A. G. Griesbeck, J. Hirt, K. Peters, E.-M. Peters, H.-G. von Schnering 619

Synthesis of cu-Amino-3-chloro-4,5-dihydro-5-methyl-5-isoxazo~eacetic Acid, a Ring-Methylated Analogue of the Antitumor Agent Acivicin (AT-125) Axel G. Griesbeck*a[*I, Joachim Hirta[*l, Karl Peters', Eva-Maria Peters', and Hans-Georg von Schnering'

Institut fur Qrganische Chemie, Universitat Wurzburg", Am Hubland, D-97074 Wurzburg, Germany

Max-Planck-Institut fur Festkorperforschungb, HeisenbergstraBe 1, D-70569 Stuttgart

Received September 5, 1994

Key Words: Acivicin / Photochemistry / 1,3-Dipolar cycloadditions / Jitrile oxi, .es

a-Amino-3-chloro-4,5-dihydro-5-methyl-5-isoxazoleacetic involving the photoisomerization of N-phthaloylvaline acid (8), a ring-methylated analogue of the potent antitumor methyl ester (1). The stereochemical course of the 1,3-dipolar agent acivicin (AT-125), is synthesized in a 6-step procedure cycloaddition is proven by means of a X-ray structure analy- in 63% overall yield from (S)-valine. Key step is the 1,3-dipo- sis of the major diastereoisomer ?a formed in the chloronitrile lar addition of bromonitrile oxide to the N,C-protected (S)- oxide cycloaddition. The absolute configuration of the major isodehydrovaline (6) available from (S)-valine in four steps ( u ) diastereomer ?a and the bromo derivative 7b is ( a S , 5 R ) .

Acivicin or AT-125, (crS,.5S)-a-amino-3-chloro-4,5-di- hydro-5-isoxazoleacetic acid (Figure l), has been isolated by Martin et al.['l from fermentation broths of streptomyceus sviceus and characterized as an antimicrobial, antimetabolic and effective antitumor agent[*].

Figure 1 . Structure of Acivicin (AT-125)

c1

Therefore, it has become the most interesting synthetic targets from the family of naturally occurring isoxazolines[31 in the last 15 years. The first total synthesis was carried out by Kelly and coworkers in 1979 starting from racemic trans- 3-hydroxy-4-aminocyclopentene[41. Several EPC syntheses have been reported using the chiral pool approa~h[~-~I . In order to gain more information about structure-activity re- lationships, analogues modified at C-3 (3-methy1I71, 3- bromo[']) have been synthesized, of which the 3-bromo compound shows similar activity compared to the parent system. In one case the 3-bromo analogue has also been reported to be a suitable substrate for transformation into aci~icin[~l. One of the most promising synthetic concepts is the 1,3-dipolar cycloaddition reaction of halonitrile oxide with C,N-protected p, y-unsaturated a-amino acids, an ap- proach which was first developed by Baldwin and cowork- ers['O]. According to this protocol the 3-chloro-5-methyl analogue has been synthesized as a racemate whereas at-

[*] New address: Institut fur Organische Chemie der Universitat zu Koln, GreinstraDe 4, D-50939 Koln.

tempts to synthesize the parent compound have failed due to the low reactivity of chloronitrile oxide[I']. Use of the more reactive bromonitrile oxide improves the efficiency of the 1,3-dipolar cycloadditiod'l and the addition of silver salts, as shown by Wade and coworkers, also activates chlo- ronitrile oxide for cycloaddition to monosubstituted ole- fins[l21. Thus, the addition of chloronitrile oxide to N- phthaloylvinylglycine in the presence of silver nitrate leads to a 37: 15 mixture (u:1)[l31 of diastereoisomeric cycload- ducts[12b]. Even lower is the d.r. in the case of the bromoni- trile oxide cycloaddition to N-protected vinylglycinol (3 :2 I:u)[~]. An excellent facial selectivity (>19:1) has been re- ported for the 1,3-dipolar cycloaddition of an enantiomer- ically pure N-glycosylnitrone to a (S)-vinylglycine deriva- tive[l41. During our studies of the photochemistry of N- phthaloyl-a-amino acid derivatives['5] we have developed a useful approach to enantiomerically pure isodehydrovali- ne[15a,bl, a suitable precursor for the synthesis of 5-methyl- acivicin. It has also been anticipated that the presence of an additional stereogenic center incorporated in the sterically demanding N-protecting group could improve the diaster- eoselectivity of the 1,3-dipolar cycloaddition step. Further- more, deprotection should be facilitated due to the easily removable imido functionality.

We have first investigated the addition of nitrile oxides to the methyl 1 -hydroxyisoindolinonecarboxylate 2 which is the product of the photoisomerization of N-phthaloylvaline methyl ester l[lSa]. As precursors for the in situ generation of nitrile oxides we have used the dihaloformaldoximes 4a, b synthesized from glyoxylic acid aldoxime 3 as described by Torsell et al.[l61. This procedure which involves a bishal- ogenation of 3 with NBS and NCS, respectively, leads to standard solutions of 4a and 4b which have immediately

Liebigs Ann. 1995,619-623 0 VCH Verlagsgesellschaft mbH, D-69451 Weinheim, 1995 0947-3440/95/0404-0619 $10.00+.25/0

620 A. G. Griesbeck, J. Hirt, K. Peters, E.-M. Peters, H.-G. von Schnering

been used for the reaction with the dipolarophile. The ni- trile oxides are liberated by reaction of 4a, b with an excess of potassium hydrogen carbonate in DME at room tem- perature. Complete decomposition of the substrate 2 occurs if chloronitrile oxide is generated by using the procedure described above. We have already noticed earlier" 5b1 that 2 is easily isomerized in the presence of weak bases and epimerized at the stereogenic center C-1 in the presence of mineral acids. Probably one of these reactions induces de- composition of the substrate due to secondary reactions which may compete with the (slow) cycloaddition step. Bromonitrile oxide adds much more rapidly to disubsti- tuted CC double bonds and, as expected, the reaction of 2 with this 1,3-dipole results in the formation of the cycload- ducts 5 in a diastereomeric ratio of 65:35 in 52% yield (Scheme 1).

Scheme 1

1 2

(i) Phthalic anhydride, 145°C (98%) (ii) MeOH, HCI, 0°C (97%)

KHCO3

5

3 4b

The fact that only two out of four possible diastereomers are formed in this cycloaddition reaction indicates that the relative (and absolute) configuration at the a-carbon atom and the C-2 carbon atom has been preserved, i.e. no epi- merization has occurred. Another indication of this as- sumption is that the only pronounced difference in 13C NMR chemical shifts is observed for the methyl signals (6 = 22.8 and 24.8). The asymmetric induction by the two existing stereogenic centers, however, is only marginal. Therefore, we have next compared the cycloaddition stereo- selectivity of nitrile oxides to the N-phthaloylisodehydrova- line methyl ester 6, which is available by oxidation of 2 with PCC[lSb]. Due to the much higher stability of 6, bromo- as well as chloronitrile oxide have been successfully applied. The cycloaddition of 6 to bromonitrile oxide generated from 4b is complete after a reaction time of 66 hours at room temperature and the isoxazolines 7b are formed in a diastereomeric ratio of 83: 17 in 85% yield. The less reactive chloronitrile oxide generated from 4a yields after a reaction time of 72 hours the isoxazolines 7a with 60% conversion (corrected yield: 73%) and in a diastereomeric ratio of 84: 16 (Scheme 2).

In order to establish the relative configuration we have compared the 13C NMR shift data of the diastereomeric

Scheme 2

,-&, 0

0

gx COOCH,

I 7s,b (a=CI, b=Br) 6

(iv) (74% 7a-18) H O N N Y X X 1 (94% 7b+8)

(iii) PCC, Alox, CHzClz (98%)

(iv) NH~NH~xHOAC, then HC1 COO-

8

mixtures of isoxazolines 7a and 7b with data published by Borowicz and Eckstein for analogous compounds { a-amino-5-methyl-3-(2',6'-dichlorophenyl)-4,5-dihydro-5- isoxazoleacetic acid 2,2,2-trichloroethyl ester} [l71. For these systems the authors have assigned the high-field-shifted 13C NMR 5-methyl group signal and the low-field-shifted 13C NMR C-4 group signal to the I diastereoisomer and the corresponding lowfield-shifted 5-methyl signal (A6 = 2.5) and the highfield-shifted C-4 signal (A6 = 1.4) to the u dias- tereois~mer['~~]. Consequently we assign the (aS5S) con- figuration to the major diastereomer 7a formed in the chlo- ronitrile oxide cycloaddition reaction with 6 (6[CH3] = 23.8, 6[CH2] =48.9). The shift difference for the two critical methyl resonances is A6 = 2.7 and the two methylene reso- nances is A6 = 1.6, also in agreement with the data given in literature[l71. A crystal structure performed by us with the major diastereomer 7a (crystallized from ace- tonekhloroform), however, has revealed that this assign- ment is incorrect (Figure 2).

Figure 2. Structure of 7a (aS,SR) in the crystal. Selected bond lengths [pm] and angles PI. Cl-Cll 158.4(7), C1-N2 145.9(5), Cl-Cl6 151.7(8), Cll-C19 152.3(10), Cll-C15 153.1(7), Cll-012 146.2(7), 012-N13 140.0(8), N13-Cl4 127.2(6), C14-CI 170.9(7), C14-Cl5 147.0(10), N2-C3 141.1(7), C3-C4 149.1(7), C3-03 120.6(9); 03-C3-N2 125.1(4), C3-N2-C1 122.5(5), N2-Cl-CI6 113.7(5), N2-Cl-CIl 111.8(4), Cl-Cll-012 104(4), Cl-Cll-Cl9 114.7(5), Cl-C11-C15 109.8(4), C19-C11-012 107.7(4), C19-Cll-Cl5 115.5(4), Cll-CI5-Cl4 99.6(4), C15-Cl4-Cl 123.6(4), Cl-Cl4-NI3 119.6(6), C15-Cl4-Nl3 116.8(6), C14-N13-012 107.3(6),

N13-012-Cll 109.7(4)

CI

010

Liebigs Ann. 1995, 619-623

Analogue of the Antitumor Agent Acivicin 62 1

Figure 3. I3C-NMR 5-methyl and C-4-methylene group signal of 7a (USSR) and 7a (crS,SS)

7s (aS,SR) = u 7s (aS,SS) = 1

Due to the fact that the chemical shift values (Figure 3) as well as the shift differences for a whole set of nitrile oxide cycloadducts[’91 with the 1,3-dipolarophiles 2 and 6 are nearly equal (AS = k0.l) and comparable with the data given in ref.[17] we assume that also the literature assign- ment must be corrected in the way described here. The cor- rect assignment for the chloronitrile oxide adduct (and analogous for the bromo derivative - vide infra) is (aSJR).

Deprotection of a mixture of the cycloadduct 7a has eas- ily been achieved in 94% yield by using a two-step pro- cedure: hydrazinolysis[20] and subsequent saponification on an acidic ion exchange resin. A major disadvantage of this sequence which we up to now have not been able to circum- vent is partial epimerization at the stereogenic center C-5 presumably occuring during hydrazinolysis. In agreement with the experiments described by Vyas et al.i91 the bromo derivative 7b can also be transformed into 5-methylacivicin 8 by treatment with aqueous HCl after hydrazinolysis. The product obtained after this procedure is identical with the mixture obtained after deprotection of the cycloadducts 7a from the chloronitrile oxide addition to 6, thus proving the correct stereochemical assignment for cycloadducts 7b.

Detailed and investigations of the stereoselectivity of nitrile oxide cycloaddition reac- t i o n ~ [ ~ ~ ] with monosubstituted chiral olefins have been pub- lished in the last decade. Houk, Jager and coworkers have developed a profound concept for the prediction of the di- rection and relative amount of asymmetric induction in these The “anti-periplanar effect” in the stag- gered (allylic substituents with respect to the forming bonds) transition state has been shown to operate for a large number of x-system~[~~] and influences both reactivity and stereoselectivity of electrophilic additions. A combi- nation of steric and electrostatic effects may be responsible for our results on the diastereoselectivity of the nitrile oxide additions to 2 and 6. Assuming that in the preferred tran- sition-state structures the largest substituent at the allylic stereogenic center is located anti-periplanar with respect to the incoming nitrile oxide oxygen and the allylic hydrogen is located “outside”, the methoxycarbonyl or the phthalimido substituent is located “inside” (Figure 4).

The transition-state I should be the preferred state, which gives the major u diastereomer which is also the main prod- uct in the cycloaddition reaction of benzenesulfonylcarbon- itrile oxide to (9-vinylglycine-derived alkenes[12bl. The dia- stereoselectivity when going from the phthalimido-substi- tuted dipolarophile 6 to the hydroxyisoindolinone-substi- tuted substrate 2 decrease as a effect of the steric demand in the latter case. The competing reaction path (structure

Figure 4. Proposed transition-state structures for the formation of the u diastereomer (I) and the I diastereomer (n); E = COOMe

I + u (7a) II + I (7a)

II with the nitrogen substituent anti-periplanar) becomes more favored.

In summary, 5-methyl-substituted acivicin can be synthe- sized in a 6-step procedure which involves photoisomeri- zation of C,N-protected (8-valine and bromonitrile oxide cycloaddition in an overall yield of 63%.

This work was supported by the Deutsche Forschungsgemein- schaft (project Gr 881/7-l), the Fonds der Chemischen Industrie, and the Degussa AG, Hanau.

Experimental IR: Perkin-Elmer 1420 spectrometer. - ’H NMR spectra: Hita-

chi Perkin-Elmer R-24 B (60 MHz), Bruker AW 80 (80 MHz), Bruker AC 200 (200 MHz), Bruker AC 250 (250 MHz), and Bruker WM 400 (400 MHz). - I3C NMR spectra: Bruker AC 200 (50.3 MHz), Bruker AC 250 (63.4 MHz), and Bruker WM 400 (100.6 MHz), TMS as internal standard. - UVNIS: Hitachi U-3200. - Column chromatography: silica gel (Merck) 60-230 mesh; pe- troleum ether (PE, 40-60°C), ethyl acetate (EA). - Optical ro- tations: Perkin-Elmer 241 MC polarimeter. - Combustion analy- ses: Institut fur Anorganische Chemie der Universitat Wurzburg.

(S)-N-Phthaloylvaline: 14.8 g (10.0 mmol) of phthalic anhydride was melted in a stoppered flask preheated to 140-145°C. During vigorous stirring 11.7 g (10.0 mmol) of (8-valine was added within 5 min. This mixture was kept at 140°C for about 10 min. During the last 5 min the flask was opened in order to evaporate the reac- tion water. After cooling the crystalline residue was recrystallized from MeOH to give 24.1 g (98%) of (3-N-phthaloylvaline as color- less needles, m.p. 116- 117°C (ref.[*5a,b] 116- 117°C). - IR (CCI,): 6=2980 cm-’ (m), 2940 (m), 1785 (s), 1725 (m), 1475 (m), 1395 (vs), 1290 (m), 1080 (m), 920 (m), 720 (m). - ‘H NMR (250 MHz,

CH3), 2.76 [dsept, J=6.7 Hz, 8.2 Hz, lH, CH(CH3),], 4.64 (d, J=8.2 Hz, lH, NCH), 7.75 (m, 2H, ar. H), 7.89 (m, 2H, ar. H), 10.54 (br. s, lH, COOH). - I3C NMR (63 MHz, CDC13): S = 19.8

(d, 2 C, ar. C), 131.8 (s, 2 C, ar. C), 134.5 (d, 2 C, ar. C), 168.0 (s, 2 C, NC=O), 174.8 (s, C=O).

(S)-N-Phthaloylvaline Methyl Ester (1): A slow stream of gase- ous HCI was bubbled through a precooled (0°C) solution of 24.8 g (10.0 mmol) of (3-N-phthaloylvaline in 150 ml of methanol for 2 min. After cooling to 0°C the solution was again saturated with gaseous HCl for 2 min. The solution was stirred for about 12 hours for complete conversion. The methanol was removed at reduced pressure to yield 25.2 g (97%) of 1 as a colorless oil: b.p. 140- 142”C/0.01 mbar (ref.[14a,b] 140- 142”C/0.01 mbar). - [aID= -66.7 (c= 1.2 in methanol). - IR (CCb): 0=2990 cm-’ (m), 2880 (w), 1770 (vs), 1740 (vs), 1390 (m), 1330 (vs), 1285 (s), 1220 (s), 1080 (m), 1025 (m), 915 (m). - UV (CH3CN): I,,, (lg E ) = 293.7 nm (3.09), 242.0 (3.78), 233.3 (3.91), 219.5 (4.36). - ‘H

CDC13): 6 = 0.92 (d, Jz6.7 Hz, 3H, CH3), 1.17 (d, J=6.7 Hz, 3H,

(q, CH3), 21.2 (q, CH3), 28.7 [d, CH(CH3)2], 57.8 (d, NCH), 123.9

Liebigs Ann. 1995, 619-623

622 A. G. Griesbeck, J. Hirt, K. Peters, E.-M. Peters, H.-G. von Schnering

NMR (250 MHz, CDC13): 6 = 0.89 (d, J = 6.8 Hz, 3H, CH,), 1.13 (d, J=6.8 Hz, 3H, CH,), 2.74 [dsept, J=6.8 Hz, 8.3 Hz, IH, CH(CH&], 3.69 (s, 3H, OCH3), 4.45 (d, J=8.3 Hz, IH, NCH), 7.73 (m, 2H, ar. H), 7.86 (m, 2H, ar. H). - I3C NMR (63 MHz,

(q, OCH3), 57.7 (d, NCH), 123.8 (d, 2 C, ar. C), 131.9 (s, 2 C, ar. C), 134.5 (d, 2 C, ar. C), 167.9 (s, 2 C, NC=O), 169.5 (s, C=O).

Methyl ( I ' R,2S)-2- (2,3-Dihydro-1 -hydroxy-3-oxo-l H-isoindol-2- yl)-3-methyl-3-butenoate (2) was prepared according to

Glyoxylic Acid Aldoxime (3) was prepared in nearly quantitative yield according to ref.['6] and was recrystallized from diethyl ether, m.p. 133-136°C (ref.[16] 134- 136°C).

Dichloroformaldoxime (4a): 6.77 g (50.6 mmol) of N-chlorosucci- nimide and 2.31 g (26.2 mmol) of glyoxylic acid aldoxime (3) in 40 ml of 1,2-dimethoxyethane (DME) were heated at reflux causing evolution of carbon dioxide. After C02 evolution had ceased, chlorination was completed. The reaction mixture was brought to room temp., and the solution was then used directly for further re- action.

Dibromoformaldoxime (4b): To a cooled solution (0-5°C) of 1.41 g (1 5.8 mmol) of glyoxylic acid aldoxime (3) in 14 ml of DME and 4.3 ml of water 5.13 g (28.9 mmol) of N-bromosuccinimide was added in portions. After the addition of NBS was complete the cooling bath was removed and the solution was warmed up to room temp. The reaction mixture was stirred until it became color- less (ca. 30 min) and then used directly for further reaction.

(S)-N-Phthaloylisodehydrovaline Methyl Ester (6) was prepared according to ref.[lsh1

Methyl a-(2,3-Dihydro-I-hydroxy-3-oxo-lH-isoindol-2-yl)-3- bromo-4,5-dihydro-5-methyl-5-isoxazoleacetate (5): A solution of dibromoformaldoxime (4b, 7.37 mmol) in 15 ml of DME (see above) was added to a mixture of 0.385 g (1.47 mmol) of 2 and 2.85 g (26.0 mmol) of KHCO, in 5 ml of DME. After 17 h the precipitate formed was filtered and the solvent was removed from the filtrate at reduced pressure. The residue was dissolved in 40 ml of ethyl acetate and the solution was washed with brine. After dry- ing with magnesium sulfate and evaporation of the solvent in vacuo the crude product was purified by column chromatography (PEI EA, 2: 1) to give 292 mg (52%) of 5 as a colorless oil. The 'H-NMR analysis of the crude reaction mixture revealed a diastereomeric excess of 30%. Rf=0.25. - IR (CCI4): 3=3600 cm-' (m), 3440 (br. m), 2980 (m), 1765 (vs), 1725 (vs), 1480 (m), 1445 (s), 1425 (s), 1220 (s), 1170 (m), 900 (m). - 'H NMR (250 MHz, CDC13) major diastereomer: 6 = 1.51 (s, 3H, CH,), 3.12 (d, J=18.1 Hz, lH, CHJ, 3.47 (d, J=18.1 Hz, lH, CH2), 3.67 (s, 3H, OCH,), 4.22 (d, J=10.0 Hz, IH, OH), 4.65 (s, lH, NCH), 6.12 (d, J=9.6 Hz, CHOH), 7.49 (m, lH, ar. H), 7.58 (m, 2H, ar. H), 7.73 (m, IH, ar. H); minor diastereomer: 6 = 1.52 (s, 3H, CH3), 3.02 (d, J= 18.0 Hz, lH, CH?), 3.62 (d, J=18.0 Hz, IH, CHJ, 3.67 (s, 3H, OCH3),4.62 (d, J=8.6 Hz, IH, OH), 4.85 (s, lH, NCH), 5.97 (d, J=8.1 Hz, lH, CHOH), 7.49 (m, IH, ar. H), 7.58 (m, 2H, ar. H), 7.73 (m, IH, ar. H). - 13C NMR (63 MHz, CDCI3) major diastereomer: 6 = 22.8 (q, 5-methyl), 51.7 (t, C-4)> 52.7 (q, OCH,), 55.4 (d, NCH), 82.7 (d, NCOH), 89.8 (s, C-5), 123.5 (d, ar. C), 123.6 (d, ar. C), 129.9 (d, ar. C), 130.2 (d, ar. C), 132.9 (d, ar. C), 138.1 (s, ar. C), 144.2 (s, C-3), 167.6 (s, NC=O), 168.1 (s, C=O); minor diastereomer: 6 = 24.8 (q, 5-methyl), 51.6 (t, C-4), 52.6 (q, OCH3), 58.9 (d, NCH), 82.7 (d, NCOH), 87.9 (s, C-5), 123.4 (d, ar. C), 123.7 (d, ar. C), 129.8 (d, ar. C), 129.9 (d, ar. C), 133.0 (d, ar. C), 137.7 (s, ar. C), 144.1 (s, C-3), 168.1 (s, NC=O), 168.7 (s, C=O). - CI5Hl5BrN2O5 (383.2): calcd. C 46.97, H 3.91, N 7.31; found C 46.83, H 3.50, N 7.11.

CDC13): 6 = 19.6 (q, CH3), 21.1 (q, CH3), 28.8 [d, CH(CH&], 52.6

Methyl 3- Chloro-4,S-dihydro-5-methyl-a-phthalimido-5-isoxazole- acetate (7a): A solution of dichloroformaldoxime (4a, 26.2 mmol) in 40 ml of DME (see above) was added dropwise at 25°C to a stirred mixture of 1.36 g (5.24 mmol) of 6 and 11.6 g (0.11 mol) of KHC03 in 30 ml of DME and 0.7 ml of water. After 72 h the solution was filtered and the solvent was evaporated from the fil- trate at reduced pressure at 25°C. The residue was dissolved in 40 ml of ethyl acetate and the solution washed twice with brine. After drying with magnesium sulfate and evaporation of the solvent in vacuo the crude product was purified by column chromatography (PEIEA, 2: 1) to give 0.78 g (44%) of 7a as a colorless oil and 0.34 g (25%) of 6. The 'H-NMR analysis of the crude reaction mixture revealed a conversion rate of 60% and a diastereomeric excess of 68%. (aSJR): Crystallization from acetonekhloroform, m.p. 162-163°C. - [aID=-40.6 (c=0.5 in acetonitrile). - IR (KBr): 0=2960 cm-' (m), 1775 (s), 1745 (vs), 1725 (vs), 1440 (m), 1380 (s), 1300 (m), 1260 (s), 1230 (m), 1075 (m). - Rf=0.39. - 'H NMR (250 MHz, CDCl,): ( d , S R ) : 6 = 1.47 (s, 3H, CH3), 3.04 (d, J=18.0 Hz, IH, CHz), 3.70 (d, J=18.0 Hz, IH, CH2), 3.71 (s, 3H, OCH,), 5.11 (s, IH, NCH), 7.71 (m, 2H, ar. H), 7.83 (m, 2H, ar.

3.72 (s, 3H, OCH,), 3.91 (d, J=18.2 Hz, IH, CH2), 5.01 (s, IH, NCH), 7.71 (m, 2H, ar. H), 7.83 (m, 2H, ar. H). - I3C NMR (50 MHz, CDC13): (aS5R): 6 = 23.8 (q, 5-methyl), 48.9 (t, C-4), 52.9 (q, OCH3), 55.8 (d, NCH), 88.2 (s, C-5), 123.8 (d, 2 C, ar. C), 131.3 (s, 2 C, ar. C), 134.5 (d, 2 C, ar. C), 148.8 (s, C-3), 166.8 (s, 2 C, NC=O), 167.1 (s, C=O). - (aS5S): 6 = 26.5 (q, 5-methyI), 47.3 (t, C-4), 52.9 (q, OCH,), 55.8 (d, NCH), 88.5 (s, C-5), 123.9 (d, 2 C, ar. C), 131.2 (s, 2 C, ar. C), 134.4 (d, 2 C, ar. C), 150.0 (s, C-31,

calcd. C 53.50, H 3.89, N 8.32; found C 53.33, H 3.73, N 8.24.

Crystallographic Features: 7a (as 5R) : C 5H ,N205C1 (3 36.73); a=1124.7(2), c=1051.7(3) pm, V=1152.0(6) X lo6 pm3, Z = 3, d(ca1cd.) = 1.456 g ~ m - ~ , trigonal, space group P32. Data collection: Siemens R3ml V diffractometer, MoK,, graphite monochromator, crystal size: 0.9 X 0.9 X 0.3 [mm], Wyckoff scan, theta range ["I: 1.75-27.5, recip. lattice segment: h = 0-14, k=0-14, I=-13-13, no. refl. measured: 2025, no. unique refl.=1758, no. refl. F > 30(F)= 1971, linear absorption coeff. [mtr-']=0.28, absorption correction: Y scan. Structural analysis and refinement: solution by direct phase determination, method of refinement: Full-matrix least-squares, hydrogen positions of riding model with fixed iso- tropic U, data-to-parameter ratio: 9.48, R=0.047, R,=0.047, weighting scheme: w = I/02(F), lar4est difference peak: 0.25 eA-,, largest difference hole: 0.27 eA-,, program used: Siemens SHELXTL PLUS on Micro VAX 11.

a-Amino-3-chioro-4,S-dihydro-5-methyl-5-isoxazoleacetic Acid (8): A solution of 1.30 g (3.84 mmol) of 7a (a diastereomeric mix- ture of 87:13) and 0.38 g (4.15 mmol) of anhydrous hydrazinium acetate in 35 ml of methanol was refluxed for 45 min and sub- sequently cooled. After concentration of the solution at reduced pressure to 10 ml, 1.6 ml of a saturated methanolic hydrogen chlor- ide solution was added to the solution and the white precipitate was filtered off. The filtrate was evaporated at 25"C/18 Torr to dry- ness and the residue dried in vacuo for about 12 hours. Water (8 ml) and 1.1 ml of concentrated hydrogen chloride were added and the solution was refluxed for 15 min, then concentrated. The resi- due was dissolved in 10 ml of water, 5 g of charcoal was added and the mixture filtrated. The solvent was removed at reduced pressure. The 'H-NMR analysis of the crude methyl ester hydrochloride of 8 revealed a diastereomeric ratio of 66:34. The residue was dis- solved in 10 ml of water and the solution placed on a Dowex SOW X 4 column (60 ml; 50- 100 mesh, H+ form, after two cycles with

H);(aS,5s):6= 1.76(~,3H,CH3),3.10(d,J=18.2H~, lH,CHz),

166.8 (s, 2 C, NC=O), 167.1 (s, C=O). - CISH~~CIN~OS (336.7):

Liebigs Ann. 1995, 619-623

Analogue of the Antitumor Agent Acivicin 623

60 ml of 2 N HCl and 60 ml of 3 M NH3). The column was washed with 10 ml of water and eluated with 3 M NH3. Evaporation of the eluant gave 0.55 g (%YO) of 8 in a diastereomeric ratio of 66:34, which was recrystallized from waterfacetone. - 'H NMR (200 MHz, DzO): ( d , 5 R ) : 6 = 1.40 (s , 3H, CH3), 3.23 (d, J=18.2 Hz, lH, CHJ, 3.77 (d, J=18.2 Hz, lH, CH,), 3.97 (s, lH, NCH); (aS,SS): 6 = 1.57 (s, 3H, CH3), 3.14 (d, Jz18.3 Hz, IH, CHZ), 3.62 (d, Jz18.3 Hz, lH, CHJ, 3.87 (s, lH, NCH). - 13C NMR (63 MHz, D20): (aS,SR): 6 = 20.5 (q, Smethyl), 49.5 (t, C-4), 60.4

6 = 25.6 (q, 5-methyl), 46.3 (t, C-4), 60.9 (d, NCH), 88.6 (s, C-5), 152.6 (s, C-3), 171.1 (s, C=O). - C6H9C1N203 (192.6): calcd. C 37.42, H 4.71, N 14.55; found C 37.55, H 4.71, N 14.52.

Methyl 3-Bromo-4,5-dihydro-5-methyl-ol-phthalimido-5-isoxazoie- acetate (7b): A solution of dibromoformaldoxime (4b, 15.8 mmol) in 14 ml of DME (vide supra) was added to a mixture of 0.82 g (3.16 mmol) of 6 and 6.13 g (55.8 mmol) of KHC03 in 12 ml of DME. After 66 h the precipitate formed was filtered off and the solvent was removed at reduced pressure. The residue was dissolved in 40 ml of ethyl acetate and the solution was washed with brine. After drying with magnesium sulfate and evaporation of the solvent in vacuo the crude product was purified by column chromatogra- phy (PE/EA, 2:l) to give 1.03 g (85Y0) of 7b as a colorless oil. The 'H-NMR analysis of the crude reaction mixture revealed a diastereomeric excess of 66%. Rf=0.29. - 'H NMR (250 MHz,

CH,), 3.68 (s, 3H, OCH3), 3.71 (d, J=18.1 Hz, lH, CH,), 5.09 (s, lH, NCH), 7.73 (m, 2H, ar. H), 7.84 (m, 2H, ar. H); (aS,5s): 6 =

OCH3), 3.93 (d, J=18.3 Hz, IH, CH2), 5.00 (s, lH, NCH), 7.73 (m, 2H, ar. H), 7.84 (m, 2H, ar. H). - I3C NMR (63 MHz, CDC13): (a$SR): 6 = 23.7 (q, Smethyl), 51.9 (t, C-4), 52.8 (q, OCH3), 55.9 (d, NCH), 87.6 (s, C-5), 123.8 (d, 2 C, ar. C), 131.3 (s, 2 C, ar. C), 134.4 (d, 2 C, ar. C), 137.1 (s, C-3), 167.0 (s, 2 C, NC=O), 167.2 (s, C=O); (cwSSS): 6 = 26.4 (q, Smethyl), 50.2 (t,

ar. C), 131.3 (s, 2 C, ar. C), 134.4 (d, 2 C, ar. C), 137.3 (s, C-3), 167.0 (s, 2 C, NC=O), 167.2 (s, C=O). - CI5Hl3BrN2O5 (381.2): calcd. C 47.26, H 3.44, N 7.34; found C 47.43, H 3.40, N 7.51.

a-Amino-3-chloro-4,5-dihydro-5-methyl-S-isoxazoleacetic Acid (8) from 7b: A solution of 1.03 g (2.69 mmol) of 7b (a diastereomeric mixture of 84:16) and 0.27 g (2.91 mmol) of anhydrous hydrazin- ium acetate in 25 ml methanol was refluxed for 45 min and sub- sequently cooled. The reaction mixture was worked up as described for the corresponding chloro compound. The yield was 0.60 g (!%YO) of 8, which was recrystallized from watedacetone. After this procedure the diastereomeric ratio had decreased from 84: 16 (7b) to 66:34 (8).

(d, NCH), 88.0 (s, C-5), 153.1 (s, C-3), 171.1 (s, C=O); (aS,5S):

CDC13): ( 4 5 R ) : 6 = 1.44 (s, 3H, CH3), 3.07 (d, J=18.1 Hz, IH,

1.74 (s, 3H, CH3), 3.12 (d, Jz18.3 Hz, lH, CH3, 3.69 (s, 3H,

C-4), 52.8 (q, OCH3), 56.9 (d, NCH), 87.9 (s, C-5), 123.8 (d, 2 C,

['I D. G. Martin, D. J. Duchamp, C. G. Chidester, Tetrahedron Lett. 1973, 2549-2552.

['I L. J. Hanka, D. G. Martin, G. L. Neil, Cancer Chemother. Rep., Part I 1973, 57, 141-147. - LZb] D. P. Houchens, A. A. Ovejera, M. A. Sheridan, R. K. Johnson, A. E. Bogden, G. L. Neil, Cancer Treat. Rep. 1979, 63, 473-476.

[31 [3a] Tricholomic acid: I. Wagner, H. Musso, Angew. Chem. 1983, 95, 827-839; Angew. Chem. Znt. Ed. Engl. 1983, 22, 816. - [3b1 Dihydromuscimol: M. De Amici, C. De Micheli, V. Misani, Tetrahedron 1990,46, 1975- 1986. - L3'] ~-(3-Zsoxazolin-5-on-2- y1)alanin: S. U. Schenk, D. Werner, Phytochernistry 1991, 30, 467-4711 .-. . ._.

K4] R. C. Kelly, I. Schletter, S. J. Stein, W. Wierenga, 1 Am. Chem. SOC. 1979,101, 1054-1056. From dehvdrodutamate: J. E. Baldwin. L. I. Kruse. J.-K. Cha. J Am. Chem. ~ O C . 1981,103,942-943.

Chem. SOC. 1981, 103, 7357-7358. C6] From glutamic acid: R. B. Silverman, M. W. Holladay, .I Am.

[q Z. Gombos. J. Nvitrai, P. Kolonits, M. Kaitar-Peredv. 1 Chem. - I

SOC., Perkin Trans. 1, 1989, 1915-'1921.

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25,487-490.

Commun. 1976, 795-796.

['I A. A. Hagedorn 111, B. J. Miller, J. 0. Nagy, Tetrahedron Lett.

['I D. M. Vyas, Y. Chiang, T. W. Doyle, Tetrahedron Lett. 1984,

[lo] J. E. Baldwin, C. Hoskins, L. I. Kruse, 1 Chem. Soc. Chem.

[''I J. E. Baldwin, J. K. Cha, L. I. Kruse, Tetrahedron 1985, 41,

[I2] P. A. Wade, M. K. Pillay, S. M. Singh, Tetrahedron Lett. 1982, 23, 4563-4566. - P. A. Wade, S. M. Singh, M. K. Pillay, Tetrahedron 1984, 40, 601 -61 1.

[I3] For the use of this nomenclature, see: D. Seebach, V. Prelog, Angew. Chem. 1982, 94, 696-702; Angew. Chem. Znt. Ed. Engl. 1982,21, 654-660.

[141 R. A. Whitney, 1 Am. Chem. SOC. 1987, 109, 276-277. - S. Mzengeza, R. A. Whitney, 1 Org Chem. 1988,53,4074-4081.

[I5] [Isa] A. G. Griesbeck, H. Mauder, Angew. Chem. 1992, 104, 97-99. Angew. Chem. Znt. Ed. Engl. 1992, 31, 73-75. - [15b] A.' G. Griesbeck, H. Mauder, I. Miiller, Chem. Ber 1992, 125, 2467-2475. - [Isc] A. G. Griesbeck, H. Mauder, I. Miiller, K. Peters, E.-M. Peters, H. G. von Schnering, Tetrahedron Lett. 1993, 34, 453-456. - A. G. Griesbeck, A. Hem, J. Hirt, V. Ptatschek, T. Engel, D. LoEler, F. W. Schneider, Tetrahedron 1994,50, 701-714.

[16] K. Halling, I. Thomson, K. B. G. Torsell, Liebigs Ann. Chem. 1989, 985-990.

[I7] P. Borowicz, Z. Eckstein, Pol. 1 Chem. 1984, 58, 65-72. - P. Borowicz, Z. Eckstein, ibid., 887-894. - [174 P. Boro-

wicz, A. Zimniak, 2. Eckstein, Bull. Acad. Pol. Sci. Sex Sci. Chim. 1982, 30, 23-29.

["I Further details of the crystal structure investigation are avail- able on request from the Fachinformationszentrum Karlsruhe, Gesellschaft fur wissenschaftlich-technische Information mbH, D-76344 Eggenstein-Leopoldshafen, on quoting the depository number CSD-401166, the names of the authors, and the jour- nal citation.

[I9] We also studied the cycloaddition of ethoxycarbonylnitrile ox- ide to 2 and 6 which gave analogous results.

[20] W. L. F. Armarengo, B. A. Milloy, W. Pendergast, 1 Chem. Soc., Perkin Trans. I 1976, 2229-2237.

12'] E.g.: L21a] A. P. Kozikowski, A. K. Ghosh, 1 Org. Chem. 1984, 49, 2762-2772. - ['lb] V. Jager, R. Schohe, E. F. Paulus, Tetra- hedron Lett. 1983. 24. 5501-5504. - I2Ic] V. Jaeer. 1. Miiller.

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S. Mzengeza, C. M. Yan

R. Schohe, M. Frey, R . Ehrler, B. Hafele, D. Ghroter, Lect.' Heterocycl. Chem. 1985, 8, 79-98. [22a] K. N. Houk, S. R. Moses, Y-D. Wu, N. G. Rondan, V. Jager, R. Schohe F. R. Fronczek, J. Am. Chem. SOC. 1984,106, 3880-3882. - 132bl K. N. Houk. H.-Y. Duh. Y-D. Wu. S. R. Moses, 1 Am. Chem. SOC. 1986,108, 2754-2755.

Adv. Heterocycl. Chem. 1994, 60, 261 -327.

1 Am. Chem. Soc. 1981, 103, 2438-2440.

[231 C. J. Easton, C. M. M. Hughes, G. P. Savage, G. W. Simpson,

LZ4] F! Caramella, N. G. Rondan, M. N. Paddon-Row, K. N. Houk,

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