Among marine organisms, sponges are the most prolific producers of marine natural products (MNPs) which are recently becoming an attractive source of drug discovery. One probable reason is the huge amount of diverse microorganisms symbiotically living inside a sponge provide an extremely rich reservoir of natural product biosynthetic genes. However, the supply of a potent MNP is often the major issue banning it from further clinical studies. My laboratory focuses on the biosynthesis of MNPs from deep-sea sponges to identify biosynthetic origin and pathways and ultimately to develop sustainable supplies for them. Leiodermatolide (LDM), isolated from the sponge Leiodermatium sp. (sampled at ~400 m), is a trans-AT (acyl transferase) type-I polyketide, in which AT domains are encoded by separate genes, instead of generally integrated into the architecture of polyketide synthases (PKSs). LDM is a potent mitosis-targeting agent and shows strong in vitro inhibitory activity against cancer cell proliferation. Notably, LDM is potent against the leukemia cell line HEL92.1.7 with a GI50 of 1.0 nM; this cell line contains the Pgp efflux transporter which is a major cause of drug resistance in chemotherapy. The in vivo antitumor activity of LDM was recently confirmed in a mouse model of metastatic pancreatic cancer. LDM arrests cell division at the G2/M phase. Notably, LDM employs a distinct mode of action that fundamentally differs from current anti-mitotic drugs including taxanes and vinca alkaloids. LDM doesn’t bind to tubulin, nor does it induce or inhibit tubulin polymerization in vitro. LDM also causes an immediate block of microtubule elongation. A fosmid library of LDM-producing sponge Leiodermatium was constructed. The library is being screened by primers specifically targeting the highly conversed KS (ketosynthase) domains of trans-AT type I PKSs and other specific structural features. A set of fosmids were obtained. Cloning, sequencing, and restriction analysis indicated that those fosmids share common sequences but each with a unique restriction pattern. A 24-kb DNA sequence was identified, which are likely involved in the biosynthesis of a trans-AT type I polyketide. Domains such as KS, ACP (acyl carrier protein), KR (keto-reductase) and DH (dehydrogenase) were identified, but lacing AT domains. Sequencing of additional fosmids is in the process. A proposed biosynthetic pathway of LDM and future strategies for sustainable supplies will also be discussed.

Biosynthesis of deep-sea marine natural products: Genes, enzymes and pathways

254th National Meeting (2017)

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