Abstract
Discovering bioactive metabolites within a metabolome is challenging because there is generally little foreknowledge of metabolite molecular and cell-targeting activities. Here, single-cell response profiles and primary human tissue comprise a response platform used to discover novel microbial metabolites with cell-type-selective effector properties in untargeted metabolomic inventories. Metabolites display diverse effector mechanisms, including targeting protein synthesis, cell cycle status, DNA damage repair, necrosis, apoptosis, or phosphoprotein signaling. Arrayed metabolites are tested against acute myeloid leukemia patient bone marrow and molecules that specifically targeted blast cells or nonleukemic immune cell subsets within the same tissue biopsy are revealed. Cell-targeting polyketides are identified in extracts from biosynthetically prolific bacteria, including a previously unreported leukemia blast-targeting anthracycline and a polyene macrolactam that alternates between targeting blasts or nonmalignant cells by way of light-triggered photochemical isomerization. High-resolution cell profiling with mass cytometry confirms response mechanisms and is used to validate initial observations.
Highlights
Discovering bioactive metabolites within a metabolome is challenging because there is generally little foreknowledge of metabolite molecular and cell-targeting activities
The biological characterization of such untargeted metabolomic arrays results in the generation of “bioactivity chromatograms”, and correlation analysis to matched extracted ion current (EIC) mass chromatograms identifies candidate metabolites linked to measured bioassay targets
Fluorescence cytometry has the advantage of cellular throughput and more robust barcoding potential[32], whereas mass cytometry has the power to track more than 35 key markers of acute myeloid leukemia (AML) cell phenotype and function simultaneously[23,33,34]
Summary
Discovering bioactive metabolites within a metabolome is challenging because there is generally little foreknowledge of metabolite molecular and cell-targeting activities. A combination of (1) phospho-flow, (2) single-cell chemical biology, and (3) cellular barcoding was matched with (4) metabolomic arrays to identify natural product secondary metabolites that target primary human leukemia cells and spare adjacent nonmalignant immune cells This activitymetabolomics platform is termed multiplexed activity metabolomics (MAM) and comprises a system for single-cell metabolome-scale analysis of bioactivity using human cells from primary tissue biopsies in a high-throughput screening-compatible microtiter format. This untargeted assay modality samples a cross section of biological responses in a heterogeneous mixture of cells representing an in vivo human tissue environment and has the potential to identify disease-relevant bioactive metabolites within metabolomic arrays
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