High Throughput Screening for Glycogen and Polyglucosan

H. Orhan AKMAN, PhDLaboratory of Dr. Salvatore DiMauro

Columbia University Medical Center

New York, NY

High Throughput Screening (HTS)

HTS is a tool for drug discovery.

Recent advancements in computer science combined with robotics and fluid delivery systems allowed to test the effects of thousands of compounds or known as chemical libraries.

Components of HTS:

1-Robotics

2-Liquid handling devices

3-Detectors

4-Data processing and control software

HTS Allows

• A researcher to quickly conduct millions of chemical, genetic or pharmacological tests.

• To identify active compounds, antibodies or genes which modulate a pathway.

• To establish starting points for drug design by understanding the interaction or role of a particular biochemical process in biology.

Preparation for HTS

• Assay preparation

• Reaction observation

• Automation systems

• Experimental design and data analysis

a. Quality control

b. Hit selection

Assay Preparation and Observation

Assay to observe the changes in quality and quantity of glycogen.

How Do We Measure the Quality of Glycogen?

• Limited digestion with diastase and staining with Per iodic Schiff base (PAS)

Klein et al. Muscle Nerve. 2004 Feb;29(2):323-8

Generating Computer Friendly Assay for HTS

Cultured Mouse Fibroblasts

Genetic and Biochemical Properties of Mouse Fibroblasts

• This fibroblast line is compound heterozygous for Y329S mutation and Exon 7 deletion

• GBE activity is less than 8% of controls.

Exon 7Intron 6 PGK-Neo

Intron 6 Intron 7

Intron 7

There is no Polyglucosan in Control MEF

Currently

We are testing 1700 FDA approved compounds.

Tests are being done in triplicates.

Assay plates are being analyzed.