Spirocyclic β-lactone secondary metabolites modulate spliceosome function

Area of Science:

• Natural Product Chemistry
• Molecular Pharmacology
• Cancer Biology

Background:

• Spirocyclic β-lactones (SβLs) are cytotoxic natural products with an unknown mechanism of action.
• Actinomycetota, specifically Streptomyces platensis, produce SβLs via cryptic gene clusters.

Purpose of the Study:

• To elucidate the mechanism of action of SβLs.
• To discover new SβLs and characterize their structure-activity relationships.
• To explore the therapeutic potential of SβLs in cancer.

Main Methods:

• Isolation and structure elucidation of SβLs from Streptomyces platensis.
• Single cell Multiplexed Activity Metabolomics to assess cell death markers.
• Cytotoxicity assays and phospho-flow cytometry for structure-activity relationship analysis.
• Global cellular thermal proteome profiling and alternative splicing analysis.

Main Results:

• Several SβLs, including a new variant, were isolated and correlated with a cryptic biosynthetic gene cluster for oxazolomycin D (OxD) family metabolites.
• SβLs possess two pharmacophores: a polyketide component for target engagement and a β-lactone warhead for cytotoxicity.
• OxD treatment strongly interacted with and modulated spliceosome-associated proteins, leading to functional changes in RNA splicing.
• Phosphoprotein-S6 phosphorylation inhibition was observed, indicating a specific cellular response.

Conclusions:

• Spirocyclic β-lactones exhibit dual pharmacophores, contributing to their potent cytotoxicity and target engagement.
• Oxazolomycin D and related SβLs modulate the spliceosome, impacting RNA splicing.
• The spliceosome-associated cellular response to OxD presents a novel therapeutic opportunity for cancers with aberrant RNA splicing.