Phenotype-Guided Natural Products Discovery Using Cytological Profiling

Abstract

Phenotype-guided natural products discovery is emerging as a useful new discovery tool that addresses challenges in early, unbiased natural product biological annotation. These high-content approaches yield screening results that report directly on the impact of test compounds on cellular processes in target organisms and can be used to predict the modes of action of bioactive constituents from primary screening data. In this study we explored the use of our recently implemented cytological profiling platform for the isolation of compounds with a specific, predefined mode of action, namely, induction of mitotic arrest. Screening of a microbially derived extract library revealed six extracts whose cytological profiles clustered closely with those of known antimitotic agents from the pure compound training set. Subsequent examination of one of these extracts revealed the presence of two separate bioactive constituents, each of which possessed a unique cytological profile. The first, diketopiperazine XR334 (3), recapitulated the observed antimitotic phenotype of the original extract, demonstrating that cytological profiling can be used for the targeted isolation of compounds with specific modes of action. The second, nocapyrone L (6), possessed a cytological profile that clustered with known calcium channel modulators, in line with previous published activities for this compound class, indicating that cytological profiling is a flexible and powerful platform for the de novo characterization of compound modes of action.

Marine natural products

“…165 The initial development of cytological screening of natural product extracts using a high content imaging approach to generate phenotype ngerprints has been extended from the original 312 extracts 166 to over 5000 pre-fractionated extracts from marine Actinobacteria and demonstrated the role that untargeted cytological screening can play in ascertaining the pathways and the mechanisms disrupted and so leading to a targeted selection of extracts based on a potential mode of action. 167…”

Kalkipyrone B, a marine cyanobacterial γ-pyrone possessing cytotoxic and anti-fungal activities

“…It will be interesting to observe if these structure-activity relationships extend to other types of biological activity. For example,the nocayrones B, H and L, γ-pyrone containing polyketides from marine Nocardiopsis strains, were recently shown to be Ca 2+ channel modulators (Ochoa et al, 2015; Kim et al, 2014). …”

Improving natural product research translation: From source to clinical trial

“…The application to chemically complex NPs of agnostic (or non-targeted) in vitro assays 69 combining cutting-edge computational approaches, optimized mass spectrometric (MS) data collection and analysis to elucidate chemical composition, and cytological profiling 70,71 now enables the generation of strong and specific hypotheses regarding bioactive NP components and the mechanisms through which they affect gene expression or cell phenotype. The application to chemically complex NPs of agnostic (or non-targeted) in vitro assays 69 combining cutting-edge computational approaches, optimized mass spectrometric (MS) data collection and analysis to elucidate chemical composition, and cytological profiling 70,71 now enables the generation of strong and specific hypotheses regarding bioactive NP components and the mechanisms through which they affect gene expression or cell phenotype.…”