Ranjala Ratnayake

Englerin A, a selective inhibitor of renal cancer cell growth, from Phyllanthus engleri.

An extract from Phyllanthus engleri was identified in a bioinformatic analysis of NCI 60-cell natural product extract screening data that selectively inhibited the growth of renal cancer cell lines. Bioassay-guided fractionation yielded two new guaiane sesquiterpenes, englerins A (1) and B (2). Englerin A showed 1000-fold selectivity against six of eight renal cancer cell lines with GI(50) values ranging from 1-87 nM. The structures of 1 and 2 and their relative stereochemistry were established by spectroscopic methods.

Englerin A stimulates PKCθ to inhibit insulin signaling and to simultaneously activate HSF1: pharmacologically induced synthetic lethality.

The natural product englerin A (EA) binds to and activates protein kinase C-θ (PKCθ). EA-dependent activation of PKCθ induces an insulin-resistant phenotype, limiting the access of tumor cells to glucose. At the same time, EA causes PKCθ-mediated phosphorylation and activation of the transcription factor heat shock factor 1, an inducer of glucose dependence. By promoting glucose addiction, while simultaneously starving cells of glucose, EA proves to be synthetically lethal to highly glycolytic tumors.

Grassypeptolides A-C, cytotoxic bis-thiazoline containing marine cyclodepsipeptides.

Grassypeptolides A−C (1−3), a group of closely related bis-thiazoline containing cyclic depsipeptides, have been isolated from extracts of the marine cyanobacterium Lyngbya confervoides. Although structural differences between the analogues are minimal, comparison of the in vitro cytotoxicity of the series revealed a structure−activity relationship. When the ethyl substituent of 1 is changed to a methyl substituent in 2, activity is only slightly reduced (3−4-fold), whereas inversion of the Phe unit flanking the bis-thiazoline moiety results in 16−23-fold greater potency. We show that both 1 and 3 cause G1 phase cell cycle arrest at lower concentrations, followed at higher concentrations by G2/M phase arrest, and that these compounds bind Cu2+ and Zn2+. The three-dimensional structure of 2 was determined by MS, NMR, and X-ray crystallography, and the structure of 3 was established by MS, NMR, and chemical degradation. The structure of 3 was explored by in silico molecular modeling, revealing subtle differences in overall conformation between 1 and 3. Attempts to interconvert 1 and 3 with base were unsuccessful, but enzymatic conversion may be possible and could be a novel form of activation for chemical defense.

Acremolides A-D, Lipodepsipeptides from an Australian Marine-Derived Fungus, Acremonium sp

An Australian estuarine isolate of an Acremonium sp. (MST-MF588a) yielded the two known compounds 19- O-acetylchaetoglobosin D ( 1) and 19- O-acetylchaetoglobosin B ( 2), as the sole cytotoxic principles, along with the known aromatic metabolite RKB 3564S ( 3), and a novel family of lipodepsipeptides, acremolides A-D ( 4- 7). Structures were assigned to 4- 7 on the basis of detailed spectroscopic analysis and chemical derivatization and by application of a new C 3 Marfeys method for amino acid analysis.

Chlorinated englerins with selective inhibition of renal cancer cell growth.

The chlorinated englerins (3-9) were isolated from Phyllanthus engleri and shown to selectively inhibit the growth of renal cancer cells. The compounds were shown to be extraction artifacts produced by exposure to chloroform decomposition products during their isolation. The most active compound, 3, was synthesized from englerin A (1).

C3 and 2D C3 Marfey’s methods for amino acid analysis in natural products

We validate the improved resolution and sensitivity of the C3 Marfeys method, including an ability to resolve all Ile isomers, against an array of amino acids commonly encountered in natural products and by comparison to an existing Marfeys method. We also describe an innovative 2D C3 Marfeys method as an analytical approach for determining the regiochemistry of enantiomeric amino acid residues in natural products. The C3 and 2D C3 Marfeys methods represent valuable tools for probing and defining the stereocomplexity of hydrolytically accessible amino acid residues in natural products.

Grassypeptolides F and G, Cyanobacterial Peptides from Lyngbya majuscula

Grassypeptolides F (1) and G (2), bis-thiazoline-containing cyclic depsipeptides with a rare β-amino acid, extensive N-methylation, and a large number of d-amino acids, are reported from an extract of the Palauan cyanobacterium Lyngbya majuscula. Both 1 and 2 were found to have moderate inhibitory activity against the transcription factor AP-1 (IC₅₀ = 5.2 and 6.0 μM, respectively).

Inhibitors of the Oncogenic Transcription Factor AP-1 from Podocarpus latifolius

An activator protein-1 (AP-1) based bioassay-guided phytochemical investigation on Podocarpus latifolius led to the isolation of three new sempervirol-type diterpenes, cycloinumakiol (1), inumakal (2), and inumakoic acid (3), along with three known norditerpenes (4-6). Compounds 4 and 6 were responsible for the observed bioactivity.

Grassypeptolides as natural inhibitors of dipeptidyl peptidase 8 and T-cell activation.

Natural products made by marine cyanobacteria are often highly modified peptides and depsipeptides that have the potential to act as inhibitors for proteases. In the interests of finding new protease inhibition activity and selectivity, grassypeptolide A (1) was screened against a panel of proteases and found to inhibit DPP8 selectively over DPP4. Grassypeptolides were also found to inhibit IL‐2 production and proliferation in activated T‐cells, consistent with a putative role of DPP8 in the immune system. These effects were also observed in Jurkat cells, and DPP activity in Jurkat cell cytosol was shown to be inhibited by grassypeptolides. In silico docking suggests two possible binding modes of grassypeptolides—at the active site of DPP8 and at one of the entrances to the internal cavity. Collectively these results suggest that grassypeptolides might be useful tool compounds in the study of DPP8 function.

Intramolecular Modulation of Serine Protease Inhibitor Activity in a Marine Cyanobacterium with Antifeedant Properties

Extracts of the Floridian marine cyanobacterium Lyngbya cf. confervoides were found to deter feeding by reef fish and sea urchins (Diadema antillarum). This antifeedant activity may be a reflection of the secondary metabolite content, known to be comprised of many serine protease inhibitors. Further chemical and NMR spectroscopic investigation led us to isolate and structurally characterize a new serine protease inhibitor 1 that is formally derived from an intramolecular condensation of largamide D (2). The cyclization resulted in diminished activity, but to different extents against two serine proteases tested. This finding suggests that cyanobacteria can endogenously modulate the activity of their protease inhibitors.