Approach to a enedyine biosynthesis tool box through metagenomic and targeted metabolomic analyses of actinomycetes
Final Report Abstract
The enediynes represent one of the most fascinating families of natural products for their unprecedented molecular architecture and extraordinary biological activities. Their structures contain a unit consisting of two acetylenic groups conjugated to a double bond or incipient double bond within a 9- or 10-membered carbocycle. The electronic rearrangement of the enediyne carbocycle produces a transient benzenoid diradical, which when positioned within the minor groove of DNA abstracts hydrogen atoms from DNA causing interstrand crosslinks (ICLs), DNA double-strand breaks (DSBs), or both. With this exquisite mode of action and the extraordinary cytotoxicity, the enediynes have been successfully translated into clinical drugs. However, the enediyne family of anticancer antibiotics is relatively small and consists of 11 structurally characterized members and four additional members isolated in their cycloaromatized form known to date. Therefore, it is a great challenge to develop innovative methods to discover new enediynes and produce them in sufficient quantities for chemical, biological, and clinical investigations. Strain prioritization and genome mining of 3,400 strains for enediyne natural products from the actinomycetes strain collection at The Scripps Research Institute (TSRI) led to the identification of 81 potential enediyne producers. Consecutive genome sequencing of 31 representative strains revealed at least 28 distinct enediyne biosynthetic gene clusters. By constructing an enediyne genome neighborhood network (GNN) we were able to predict new structural features leading to the discovery of one new enediyne natural product. Tiancimycin (TNM) is a 10-membered enediyne discovered from Streptomyces sp. CB03234. TNM exhibits potent cytotoxicity against a broad spectrum of cancer cell lines and kills selected cancer cells more rapidly and completely than the payloads used in FDA-approved antibody drugs. We were able to improve the production of TNMs in sufficient quantities and elucidate the biosynthesis of TNM by genetic manipulation. C-1027, a 9-membered enediyne was first discovered in Streptomyces globisporus in 1993. Genome mining of the 81 potential enediyne producers led to the discovery of four new C-1027 producers with titers of up to 11-fold higher than the original producer. These new C-1027 producing strains will supply C-1027 for current preclinical studies, future clinical trials, and eventual commercialization, and facilitate future biosynthetic studies for engineered production of designer C-1027 analogues with improved pharmacokinetic properties or new modes of action. Parts of the findings made during the Forschungsstipendium have been highlighted in the News & Views section of TSRI.
Publications
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(2016) Crystal structure of SgcJ, an NTF2-like superfamily protein involved in biosynthesis of the 9-membered enediyne antitumor antibiotic C-1027, J. Antibiot., 69:731-740
Huang, T.; Chang, C.-Y.; Lohman, J. R.; Rudolf, J. D.; Kim, Y.; Chang, C.; Yang, D.; Ma, M.; Yan, X.; Crnovcic, I.; Bigelow, L.; Clancy, S.; Bingman, C. A.; Yennamalli, R. M.; Babnigg, G.; Joachimiak, A.; Phillips Jr., J. N.; Shen, B.
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(2016) Engineered production of cancer targeting peptide (CTP)-containing C-1027 in Streptomyces globisporus and biological evaluation. Bioorg. Med. Chem., 24:3884-3892
Li, W.; Li, X.; Huang, T.; Teng, Q.; Crnovcic, I.; Rader, C.; Shen, B.
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(2016) Strain prioritization and genome mining for enediyne natural products. mBio., 7:e2104-16
Yan, X.; Ge, H.; Huang, T.; Hindra; Yang, D.; Teng, Q.; Crnovcic, I.; Li, X.; Rudolf, J. D.; Lohman, J. R.; Gansemans, Y.; Zhu, X.; Huang, Y.; Zhao, L.-X.; Jiang, Y.; Van Nieuwerburgh, F.; Rader, C.; Duan, Y.; Shen, B.
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(2017) "Chapter 3, The Role of Combinatorial Biosynthesis in Natural Products Discovery" in Chemical Biology of Natural Products, Cragg, G.; Newman, D.; Grothaus, P. Eds, Taylor and Francis, pp87-125
Rudolf, J. D.; Crnovcic, I.; Shen, B.
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(2017) Genome mining of Micromonospora yangpuensis DSM 45577 as a producer of an anthraquinone-fused enediyne. Org. Lett., 19:6192-6195
Yan, X.; Chen, J.-J.; Adhikari, A.; Yang, D.; Crnovcic, I.; Wang, N.; Chang, C.-Y.; Rader, C.; Shen, B.
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(2018) Discovery of alternative producers of the enediyne antitumor antibiotic C-1027 with high titers. J. Nat. Prod., 81:594-599
Yan, X.; Hindra; Ge, H.; Yang, D.; Huang, T.; Crnovcic, I.; Chang, C.-Y.; Fang, S.; Annaval, T.; Zhu, X.; Huang, Y.; Zhao, L.-X.; Jiang, Y.; Duan, Y.; Shen, B.
