Synthesis of actinomycin and analogs. 12. Synthesis and some properties and antitumor effects of the actinomycin lactam analog, [di-(1'-L-.alpha.,.beta.-diaminopropionic acid)]actinomycin D

Abstract

A lactam analog of actinomycin D (AMD) was synthesized as a potential antitumor chemotherapeutic agent. Both L-threonine residues were replaced by L-.alpha.,.beta.-diaminopropionic acid. Starting with N.alpha.-benzyloxycarbonyl-N.beta.-tert-butyloxycarbonyl-L-.alpha.,.beta.-diaminopropionic acid methyl ester hydrochloride, the linear intermediate N.alpha.-benzyloxycarbonyl-N.beta.-(tert-butyloxycarbonylsarcosyl-L-N-methylvalyl)-L-.alpha.,.beta.-diaminopropionyl-D-valyl-L-proline p-nitrophenyl ester was prepared by conventional methods of peptide synthesis in solution. Selective cleavage of the N.beta.-tert-butyloxycarbonyl group and lactam formation afforded the desired cyclic pentapeptide derivative. The chromophore precursor, N.alpha.-(2-nitro-3-benzyloxy-4-methylbenzoyl) substituent, was introduced via its symmetric anhydride. Catalytic reduction followed by ferricyanide-mediated phenoxazinone formation provided the lactam analog, [di(1''-L-.alpha.,.beta.-diaminopropionic acid)]actinomycin D ([Dpr1]2-AMD). Its binding to natural and synthetic DNA and that of an analogous L-threo-.alpha.,.beta.-diaminobutyric acid containing lactam ([Dbu1]2-AMD) compared with the binding of AMD (in which the peptides are in lactone form) was studied by circular dichroic (CD) spectroscopy. The visible and UV CD spectra of free AMD differed from those of the free lactam analogs, indicating that the asymmetric environment of the pentapeptide rings in the region of the chromophore differs in free actinomycin lactone and lactams. In the presence of calf thymus DNA, PM2 DNA and the synthetic d(A-T)-like copolymers containing 2,6-diaminopurine (DAP), poly[d(DAP-T)], and poly[d(DAP-A-T)], the rotational strengths of the optically active transitions in the visible region of the actinomycins increased, and the CD spectra in the presence of the various DNA duplexes were qualitatively similar. The CD spectra of bound actinomycin lactams resembled the spectrum of bound AMD. The lactone and lactam actinomycins apparently acquired a similar environment when bound to DNA. [Dpr1]2-AMD was less cytotoxic than AMD in antibacterial [Bacillus subtilis, Staphylococcus aureus and Sarcina subflava] assays but exhibited somewhat higher toxicity in mice than AMD. At optimal dose levels the lactam analog had little or no antitumor activity in 3 murine tumor systems [L1210 leukemia, B16 melanoma and P388 leukemia].