On March 18, 2019, a groundbreaking study by Cochrane et al. published in ACS Combinatorial Science introduced activity-based DNA-encoded library (DEL) screening as a transformative paradigm shift in drug discovery. This research addressed key limitations of traditional affinity-based DEL methods by developing an integrated microfluidic platform that enables functional screening of complex targets like ion channels and protein complexes. The system leverages droplet compartmentalization, photochemical compound release, and real-time fluorescence detection to achieve high-throughput screening (HTS) at 30,000 beads per hour while using minimal reagents (< 10 μg enzyme). Validation through autotaxin inhibitor identification demonstrated practical efficacy, with 20 of 35 hits confirming activity and IC₅₀ values as low as 3.5 μM. This approach expands DEL applications to previously inaccessible target classes, combining chemical diversity with functional insights to accelerate therapeutic discovery.

Introduction

In the evolving landscape of drug discovery, DEL technology has emerged as a powerful tool for exploring vast chemical spaces. While traditional DEL screening relies on affinity-based selections, recent breakthroughs have expanded this paradigm to include activity-based screening - a transformative approach that opens new possibilities for investigating complex biological targets.

The Limitations of Conventional Screening Methods

HTS has long been the cornerstone of drug discovery, enabling researchers to test thousands of compounds against biological targets. However, HTS faces significant challenges, including high costs, substantial compound storage requirements, and limitations in accessing novel chemical diversity. Traditional DEL technology, while excellent for affinity-based selections, has been restricted to soluble, stable targets, leaving many important target classes – such as ion channels, receptors, and protein complexes - largely unexplored.

A Revolutionary Approach: Activity-Based DEL Screening

The research team led by Wesley G. Cochrane and Brian M. Paegel developed an integrated microfluidic platform that successfully bridges this technological gap. Their system combines the chemical diversity of DELs with the functional insights of activity-based screening, creating a powerful new tool for drug discovery.

The microfluidic circuit incorporates several innovative components that enable efficient activity-based screening:

• Droplet Generation and Compartmentalization

The system generates water-in-oil droplets at 60 Hz, each serving as a miniature reaction vessel for screening individual library members.

• Photochemical Compound Release

Library compounds are photochemically cleaved from solid-phase beads directly within the droplets, ensuring precise dosing and controlled release.

• Real-time Fluorescence Detection

A sophisticated detection system monitors enzymatic activity in each droplet, identifying hits through dynamic statistical hypothesis testing.

• High-throughput Sorting

The platform achieves an impressive throughput of 30,000 beads per hour, with the capability to sort hit-containing droplets for further analysis.

Fig.1 Architecture of a microfluidic circuit for HTS. (Cochrane, et al., 2019)

Application to Autotaxin Inhibition

The researchers demonstrated their platform's effectiveness by screening a 67,100-member DEL for inhibitors of autotaxin (ATX), a clinically relevant phosphodiesterase enzyme. The screen identified 35 high-priority hit compounds, with 20 validating as active ATX inhibitors upon resynthesis. Five selected compounds showed promising IC₅₀ values ranging from 3.5 to 10 μM.

Synergy with CD BioGlyco's DEGL Platform

The advancements demonstrated in this research align perfectly with CD BioGlyco's comprehensive DEGL services. Our platform offers several complementary advantages:

CD BioGlyco provides multiple DEGL design strategies, including:

• Chemistry-driven DEGL Design
• Biology-driven DEGL Design
• AI-based Library Optimization

Our services incorporate cutting-edge screening technologies that complement activity-based approaches:

• Magnetic bead-based affinity screening
• Cell-based DEGL screening
• Microfluidic processor-based screening

Advantages of the Integrated Approach

• Expanded Target Scope

Enables investigation of previously inaccessible target classes, including signaling pathways and cellular response systems.

• Reduced Resource Consumption

The miniaturized format uses vanishingly small quantities of reagents (<10 μg enzyme, <20 nmol substrate) and library material.

• Enhanced Statistical Power

Increased sampling capability overcomes Poisson-derived variances inherent in combinatorial libraries.

• Functional Insights

Provides direct information about compound activity rather than mere binding affinity.

Future Directions and Applications

The success of activity-based DEL screening opens exciting new possibilities for drug discovery:

• Phenotypic screening: Integration with cellular assays for more biologically relevant screening
• Complex pathway analysis: Investigation of multi-component biological systems
• Novel target identification: Discovery of compounds against challenging targets like protein-protein interactions

CD BioGlyco remains at the forefront of these developments, continuously expanding our DEGL capabilities to incorporate the latest technological advancements.

Conclusion

The pioneering work on activity-based DEL screening represents a significant milestone in drug discovery technology. By combining the chemical diversity of DNA-encoded libraries with functional activity assessment, researchers can now explore biological targets with unprecedented depth and efficiency. CD BioGlyco's DEGL platform builds upon these innovations, offering comprehensive services that leverage the latest advances in library design, screening, and validation.

As the field continues to evolve, the integration of activity-based screening approaches with sophisticated DEGL technologies promises to accelerate the discovery of novel therapeutic agents and deepen our understanding of complex biological systems.

Explore CD BioGlyco's DEGL services to leverage these cutting-edge technologies for your research needs. Visit our platform to learn more about our comprehensive DNA-encoded glycan library solutions.