Leonard A. McDonald

Biomedical Potential of Marine Natural Products

Marine natural products, the secondary or nonprimary metabolites produced by organisms that live in the sea, have received increasing attention from chemists and pharmacologists during the last two decades. Interest on the part of chemists has been twofold: natural products chemists have probed marine organisms as sources of new and unusual organic molecules, while synthetic chemists have followed by targeting these novel structures for development of new analogs and new synthetic methodologies and strategies (Albizati et al., 1990). The rationale for investigating the chemistry of marine organisms has changed over the past several decades. Early investigations were largely of a “phytochemical” nature, reporting detailed metabolite profiles similar to those reported for terrestrial plants in previous decades. However, analogous to investigations of terrestrial plants, more recent studies of marine organisms have focused on their potential applications, particularly to the treatment of human disease and control of agricultural pests (Fautin, 1988). Pharmacological evaluations of marine natural products have likewise undergone an evolution over the past two decades: beginning with the early investigations of toxins, followed by studies of cytotoxic and antitumor activity, to the present day, where a myriad of activities based on whole-animal models and receptor-binding assays are being pursued. The intent of this chapter is to look back at the evolution of biomedically oriented natural product studies of marine organisms, to chronicle the key developments, discoveries, and advances in the level of sophistication that have fueled further interest in this field, and finally to look forward at the future biomedical potential of marine natural products.

Binding of rapamycin analogs to calcium channels and FKBP52 contributes to their neuroprotective activities

Rapamycin is an immunosuppressive immunophilin ligand reported as having neurotrophic activity. We show that modification of rapamycin at the mammalian target of rapamycin (mTOR) binding region yields immunophilin ligands, WYE-592 and ILS-920, with potent neurotrophic activities in cortical neuronal cultures, efficacy in a rodent model for ischemic stroke, and significantly reduced immunosuppressive activity. Surprisingly, both compounds showed higher binding selectivity for FKBP52 versus FKBP12, in contrast to previously reported immunophilin ligands. Affinity purification revealed two key binding proteins, the immunophilin FKBP52 and the β1-subunit of L-type voltage-dependent Ca2+ channels (CACNB1). Electrophysiological analysis indicated that both compounds can inhibit L-type Ca2+ channels in rat hippocampal neurons and F-11 dorsal root ganglia (DRG)/neuroblastoma cells. We propose that these immunophilin ligands can protect neurons from Ca2+-induced cell death by modulating Ca2+ channels and promote neurite outgrowth via FKBP52 binding.

Pyranonaphthoquinone Lactones: A New Class of AKT Selective Kinase Inhibitors Alkylate a Regulatory Loop Cysteine

The naturally occurring pyranonaphthoquinone (PNQ) antibiotic lactoquinomycin and related aglycones were found to be selective inhibitors of the serine-threonine kinase AKT. A set of synthetic PNQs were prepared and a minimum active feature set and preliminary SAR were determined. PNQ lactones inhibit the proliferation of human tumor cell lines containing constitutively activated AKT and show expected effects on cellular biomarkers. Biochemical data are presented supporting a proposed bioreductive alkylation mechanism of action.

Discovery of lactoquinomycin and related pyranonaphthoquinones as potent and allosteric inhibitors of AKT/PKB: mechanistic involvement of AKT catalytic activation loop cysteines

The serine/threonine kinase AKT/PKB plays a critical role in cancer and represents a rational target for therapy. Although efforts in targeting AKT pathway have accelerated in recent years, relatively few small molecule inhibitors of AKT have been reported. The development of selective AKT inhibitors is further challenged by the extensive conservation of the ATP-binding sites of the AGC kinase family. In this report, we have conducted a high-throughput screen for inhibitors of activated AKT1. We have identified lactoquinomycin as a potent inhibitor of AKT kinases (AKT1 IC50, 0.149 ± 0.045 μmol/L). Biochemical studies implicated a novel irreversible interaction of the inhibitor and AKT involving a critical cysteine residue(s). To examine the role of conserved cysteines in the activation loop (T-loop), we studied mutant AKT1 harboring C296A, C310A, and C296A/C310A. Whereas the ATP-pocket inhibitor, staurosporine, indiscriminately targeted the wild-type and all three mutant-enzymes, the inhibition by lactoquinomycin was drastically diminished in the single mutants C296A and C310A, and completely abolished in the double mutant C296A/C310A. These data strongly implicate the binding of lactoquinomycin to the T-loop cysteines as critical for abrogation of catalysis, and define an unprecedented mechanism of AKT inhibition by a small molecule. Lactoquinomycin inhibited cellular AKT substrate phosphorylation induced by growth factor, loss of PTEN, and myristoylated AKT. The inhibition was substantially attenuated by coexpression of C296A/C310A. Moreover, lactoquinomycin reduced cellular mammalian target of rapamycin signaling and cap-dependent mRNA translation initiation. Our results highlight T-loop targeting as a new strategy for the generation of selective AKT inhibitors. [Mol Cancer Ther 2007;6(11):OF1–11]

Spiroxins, DNA cleaving antitumor antibiotics from a marine-derived fungus

The spiroxins (1–5) were purified from the culture extract of a marine-derived fungus. Their unique bisnaphthospiroketal structures were established by NMR spectroscopy. In addition to cytotoxicity, these compounds showed antibiotic activity and were active in a mouse xenograft model against human ovarian carcinoma. The mechanism of action of these compounds was shown to be due, in part, to their effect on DNA.

Botryllamides A-D, new brominated tyrosine derivatives from styelid ascidians of the genus Botryllus

Abstract Four new bromotyrosine derivatives, botryllamides A-D (1–4) were isolated from the styelid ascidian Botryllus sp. from Siquijor Is., Philippines, and from Botryllus schlosseri from the Great Barrier Reef, Australia. Their structures were deduced from 1D and 2D NMR spectral data.

Discovery of (2S,4R)-1-(2-Aminoacetyl)-4-benzamidopyrrolidine-2-carboxylic Acid Hydrochloride (GAP-134)13, an Orally Active Small Molecule Gap-Junction Modifier for the Treatment of Atrial Fibrillation

Rotigaptide (3) is an antiarrhythmic peptide that improves cardiac conduction by modifying gap-junction communication. Small molecule gap-junction modifiers with improved physical properties were identified from a Zealand Pharma peptide library using pharmaceutical profiling, established SAR around 3, and a putative pharmacophore model for rotigaptide. Activity of the compounds was confirmed in a mouse cardiac conduction block model of arrhythmia. Dipeptide 9f (GAP-134) was identified as a potent, orally active gap-junction modifier for clinical development.

The lecanindoles, nonsteroidal progestins from the terrestrial fungus Verticillium lecanii 6144.

Four new indolosesquiterpenes, lecanindoles A-D (1-4), were isolated from fermentations of the terrestrial fungus Verticillium lecanii 6144. The structures of compounds 1-4 were elucidated from analysis of spectroscopic data. Compound 2 was reduced to give 4 and its isomer 5. Compound 4 was found to be a potent and selective progesterone receptor agonist with an EC50 of 1.1 +/- 0.4 nM in a cell-based luciferase reporter assay.

Fibrosterol Sulfates from the Philippine Sponge Lissodendoryx (Acanthodoryx) fibrosa; Sterol Dimers that Inhibit PKCζ

Three new sulfated sterol dimers, fibrosterol sulfates A-C (1-3), have been isolated from the sponge Lissodendoryx (Acanthodoryx) fibrosa, collected in the Philippines. The structures were assigned on the basis of extensive 1D and 2D NMR studies as well as analysis by HRESIMS. Compounds 1 and 2 inhibited PKCzeta with IC(50) values of 16.4 and 5.6 microM, respectively.

Stereoselective synthesis of an active metabolite of the potent PI3 kinase inhibitor PKI-179.

The synthesis and stereochemical determination of 1-(4-(4-((1R,5R,6R)-6-hydroxy-3-oxa-8-azabicyclo[3.2.1]octan-8-yl)-6-morpholino-1,3,5-triazin-2-yl)phenyl)-3-(pyridin-4-yl)urea (2), an active metabolite of the potent PI3 kinase inhibitor PKI-179 (1), is described. Stereospecific hydroboration of the double bond of 2,5-dihydro-1H-pyrrole 8 gave the 2,3-trans alcohol 9 exclusively. The configuration of the 3-hydroxyl group in 9 was inverted by an oxidation and stereoselective reduction sequence to give the corresponding 2,3-cis isomer 23. Both exo (21) and endo (27) isomers of the metabolite 2 were prepared via a practical synthetic route from 9 and 23, respectively, and the stereochemistry of 2 was determined to be endo. The endo isomer (27) was separated into two enantiomers 28 and 29 by chiral HPLC. Compound 2 was found to be enantiomerically pure and identical to the enantiomer 28. The absolute stereochemistry of the enantiomer 28 was determined by Moshers method, thus establishing the stereochemistry of the active metabolite 2.