg)
was slowly added and the mixture stirred for 1 h. CH2Cl2 (40
mL) was added to the reaction mixture, and after separation
of the layers, the aqueous phase was further extracted with
the organic solvent (3 × 40 mL). The combined organic extracts
were washed with 2N KOH (50 mL), dried over anhydrous
sodium sulfate, and concentrated. The crude product obtained
was purified by flash chromatography; elution with CH2Cl2
gave unreacted acronycine, while further elution with CH2-
Cl2-MeOH, 97:3, provided cis-1,2-dihydroxy-1,2-dihydroacro-
nycine as a yellow powder (222 mg, 40%).
HP LC Con d ition s for th e An a lytica l Deter m in a tion of
th e En a n tiom er ic Excesses a n d for th e P r ep a r a tion of
En a n tiom er ica lly P u r e Sa m p les on a P r ep a r a tive Sca le.
Column PHENOMENEX: CHIREX (S)-indoline carboxylic acid
and (R)-1-R-naphthylethylamine (1/4′′ × 25 cm). Int. diam, 4.6
mm; eluent, n-heptane-CH2Cl2-EtOH: 130/40/30; flow rate,
1 mL/min; solutions, 1 mg/mL in tetrahydrofuran (1 µL
injected); detection, 275 nm.
Using chiral ligands (DHQ)2-PYR and (DHQD)2-PYR, the
ee of diols 2 (1R,2R) and 3 (1S,2S) was determined to be 40%
and 70%, respectively. Enantiomerically pure samples of 2 and
3 were obtained by repetitive semipreparative HPLC, per-
formed on the diol mixtures obtained from reaction with
(DHQ)2-PYR and (DHQD)2-PYR, respectively. HPLC purified
samples of 2, [R]20D -37.8° (c 0.5, MeOH), and 3, [R]20D +37.8°
(c 0.5, MeOH), were recrystallized from MeOH to afford yellow
needles: mp 232-234 °C; 1H NMR (DMSO-d6, 300 MHz) δ 8.04
(1H, dd, J ) 8, 2 Hz, H-8), 7.66 (1H, td, J ) 8,2 Hz, H-10),
7.41 (1H, dd, J ) 8, 1 Hz, H-11), 7.18 (1H, td, J ) 8,1 Hz,
H-9), 6.16 (1H, s, H-5), 5.05 (2H, m, H-1, OH-2), 4.55 (1H, d,
J ) 9 Hz, OH-1), 3.80 (3H, s, OMe), 3.77 (3H, s, NMe), 3.64
(1H, t, J ) 5 Hz, H-2), 1.42 (3H, s, Me), 1.39 (3H, s, Me);13C
NMR (DMSO-d6, 75 MHz) δ 176.7 (C-7), 162.1 (C-6), 160.2 (C-
4a), 150.1 (C-12a), 145.3 (C-11a), 133.7 (C-10), 126.7 (C-8),
125.5 (C-7a), 121.1 (C-9), 117.7 (C-11), 111.6 (C-6a), 104.4 (C-
12b), 95.0 (C-5), 78.6 (C-3), 71.2 (C-2), 65.1 (C-1), 56.8 (OCH3),
42.6 (NCH3), 26.2 [C-3(CH3b)], 23.3 [C-3(CH3a)]; EIMS m/z 355
[M•+], 337.
(2R)-2-Hyd r oxy-1,2-d ih yd r oa cr on ycin e (4) a n d (2S)-2-
Hyd r oxy-1,2-d ih yd r oa cr on ycin e (5). To a stirred solution
of 2 (60 mg, 0.17 mmol) in 1,2-dichloroethane (15 mL) at room
temperature were added solid zinc iodide (81 mg, 0.26 mmol)
and sodium cyanoborohydride (80 mg, 1,29 mmol). The reac-
tion mixture was stirred at room temperature for 48 h, filtered
through Celite, and washed with CH2Cl2 (40 mL). The com-
bined filtrates were evaporated to dryness, and the residue
was chromatographed (eluent: CH2Cl2-MeOH, 98:2, v/v) to
give unreacted diol 2 and 4 (20 mg, 35%): [R]20D -14.9° (c 0.4,
CHCl3) [lit.14 [R]20D -15° (c 0.4, CHCl3)] as a yellow amorphous
solid; 1H NMR (CDCl3, 300 MHz) δ 8.30 (1H, dd, J ) 8, 2 Hz,
H-8), 7.58 (1H, td, J ) 8, 2 Hz, H-10), 7.30 (1H, dd, J ) 8, 2
Hz, H-11), 7.20 (1H, td, J ) 8, 2 Hz, H-9), 6.26 (1H, s, H-5),
3.93 (3H, s, OMe), 3.82 (1H, dd, J ) 6, 5 Hz, H-2), 3.75 (3H, s,
NMe), 3.12 (1H, dd, J ) 16, 5 Hz, H-1a), 2.88 (1H, dd, J ) 15,
6 Hz, H-1b), 2.11 (1H, br s, D2O exch, OH-2), 1.62 (3H, s, Me),
1.46 (3H, s, Me); 13C NMR (CDCl3, 75 MHz) δ 177.55 (C-7),
160.64 (C-4a), 158.45 (C-6), 150.31 (C-12a), 145.55 (C-11a),
132.44 (C-10), 126.62 (C-8), 125.37 (C-7a), 121.51 (C-9), 116.18
(C-11), 110.92 (C-6a), 98.91 (C-12b), 94.90 (C-5), 77.42 (C-3),
69.23 (C-2), 55.87 (OCH3), 44.02 (NCH3), 31.44 (C-1), 25.19
[C-3(CH3b)], 21.43 [C-3(CH3a)]; DCIMS m/z 340 [M + H]+. In
a similar way, benzylic reduction of 3 gave 5 (22 mg, 38%);
[R]20 +15.2° (c 0.4, CHCl3) [lit.14 [R]20 +15° (c 0.4, CHCl3)].
a
Key: (i) K2OsO2, 2H2O, chiral ligand, K3Fe(CN)6, K2CO3, t-BuOH-H2O,
rt; (ii) NaBH3CN/ZnI2, ClCH2CH2Cl, rt; (iii) Ac2O, Py, rt.
Ta ble 1. Cytotoxic Activitya
compound
acronycine
IC50 (µM)
10.4
3.4
racemic cis-diacetate
6
7
3.1
3.7
a
Inhibition of L-1210 cell proliferation measured by the MMT
assay (mean of two values obtained in independent experiments).
HRMS were recorded on a Micromass ZAB2-SEQ spectrom-
eter. Flash chromatography was performed on Si gel, type 60
Å column chromatograph Chromagel, 35-70 µm, with an
overpressure of 300 mBars. Chromatography was performed
on Si gel, type 60 Å C. C. Chromagel, 20-45 µm. Acronycine
was prepared according to the method of Hlubucek.17
(1R,2R)-1,2-Dih yd r oxy-1,2-d ih yd r oa cr on ycin e (2) a n d
(1S,2S)-1,2-Dih yd r oxy-1,2-d ih yd r oa cr on ycin e (3). P r ep a -
r a t ion of t h e Ch ir a l Diols. Potassium osmate dihydrate
(K2OsO2), 2H2O (5 mg, 1 mol % of olefin), and (DHQ)2-PYR
D
D
(1R,2R)-1,2-Dia cetoxy-1,2-d ih yd r oa cr on ycin e (6) a n d
(1S,2S)-1,2-Dia cetoxy-1,2-d ih yd r oa cr on ycin e (7). A cooled

492 J ournal of Natural Products, 1999, Vol. 62, No. 3
Notes
mixture of Ac2O (0.5 mL, 5 mmol) and dry pyridine (0.5 mL)
was added to 2 (100 mg, 0.28 mmol) and the reaction mixture
stirred at room temperature for 24 h and poured into cold H2O
(10 mL). The precipitate obtained was filtered, washed with
H2O, and dried in vacuo under P2O5 to afford 6 (110 mg,
Refer en ces a n d Notes
(1) Hughes, G. K.; Lahey, F. N.; Price, J . R.; Webb, L. J . Nature 1948,
62, 223-224.
(2) Svoboda, G. H. Lloydia 1966, 29, 206-224.
(3) Svoboda, G. H.; Poore, G. A.; Simpson, P. J .; Boder, G. B. J . Pharm.
90%): [R]20 -67.2° (c 0.16, MeOH); 1H NMR (CDCl3, 300
Sci. 1966, 55, 758-768.
(4) Suffness, M.; Cordell, G. A. In The Alkaloids; Brossi, A., Ed.;
Academic: New York, 1985; pp 1-355.
D
MHz) δ 8.85 (1H, dd, J ) 8, 2 Hz, H-8), 7.64 (1H, td, J ) 8, 2
Hz, H-10), 7.26 (2H, m, H-9 and H-11), 6.55 (1H, d, J ) 5 Hz,
H-1), 6.31 (1H, s, H-5), 5.48 (1H, d, J ) 5 Hz, H-2), 4.00 (3H,
s, OMe), 3.63 (3H, s, NMe), 2.04 (3H, s, CH3CO-O-C1), 1.97
(5) Dorr, T. R.; Liddil, J . D.; Von Hoff, D. D.; Soble, M.; Osborne, C. K.
Cancer Res. 1989, 49, 340-344.
(6) Scarffe, H. J .; Beaumont, A. R.; Crowther, D. Cancer Treat. Rep. 1983,
67, 93-94.
(3H, s, CH3CO-O-C2), 1.56 (3H, s, Me), 1.47 (3H, s, Me); 13
C
NMR (CDCl3, 75 MHz) δ 177.4 (C-7), 170. 9 (CH3CO-O-C2),
165.3 (CH3CO-O-C1), 162.7 (C-6), 159.8 (C-4a), 149.3 (C-12a),
144.9 (C-11a), 132.8 (C-10), 126.9 (C-8), 125.6 (C-7a), 121.9
(C-9), 115.7 (C-11), 111.9 (C-6a), 97.5 (C-12b), 94.7 (C-5), 76.2
(C-3), 69.3 (C-2), 65.7 (C-1), 56.2 (C6-OCH3), 42.4 (NCH3), 24.4
[C-3(CH3b)], 23.4 [C-3(CH3a)], 20.9 (CH3CO-O-C2), 20.6 (CH3-
CO-O-C1); EIMS m/z 439 [M•+]; HREIMS m/z 439.1635 (calcd
for C24H25NO7, 439.1631). Acetylation of 3 (100 mg, 0.28 mmol)
(7) Elomri, A.; Mitaku, S.; Michel, S.; Skaltsounis, A.-L.; Tillequin, F.;
Koch, M.; Pierre´, A.; Guilbaud, N.; Leonce, S.; Kraus-Berthier, L.;
Rolland, Y.; Atassi, G. J . Med. Chem. 1996, 39, 4762-4766.
(8) Tillequin, F.; Michel, S.; Skaltsounis, A.-L. In Alkaloids: Chemical
& Biological Perspectives, Vol. 12; Pelletier, S. William, Ed.; Elsevi-
er: New York, 1998; pp 1-102.
(9) Tillequin, F. Recent Res. Devel. Phytochem. 1997, 1, 675-687.
(10) Wang, Z.-M.; Kakiuchi, K.; Sharpless, K. B. J . Org. Chem. 1994, 59,
6895-6897.
under the same conditions afforded 7 (117 mg, 95%): [R]20
D
(11) Lau, C. K.; Dufresne, C.; Belanger, P. C.; Pietre, S.; Scheigetz, J . J .
+67.2° (c 0.16, MeOH) HREIMS m/z 439.1633.
Org. Chem. 1986, 51, 3038-3043.
(12) Skaltsounis, A.-L.; Mitaku, S.; Gaudel, G.; Tillequin, F.; Koch, M.
Heterocycles 1992, 34, 121-128.
Cell Cu ltu r e a n d Cytotoxicity. L-1210 cells were culti-
vated in RPMI 1640 medium (Gibco) supplemented with 10%
fetal calf serum, 2 mM L-glutamine, 100 units/mL penicillin,
100 µg/mL streptomycin, and 10 mM HEPES buffer (pH 7.4).
Cytotoxicity was measured by the microculture tetrazolium
assay as described previously.18 Cells were exposed to graded
concentration of drug (nine serial dilutions in triplicate) for
48 h. Results are expressed as IC50 determined by the
concentration that reduced by 50% the optical density of
treated cells with respect to the optical density of untreated
controls.
(13) Mitaku, S.; Skaltsounis, A.-L.; Tillequin, F.; Koch, M. Synthesis 1992,
1068-1070.
(14) Mitaku, S.; Skaltsounis, A.-L.; Tillequin, F.; Koch, M.; Rolland, Y.;
Pierre´, A.; Atassi, G. Pharm. Res. 1996, 13, 939-943.
(15) Mitaku, S.; Skaltsounis, A.-L.; Tillequin, F.; Koch, M. J . Nat. Prod.
1986, 49, 1091-1095.
(16) Mitaku, S.; Skaltsounis, A.-L.; Tillequin, F.; Koch, M.; Pusset, J . Ann.
Pharm. Fr. 1989, 47, 149-156.
(17) Hlubucek, J .; Ritchie, E.; Taylor, N. C. Aust. J . Chem. 1970, 23, 1881-
1889.
(18) Pierre´, A.; Dunn, T. A.; Kraus-Berthier, L.; Leonce, S.; Saint-Dizier,
D.; Regnier, G.; Dhainaut, A.; Berlion, M.; Bizzari, J .-P.; Atassi, G.
Invest. New Drugs 1992, 10, 137-141.
Ack n ow led gm en t. We are grateful to Drs. B. Serkiz and
J . P. Volland (Institut de Recherches Servier) for their kind
help in the choice and optimization of the HPLC conditions.
NP980420Q