Cellular DNA is subjected to ongoing damage in the course of normal metabolism and division. This damage is ameliorated by a complex network of pathways, known as the DNA damage response (DDR). Many cancer cell lines possess mutations in DNA DDR genes, which contributes to genomic instability and tumor progression. These mutations also in some cases sensitize these cell lines to DNA damaging agents, which provides an opportunity to eradicate cancer cells in the presence of non-transformed tissues. This proposal focuses on the synthesis, study, and preclinical evaluation of new anticancer agents against DNA repair deficient tumors that are based on the natural product lomaiviticin A. Lomaiviticin A is a complex dimeric metabolite that contains two diazofluorene functional groups and which is produced in vanishingly small quantities by a strain of Salinispora pacifica. Lomaiviticin possesses half-maximal inhibitory potencies in the low nanomolar-picomolar range against a panel of cultured human cancer cell lines. It was shown that the cytotoxic effects of lomaiviticin derive from the production of highly toxic double-strand breaks (DSBs) in DNA via the intermediacy of reactive carbon-centered free radical species. In accord with this, it was also established that cell lines possessing mutations in various DNA DSB repair factors, such as breast cancer type 2 (BRCA2), phosphatase and tensin homolog (PTEN), and ataxia-telangiectasia mutated (ATM), are highly sensitized toward lomaiviticin (LC50 values as low as <800 fM), suggesting application of lomaiviticin to treat these tumor types. However, large quantities of lomaiviticin are not available from the natural source, and despite this promising activity, a synthetic route that allows for manipulation, simplification, and optimization of its structure does not exist. The first objective of this application is to address this through completion of the synthesis of lomaiviticin A. To date, a short and flexible synthetic sequence to the monomeric units of lomaiviticin has been developed, and the successful coupling of these intermediates to provide the full carbon skeleton of the target has been achieved. The synthesis will be completed by implementing innovative strategies to control the stereoselectivity in the union of these intermediates. This proposal also seeks to obtain an understanding of the mechanism of generation of carbon-centered free radicals from lomaiviticin by studying the reactivity of isotopically-labeled derivatives toward thiol-based nucleophiles, nucleotides, and DNA duplexes in vitro, by NMR spectroscopy. The proposal also seeks to elucidate the mode of interaction of lomaiviticin with DNA by a host-guest crystallization method, which is an established and efficient approach to obtaining X-ray data of small molecule-DNA complexes. Using this structural data, synthetic analogs that are amenable to scale-up to support preclinical evaluation will be prepared and evaluated against a panel of DNA DSB repair-deficient cell lines. Ultimately, these studies will set the stage for in vivo evaluation of new anticancer agents that operate by a well- understood mechanism of action against tumors that are deficient in DNA DSB repair.
A large number of clinically-prescribed anticancer agents find their origins in natural products, molecules that are produced by plants, bacteria, and animals. This proposal focuses on the creation and study of novel anti- cancer agents based on the natural product lomaiviticin A. These compounds will be evaluated for the treatment of certain tumor types that possess deficiencies in DNA repair.
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