lactimidomycin and isomigrastatin

Lactimidomycin (LTM, 1) and iso-migrastatin (iso-MGS, 2) belong to the glutarimide-containing polyketide family of natural products. We previously cloned and characterized the mgs biosynthetic gene cluster from Streptomyces platensis NRRL 18993. The iso-MGS biosynthetic machinery featured an acyltransferase (AT)-less type I polyketide synthase (PKS) and three tailoring enzymes (MgsIJK). We now report cloning and characterization of the ltm biosynthetic gene cluster from Streptomyces amphibiosporus ATCC 53964, which consists of nine genes that encode an AT-less type I PKS (LtmBCDEFGHL) and one tailoring enzyme (LtmK). Inactivation of ltmE or ltmH afforded the mutant strain SB15001 or SB15002, respectively, that abolished the production of 1, as well as the three cometabolites 8,9-dihydro-LTM (14), 8,9-dihydro-8S-hydroxy-LTM (15), and 8,9-dihydro-9R-hydroxy-LTM (13). Inactivation of ltmK yielded the mutant strain SB15003 that abolished the production of 1, 13, and 15 but led to the accumulation of 14. Complementation of the ΔltmK mutation in SB15003 by expressing ltmK in trans restored the production of 1, as well as that of 13 and 15. These results support the model for 1 biosynthesis, featuring an AT-less type I PKS that synthesizes 14 as the nascent polyketide intermediate and a cytochrome P450 desaturase that converts 14 to 1, with 13 and 15 as minor cometabolites. Comparative analysis of the LTM and iso-MGS AT-less type I PKSs revealed several unusual features that deviate from those of the collinear type I PKS model. Exploitation of the tailoring enzymes for 1 and 2 biosynthesis afforded two analogues, 8,9-dihydro-8R-hydroxy-LTM (16) and 8,9-dihydro-8R-methoxy-LTM (17), that provided new insights into the structure-activity relationship of 1 and 2. While 12-membered macrolides, featuring a combination of a hydroxyl group at C-17 and a double bond at C-8 and C-9 as found in 1, exhibit the most potent activity, analogues with a single hydroxyl or methoxy group at C-8 or C-9 retain most of the activity whereas analogues with double substitutions at C-8 and C-9 lose significant activity.

Topics: Antibiotics, Antineoplastic; Bacterial Proteins; Base Sequence; Bioreactors; Cell Line, Tumor; Cell Survival; Drug Design; Gene Silencing; Humans; Macrolides; Models, Biological; Molecular Sequence Data; Molecular Structure; Multigene Family; Mutant Proteins; Neoplasms; Piperidones; Polyketide Synthases; Polyketides; Protein Engineering; Recombinant Proteins; Stereoisomerism; Streptomyces; Structure-Activity Relationship

Lactimidomycin (1) was described in the literature as an exquisitely potent cell migration inhibitor. Encouraged by this claim, we developed a concise and scalable synthesis of this bipartite glutarimide-macrolide antibiotic, which relies on the power of ring-closing alkyne metathesis (RCAM) for the formation of the unusually strained 12-membered head group. Subsequent deliberate digression from the successful path to 1 also brought the sister compound isomigrastatin (2) as well as a series of non-natural analogues of these macrolides into reach. A careful biological re-evaluation of this compound collection showed 1 and progeny to be potently cytotoxic against a panel of cancer cell lines, even after one day of compound exposure; therefore any potentially specific effects on tumor cell migration were indistinguishable from the acute effect of cell death. No significant cell migration inhibition was observed at sub-toxic doses. Although these findings cannot be reconciled with some reports in the literature, they are in accord with the notion that lactimidomycin is primarily a ribosome-binder able to effectively halt protein biosynthesis at the translation stage.

Topics: Anti-Bacterial Agents; Cell Movement; Female; Humans; Macrolides; Magnetic Resonance Spectroscopy; Piperidones

Migrastatin (1), iso-migrastatin (5) and lactimidomycin (7) are all glutarimide-containing polyketides known for their unique structures and cytotoxic activities against human cancer cell lines. Migrastatin, a strong inhibitor of tumor cell migration, has been an important lead in the development of antimetastatic agents. Yet studies of the related 12-membered macrolides iso-migrastatin, lactimidomycin, and related analogues have been hampered by their limited availability. We report here the production, isolation, structural characterization, and biological activities of iso-migrastatin, lactimidomycin, and 23 related congeners. Our studies showed that, as a family, the glutarimide-containing 12-membered macrolides are extremely potent cell migration inhibitors with some members displaying activity on par or superior to that of migrastatin as exemplified by compounds 5, 7, and 9-12. On the basis of these findings, the structures and activity of this family of compounds as cell migration inhibitors are discussed.

Topics: Animals; Biological Products; Cell Line, Tumor; Cell Movement; Cell Survival; Humans; Macrolides; Mice; Molecular Structure; Piperidones; Structure-Activity Relationship

Iso-Migrastatin (10) has been shown to be the main natural product of Streptomyces platensis, which undergoes a facile, H2O-mediated rearrangement into dorrigocin A (2), 13-epi-dorrigocin A (11), dorrigocin B (3), and migrastatin (1). Eight new congeners (12-19) of 10 were characterized. They can undergo the same H2O-mediated rearrangement into the corresponding 1, 2, 3, and 11 analogues (20-43) or 1,4-Michael addition with cysteine to afford the corresponding analogues (44-51) of NK30424 A and B (5, 6). This study generated a 47-member library of glutarimide polyketides, setting the stage to investigate the SAR for this family of natural products. These results also established the absolute stereochemistry of 5 and 6 and shed new light into the post-polyketide synthase steps for 10 biosynthesis.

Topics: Antibiotics, Antineoplastic; Lactones; Macrolides; Magnetic Resonance Spectroscopy; Molecular Structure; Piperidones; Streptomyces