The molecular approach to curing diseases

For this reason, the approach of today's pharmaceutical research is far more simplistic. The aim is to regulate a single protein. In some cases we aim at completely blocking an enzyme. To this end, we can provide a drug molecule that effectively competes with the natural substrate of the enzyme. The drug molecule, the so-called inhibitor, has to be made up such that it binds more strongly to the protein than the substrate. Then, the binding pockets of most enzyme molecules will contain drug molecules and cannot catalyze the desired reaction in the substrate. In some cases, the drug molecule even binds very tightly (covalently) to the enzyme (suicide inhibitor). This bond persists for the remaining lifetime of the protein molecule. Eventually, the deactivated protein molecule is broken down by the cell and a new identical enzyme molecule takes its place. Aspirin is an example of a suicide inhibitor. The effect of the drug persists until the drug molecules themselves are removed from the cell by its metabolic processes, and no new drug molecules are administered to replace them. Thus, one can control the effect of the drug by the time and dose it is administered.

There are several potent inhibitors of DHFR. One of them is methotrexate. Figure 1.5 shows methotrexate (color) both unbound (left) and bound (right) to DHFR (black and white). Methotrexate has been administered as an effective cytostatic cancer drug for over two decades.

7.2 The molecular approach to curing diseases I 9

7.2 The molecular approach to curing diseases I 9

Methotrexate (colored by surface potential, see Figure 3) and bound DHFR (gray)

There are many other ways of influencing the activity of a protein by providing a drug that binds to it. Drugs interact with all kinds of proteins:

• with receptor molecules that are located in the cell membrane and fulfill regulatory tasks

• with ion channels and transporter systems (again proteins residing in the cell membrane) that monitor the flux of molecules into and out of the cell

The mode of interaction between drug and protein does not always have the effect to block the protein, but we are generally looking for drugs that bind tightly to the protein.

Most drugs that are on the market today modify the enzymatic or regulatory action of a protein by strongly binding to it as described above. Among these drugs are long-standing, widespread and highly popular medications and more modern drugs against diseases such as AIDS, depression, or cancer. Even the life-style drugs that came into use in the past few years, such as Viagra and Xenical, belong to the class of protein inhibitors.

In this view, the quest for a molecular therapy of a disease decomposes into two parts:

Question 1: Which protein should we- target? As we have seen, there are many thousands candidate proteins in the human. We are looking for one of them that, by binding the drug molecule to it, provides the most effective remedy of the disease. This protein is called the target protein. Question 2: Which drug molecules should be used to bind to the target protein? Here, the molecular variety is even larger. Large pharmaceutical companies have compound archives with millions of compounds at their disposal. Every new target protein raises the questions, which of all of these compounds would be the best drug candidate. Nature is using billions of molecules. With the new technology of combinatorial chemistry, where compounds can be synthesized systematically from a limited set of building blocks, this number of potential drug candidates is also becoming accessible to the laboratory.

We will now give a short summary of the history of research on both of these questions.

Finding protein targets

Question 1 could not really be asked realistically until a few years ago. Historically, few target proteins were known at the time that the respective drug has been discovered. The reason is that new drugs were developed by modifying known drugs, based on some intuitive notion of molecular similarity. Each modification was immediately tested in the laboratory either in vitro or in vivo. Thus, the effectiveness of the drug could be assessed without even considering the target protein. To this day, all drugs that are on the marketplace world-wide target to an estimated set of 500 proteins [1]. Thus the search for target proteins is definitely the dominant bottleneck of today's pharmaceutical research.

Today, new experimental methods of molecular biology, the first versions of which have been developed just a few years ago, afford us with a fundamentally new way - the first systematic way - of looking for protein targets. We exemplify this progress at a specific DNA chip technology [2]. However, the general picture extends to many other experimental methods under development.

Figure 1.6 shows a DNA-chip that provides us with a differential census of the proteins manufactured by a yeast cell in two different cell states, one governed by the presence of glucose (green) and one by the absence of glucose (red). In effect the red picture is that of a starving yeast cell, whereas the green picture show the "healthy" state. Each bright green dot stands for a protein that is manufactured (expressed) in high numbers in the "healthy" state. Each bright red dot stands for a protein that is expressed

A DNAchip (from http://cmgm.stanford.edu/pbrown/ explore/)

A DNAchip (from http://cmgm.stanford.edu/pbrown/ explore/)

highly in the starving cell. If the protein occurs frequently in both the healthy and the starving state, the corresponding dot is bright yellow (resulting from an additive mixture of the colors green and red). Dark dots stand for proteins that are not frequent, the tint of the color again signifies whether the protein occurs more often in the healthy cell (green), equally often in both cells (yellow) or more often in the diseased cell (red).

At this point it is of secondary importance, exactly which experimental procedures generate the picture of Figure 1.6. What is important is, how much information is attached to colored dots in the picture. Here, we can make the following general statements.

1. The identity of the protein is determined by the coordinates of the colored dot. We will assume, for simplicity, that dots at different locations also represent different proteins. (In reality, multiple dots that represent the same protein are introduced, on purpose, for the sake of calibration.) The exact arrangement of the dots is determined before the chip is manufactured. This involves identifying a number of proteins to be represented on the chip and laying them out on the chip surface. This layout is governed by boundary conditions and preferences of the experimental procedures and is not important for the interpretation of the information. 2. Only rudimentary information is attached to each dot. At best, the experiment reveals the complete sequence of the gene or protein. Sometimes, only short segments of the relevant sequence are available.

Given this general picture, the new technologies of molecular biology can be classified according to two criteria, as shown by the next subsections.

Continue reading here: Genomics vs proteomics

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