Salinosporamide A: A Highly Cytotoxic Proteasome Inhibitor from a Novel Microbial Source, a Marine Bacterium of the New Genus Salinospora

Abstract

We recently reported the cultivation and phylogenetic characterization of a new group of obligate marine actinomycete bacteria that is widely distributed in ocean sediments.[1] Analogous soil-derived actinomycetes have been the single most significant source of naturally occurring microbial antibiotics, [2] thus the discovery of a major new group of these bacteria in marine sediments suggests that the ocean represents an overlooked habitat from which to isolate these important microorganisms. Given that the rate of discovery of new biologically active compounds from common soil actinomycetes has been falling, [3] obligate marine actinomycetes represent a new resource for structurally diverse secondary metabolites.To date, we have isolated in excess of 2500 strains belonging to this new taxon for which we have proposed the genus name ™Salinospora∫ (a formal taxonomic description is in progress). ™Salinospora∫ strains, with previously undescribed 16S rRNA gene sequences, have been recovered from five distinct tropical/subtropical ocean systems [1] and from depths as great as À1100 m, which indicates that they represent a widely distributed and taxonomically diverse group of sediment bacteria. All isolates display an obligate requirement of ionic sodium for growth, thus indicating a high level of marine adaptation.In preliminary screening, a high percentage of the organic extracts of cultured ™Salinospora∫ strains possessed antibiotic and anticancer activities, which suggests that these bacteria are an excellent resource for drug discovery. Herein we report the results of our first chemical investigation of a member of the ™Salinospora∫ group and show that strain CNB-392 produces the chemically unique and highly bioactive metabolite salinosporamide A (1, Scheme 1). Salinosporamide A exhibits potent cancer cell cytotoxicity and appears to exert its cytotoxic effects through inhibition of the 20S proteasome.™Salinospora∫ strain CNB-392 was isolated from a heattreated marine sediment sample that was plated on a seawater-based agar nutrient medium. Liquid shake flask cultivation of this strain, followed by solid-phase extraction with Amberlite resin (XAD-16) and elution with acetone, resulted in a crude extract that was highly cytotoxic in vitro toward HCT-116 human colon carcinoma (IC 50 ca. 80 ng mL À1 ). Cytotoxicity-guided fractionation of the crude extract led to the isolation of salinosporamide A (1) as a colorless crystalline solid (yield: 7 mg L À1 ). The complete structural assignment of 1 was accomplished by spectral analysis and by a single-crystal X-ray diffraction study.Analysis of the low-resolution mass spectrum of salinosporamide A showed a characteristic [Mþ2] þ peak indicative of the presence of a chlorine atom. High-resolution massspectral analysis provided the molecular formula Comprehensive analysis of 2D NMR data, including the results of COSY, HMQC, and HMBC experiments, enabled the complete planar structure of salinosporamide A to be assigned, as in 1, to a 2-aza-6-oxabicyclo[3.2.0]heptane-3,7...

A novel proteasome inhibitor NPI-0052 as an anticancer therapy

“…Initial screening of NPI-0052 against the NCI panel of 60 tumour cell lines showed GI 50 of o10 nM in all cases (Feling et al, 2003). In concert with these observations, our data showed that (1) NPI-0052 induces apoptosis in MM cells sensitive and resistant to conventional and Bortezomib therapies; and (2) IC50 of NPI-0052 for MM cells is within the low nanomolar concentration (Chauhan et al, 2005a).…”

“…Specifically, delineation of mechanisms mediating interaction of proteasome inhibitors with 20S proteasome will allow designing potent and therapeutically efficacious proteasome inhibitors. In this context, our recent study characterised a novel proteasome inhibitor NPI-0052, a small molecule derived from fermentation of Salinospora, a new marine Gram-positive actinomycete (Feling et al, 2003;Macherla et al, 2005;Chauhan et al, 2005a). Importantly, NPI-0052 is distinct from Bortezomib, orally bioactive, and an equipotent antitumour agent (Chauhan et al, 2005a).…”

“…A recent report describing the crystal structure of NPI-0052 in complex with the 20S proteasome revealed an important consequence of betalactone ring opening and a mechanism of irreversible binding vis-à-vis omuralide (Groll et al, 2006). Collectively, these structural differences account for the enhanced in vitro and in vivo potency of NPI-0052 compared to Omuralide (Feling et al, 2003;Macherla et al, 2005;Groll et al, 2006).…”

GC–MS Analysis of Bio-active Molecules Derived from Paracoccus pantotrophus FMR19 and the Antimicrobial Activity Against Bacterial Pathogens and MDROs

“…[6]; salinosporamide-A, an anticancer metabolite (Salinispora sp.) [7]; marinomycins (Marinophilus sp.) [8]; abyssomicin-C (Verrucosispora sp.)…”

Rapid Enrichment of Dehalococcoides-Like Bacteria by Partial Hydrophobic Separation

“…Enrichments or isolates facilitate better assessment of the physiology, structure, and function of populations within a complex microbial community. In fact, the study of enrichments and isolates has resulted in discoveries as diverse as determining the enzymes responsible for degradation of the pollutant vinyl chloride (4), isolating possible anticancer agents (5), and understanding the cell structures of bacteria within candidate phyla with no cultured representatives (6). Unfortunately, while culturing techniques have improved, they still rely heavily on knowledge of an organism's ideal growth conditions and tend to enrich the fastest growing organisms in a sample (7).…”