Label-free human cell-based screening platform for SGLT1 and GLUT5 carbohydrate transporter modulators

Overview

This research proposes the development of a stable human cell-based screening system that uses auto-fluorescent, self-reporting SGLT1 and GLUT5 carbohydrate transporters to identify compounds that modulate sugar transport activity. SGLT1 and GLUT5 are key human carbohydrate transporters involved in glucose and fructose uptake, making them important targets for metabolic disease drug discovery. The proposed platform enables real-time measurement of carbohydrate influx through fluorescence monitoring, providing a label-free alternative to conventional glucose flux assays that rely on radioactivity, external labelling, or surrogate molecules. The stable nature of the cell lines ensures a practically inexhaustible screening resource suitable for high-throughput workflows.

Technical specifications

Key features:

• Recombinant stable human cell lines (HEK293/CaCO2) expressing active chimeric SGLT1 and GLUT5 transporters
• Auto-fluorescent reporter system using a modified GFP variant fused with a carbohydrate-binding protein, positioned proximal to the cytosolic side of the transporter pore
• Real-time fluorescence readout of carbohydrate influx without external labelling, radioactivity, or surrogate substrates
• Opto-genetic, label-free detection mechanism that measures transporter activity before carbohydrate metabolism
• Compatible with high-throughput screening in multi-well plate format (>25 compounds per run)
• Protein engineering approach leveraging published SGLT1 structural data for precise reporter placement

Technology readiness level

The platform is currently at an early research stage (TRL 2-3). The research team has demonstrated prior expertise in constructing functional GFP fusion proteins and designing reporter molecules for live cell imaging of intracellular metabolites, though no specific SGLT1/GLUT5 work has been completed. A 24-month validation roadmap is planned: Milestone 1 (0-12 months) focuses on cloning, transfection, and stable cell line construction with clone screening via GFP fluorescence, Western blot, and FACS. Milestone 2 (13-18 months) validates transporter activity using commercial glucose flux assay kits and demonstrates equivalence between GFP fluorescence measurements and surrogate substrate methods using known pharmacological modulators. Milestone 3 (19-24 months) establishes high-throughput screening methodology in multi-well plate format.