Knock-down vs knock-in

A brief overview of Galapagos

Figure 1: Comparison of a specific protein’s amount before and after knock - in or kn ock - down.

Knock-down viruses work by making small pieces of RNA (siRNA), which bind to a specific RNA sequence that produces a particular protein. After the siRNA binds the RNA, the RNA is cut by the cell’s machinery, therefore stopping the RNA from making anymore of the protein. This process is called RNA interference (RNAi) and results in a reduction in the amount of the specified protein. For the knock-in viruses, a specific piece of DNA (which contains the instructions to produce a specific protein) is inserted into the virus. After the virus infects the cell, the cell starts to produce more of the corresponding protein.

DNA DNA

RNA RNA

protein protein

DNA DNA

RNA RNA

protein protein

Knock Knock - - in in Knock Knock - - down down

DNA DNA

RNA RNA

protein protein

SilenceSelect SilenceSelect ® ® FLeXSelect FLeXSelect ® ®

amount of protein amount of protein in normal cells in normal cells

amount of protein amount of protein after knock after knock - - in in

amount of protein amount of protein after knock after knock - - down down

Galapagos is a drug discovery and development company focused on finding new ap- proaches to tackle difficult to cure diseases. This summary gives a brief introduction to Galapagos’ technology and explains how its unique approach could help bring new drugs to the market. The human genome , or blueprint for all of the cells in the human body, is made up of tens of thousands of genes. Genes are composed of DNA and contain the instructions for building all of the cell’s proteins. Protei ns control the structure and function of all of the cells that make up the human body. Before a cell can use the DNA to make protein, the DNA must first be re - written into RNA . The cell’s machinery then uses the RNA to produce protein. Therefore, the proce ss of making protein in the cell goes from DNA to RNA to protein (see the yellow box in Figure 1). Nearly all diseases and disorders are caused by a disruption in the normal function of certain proteins. Therefore, the main goal of pharmaceutical compani es is to design drugs that alter the activity of these proteins so that normal function returns and the cause of the disease is minimized or eliminated. One of the main obstacles in discovering new drugs is knowing exactly which of the body’s thousands of proteins play a key role in a particular disease. Once these proteins are discovered, they become targets for drug design. Finding these targets is one of the critical steps in the drug discovery process.

In order to study proteins in human cells, Galapa gos takes advantage of the distinctive properties of adenoviruses . Adenovirus is the virus that causes the common cold and has the capability to infect almost every type of human cell, which is the reason it is so contagious. The adenoviruses Galapagos wor ks with have been engineered to act as a shuttle vehicle, allowing the delivery of specific pieces of DNA into human cells. Additionally, these viruses have been made replication incompetent , meaning they are unable to reproduce outside of the laboratory e nvironment, and are therefore a safe vehicle for the delivery of DNA into human cells. Figure 1 shows how the viruses deliver these pieces of DNA into human cells in the laboratory, causing the cells to either make more of a certain protein ( knock - in ) or t o block the production of new protein ( knock - down ).

Galapagos combines the ability to alter the amount of protein in human cells with a high - throughput screening method. Using this method, Galapagos is able to run many small-scale experiments simultaneously. Operating on a grid system, where each square corresponds to a small test tube with cells that have undergone a change in a specific protein, Galapagos can screen many thousands of proteins in a short period of time.

Figure 2: High - throughput screening technique: using adenoviral collections to screen human cells.

Viruses containing specific pieces of DNA are Viruses containing specific pieces of DNA are arranged on a grid. arranged on a grid.

Each virus delivers this specific piece of DNA Each virus delivers this specific piece of DNA into the cells, causing a particular protein to be into the cells, causing a particular protein to be knocked knocked - - in or knocked in or knocked - - down. Therefore, each down. Therefore, each square on the grid contains cells that have square on the grid contains cells that have undergone a change in the amount of undergone a change in the amount of a specific protein. a specific protein.

Experiments are designed to monitor (by, for Experiments are designed to monitor (by, for example, a color change) the effect the protein example, a color change) the effect the protein change has on the cells. Because this change change has on the cells. Because this change corresponds to a specific protein, the protein corresponds to a specific protein, the protein can be further investigated can be further investigated and eventually and eventually become a drug target. become a drug target.

Based on their function and design, proteins are divided into several classes. Researchers in the pharmaceutical industry generally agree that certain classes of proteins respond to drugs better than other classes. These classes of proteins are referred to as drugable and form the basis of most drug research programs. Galapagos has constructed a focused collection of viruses that either knock - in ( FLeXSelect ® ) or knock - down ( SilenceSelect ® ) the dru gable proteins. With these collections, Galapagos has screened human cells that are affected by diseases like osteoarthritis, osteoporosis and rheumatoid arthritis, and has identified proteins that control the disease. By further testing (validating) these proteins in more advanced studies, Galapagos has found a number of targets suitable for new drug development. Galapagos is currently conducting drug discovery research based on the targets discovered using this technology. Once a target is validated, it is tested against large collections of chemical small molecules to identify the structures that interact (block or activate) with the target. These chemical structures are then optimized to obtain “drug - like” characteristics followed by testing of the dru g candidate in the clinic. This process of drug discovery is similar to the approach taken by large pharmaceutical companies and has resulted in breakthrough medicines such as Gleevec , a block- ®

buster oncology drug developed by Novartis.

Galapagos’ uniquene ss lies in using human cells, which gives a more realistic idea of the effect that protein might have on the disease in the human body than studying proteins in engineered cells (cell lines) and animal cells, as other companies do. Moreover, Galapagos conc entrates its efforts on the drugable proteins and can efficiently screen these proteins in human cells. Galapagos believes that this unique approach to target identification and validation increases the chances of success in bringing new drugs to the marke t. In addition to forming the basis of Galapagos’ proprietary & alliance programs, these adenoviral collections and screening technologies are also available to not-for-profit organizations and biotech & pharma companies through BioFocus, a Galapagos service division.

Numerous partners have already applied Galapagos’ technology across a number of disease areas, aiding the scientific and pharmaceutical communities to better understand the cause of disease and further progressing the development of new drugs.