Modular Strategies for Structure and Function Employed by Marine Cyanobacteria: Characterization and Synthesis of Pitinoic Acids

Abstract

Novel bioactive lipids were identified from a Guamanian cyanobacterium; the Pseudomonas aeruginosa quorum sensing inhibitor pitinoic acid A (1) and the anti-inflammatory pitinoic acids B (2) and C. The structure of 2 was confirmed by synthesis, which also allowed for biological evaluation. Since 2 is an ester of pitinoic acids A and C, it represents a prodrug strategy to liberate dual biological activity for the management of P. aeruginosa infections and their associated inflammation.Pseudomonas aeruginosa is a Gram-negative bacterium that causes opportunistic infections in different host tissues and organs. For example, cystic fibrosis (CF) infections and microbial keratitis (MK) are two of the most serious lung and eye infections, respectively, caused by this organism. 1, 2 Bacterial quorum sensing (QS) has been shown to play a major role in bacterial pathogenesis, where the infection and tissue destruction are facilitated by several QS-regulated virulence factors including elastase (LasB) and the pigment pyocyanin. 2,3 Tissue damage is also aggravated by the host's defense response, where the release of pro-inflammatory mediators and the prolonged inflammatory response are other contributors to tissue pathology. 2, 4-8 Accordingly, inhibiting QS pathways in P. aeruginosa and simultaneously controlling the associated inflammatory response is a promising strategy for the treatment of infections caused by this organism.luesch@cop.ufl.edu. Supporting Information Available. 1D and 2D NMR spectra for compounds 1 and 2, supporting figures including spectral data and characterization of synthetic intermediates, experimental procedures and additional RT-qPCR data. This material is available free of charge via the Internet at http://pubs.acs.org. During our efforts to discover novel bioactive compounds from marine cyanobacteria, we explored a population of a marine cyanobacterium morphologically similar to Lyngbya sp. NIH Public Accesscollected from a channel at the north end of Piti Bay at Guam, through NMR-guided fractionation. We isolated and characterized the novel lipids pitinoic acid A (1) (5-methylene decanoic acid) and its chlorinated ester, pitinoic acid B (2). The fatty acid 1 inhibits QS in P. aeruginosa, while the ester 2 prevents the induction of pro-inflammatory cytokine expression in LPS-induced THP-1 macrophages. In addition, we were able to identify the presence of pitinoic acid C, the alcohol moiety in 2, in one HPLC-fraction, which also maintained the anti-inflammatory effect detected for 2. Based on this, 2 appears to be a nature-designed prodrug which could deliver two essential moieties with different bioactivities upon hydrolysis; the QS-inhibitory module pitinoic acid A (1) and the antiinflammatory module pitinoic acid C.The EtOAc-MeOH extract was subjected to solvent-solvent partitioning followed by fractionation using silica gel chromatography. 1 H NMR profiles of the generated silica fractions showed a major simple fatty acid (1) HPLC purification of another silica fraction that elut...