Absolute configuration of (+)-cyclophosphamide. A crystal and molecular structure determination by x-ray diffraction

Abstract

The crystal and molecular structure of enantiomerically homogeneous cyclophosphamide (C7H I 5N202PC12) has been determined by x-ray diffraction with the absolute configuration being established by the anomalous dispersion of the CI and P atoms. It is found that the dextrorotatory enantiomer of cyclophosphamide ([a]&'2.3O (c 3.0, methanol)) has the R configuration a t phosphorus. The compound crystallized in the rhombohedral space group R3 with the three molecules in the cell related by the threefold axis forming a trimeric unit by NH-O=P hydrogen bonding. Cell parameters are a = 10.520 ( 5 ) 8, and a = 108.9 (1)'. The conformation of the enantiomerically homogeneous cyclophosphamide a s compared to the racemate differs mainly in the orientation of one of the chloroethyl chains. Cyclophosphamide(2-[bis(2-chloroethyl)amino]-2H-1,3,2-0xazaphosphorinane 2-oxide, 1) is a widely used anticancer,drug which is prepared synthetically and administered clinically in racemic form (Cytoxan).2 The broad spectrum of activity3 exhibited by 1 has led to considerable interest in its metabolism, and a substantial amount of chemical and biochemical data supports the degradative pathway shown in Scheme I. Fragmentation of enzymatically produced 4-hydroxycyclophosphamide ( 2 ) and/or its putative aldehyde tautomer, aldophosphamide (3), affords acrolein and phosphoramide mustard (4), generally regarded as the ultimate DNA cross-linking agent. Competing enzymatic conversion of 2 and/or 3 into 4-ketocyclophosphamide ( 5 ) and carboxyphosphamide (6) is associated with drug deto~ification.~ Since biological systems normally exhibit a marked enantiomeric selectivity, it was expected5 and recently found6 that the antipodal forms of 1 exhibit significantly different therapeutic indices7 with (-)-I being more effective against PC6 mouse Karle et al. / (+)-Cyclophosphamide Absolute Configuration

X-ray structure and absolute configuration of (−)-(2S,4R)-4-methycyclophosphamide {2-[bis(2-chloroethyl)amino]-4-methyltetrahydro-2H-1,3,2-oxazaphosphorine 2-oxide}

“…Similar values have been reported for other cyclophosphamides, apart from (+)-CPA (Karle et al, 1977) and 4-hydroperoxy-CPA (Camerman, Smith & Camerman, 1977) in which angles of 355 and 357 ° were found. The 2-chloroethyl chains are not significantly extended as the Cl(1)...Cl(2) distance is only 4.994 (5),/k, similar to those found in other 4-substituted cyclophosphamides, e.g.…”

Structure of trofosfamide

“…This may indicate, particularly for the C(16)-C1 (17) alkylating agent. Similar long C-CI distances were observed in the cyclophosphamide structure (Karle et al, 1977;Adamiak et al, 1977). The packing diagram for TP is shown in Fig.…”

“…The six-membered ring is in the chair conformation and the configuration at the P is phosphoryl O axial and the bis(chloroethylamine) group equatorial. Comparison of the conformation of the chloroethyl alkylating groups of the present molecule with those found in 4-ketocyclophosphamide (Camerman & Camerman, 1973), cyclophosphamide (Garcia-Blanco & Perales, 1972;Karle, Karle, Egan, Zon & Brandt, 1977;Adamiak, Saenger, Kinas & Stec, 1977) and 4-hydroperoxycyclophosphamide (Camerman, Smith & Camerman, 1977) indicates that the exocyclic mustard groups, in particular, are very flexible and do not assume one favored conformation. It is likely that the important steric requirement for the chloroethyl chains is the ability for their reactive ends to be far enough apart to be able to alkylate separate bases on the DNA helices.…”

13C and 31P high resolution solid state NMR studies of cyclophosphamide and its analogues

“…These results obtained from SS NMR are complementary with those from liquid phase [35][36][37][38] and X-ray crystallography [39][40][41][42][43][44].…”