A Yeast‐Based Assay for Inhibitors of l‐Lactate Transport Utilizing Fluorescent Biosensors

Inhibitors of ʟ-lactate transport are in development as a novel mode of action in antitumor therapy and malaria. Previously, we used radiolabeled ʟ-lactate to assay transport via the human monocarboxylate transporter 1, MCT1, and the structurally unrelated malaria parasite's transporter, PfFNT. We encountered a sensitivity limit at IC50 around 100 nM possibly resulting from the required high cell number per sample. Here, we describe a sensitive background-free high-throughput assay in yeast based on fluorescent iLACCO biosensors. We used iLACCO for co-expression and fusions with the transporter protein. Uptake of ʟ-lactate produced strong intensiometric fluorescent responses that could be monitored in cell suspensions using a fluorometer and in individual cells by fluorescence microscopy. The signal decreased dose-dependently in the presence of specific MCT1 and PfFNT inhibitors. Re-evaluation of 36 PfFNT inhibitors yielded IC50 values below 100 nM now matching previous data on Ki compound affinity to isolated transporter protein.

Rechte

Nutzung und Vervielfältigung:


CC BY-NC-ND 4.0

Bitte beachten Sie, dass einzelne Bestandteile der Publikation anderweitigen Lizenz- bzw. urheberrechtlichen Bedingungen unterliegen können.

Zitieren

Zitierform:
Tiedjens, F., Menzel, M., Stahnke, P., Grotewold, H., Uzun, C., Yildirim, D., Beitz, E., 2025. A Yeast‐Based Assay for Inhibitors of l‐Lactate Transport Utilizing Fluorescent Biosensors. ChemMedChem 20. https://doi.org/10.1002/cmdc.202400918
Zitierform konnte nicht geladen werden.