Someone recently bought our

students are currently browsing our notes.


Honours Directed 2 Notes

Pharmacology Notes > Drug Development (BIOL10822) Notes

This is an extract of our Honours Directed 2 document, which we sell as part of our Drug Development (BIOL10822) Notes collection written by the top tier of University Of Manchester students.

The following is a more accessble plain text extract of the PDF sample above, taken from our Drug Development (BIOL10822) Notes. Due to the challenges of extracting text from PDFs, it will have odd formatting:

Drugs: From Molecules to Man - Lecture 10

*HD2* Molecular Techniques in Drug Discovery and Development

Expression systems (Xenopus, insect cells, yeast, bacteria, mammalian cell lines).
Transfection methods.
Role of cytochrome P450 enzymes.
CYP polymorphisms.
Effect of polymorphisms on drug metabolism and response.
Drug and diet interactions with CYP enzymes.

Drug Diversity

Most drug targets are members of protein families.
For example, there are four main classes of adrenoceptor.
These families of proteins arose via the process of gene duplication, redundancy and subsequent mutation.

Difficulties of Drug Design due to Diversity

If there are related proteins there is the potential for these drugs to interact with not just the specified target, but also its relatives, making it much harder to design drugs.

For example, it is known that noradrenaline binds to all four classes of adrenoceptor.
This causes many problems in drug development as its desirable to make drugs that are as selective as possible for our target.

It would be better in terms of finding new drugs if it were possible to test them against the target protein with no other related proteins present (isolating target protein).
If structural studies or drug screening are being carried out, we would also want to have large amounts of our target protein available.

One way around this problem is to isolate a tissue that has the target of interest.
However, finding such a drug can often be handicapped by the inability to get a source of the specific target protein, as the tissue often has a mixture of proteins that might interact with the drugs we are screening.

For example, there may be Alpha-2-adrenoceptors and Beta-1-adrenoceptors present in the same tissue, and the specific target protein might be present at very low levels.
The situation can be even more difficult if the specific protein is expressed in a very specific set of cells, such as in the brain. It may be very difficult to obtain these cells, especially if we want to work with human tissue.

• Expression Systems
The problem of the target protein diversity can be overcome by the use of Expression Systems.

The most current expression systems are cell or oocyte based.

The ideal expression system has:

An appropriate level of expression for purpose.
Has Fidelity of expression.
Is easy to use.
Is cheap.

The Fidelity of expression refers to the ability of the system to make the protein correctly, as you don't just need the amino acids to be in the right order, you also require the protein to be folded correctly, to be put into a membrane, and to get the correct post-translational modifications (eg: glycosylation).

Not all expression systems are able to do this, the basic rule is that the more closely related the organism from which the protein originates is to the organism from which the expression system originates, the better it will work.
So better fidelity will be achieved if you express a human protein in a human cell than if you tried to do it in an E. coli cell.

Protein Production

There are two main reasons why it may be desirable to express a protein.
Firstly, you may want to produce it in large quantities to do structural studies or use it as a drug (eg: Insulin).

If this is the goal, then using bacterial, yeast or insect cell based systems is likely to work best. However, you might need to do things to the protein after you have purified it, in order for it to fold correctly or get the proper post-translational modifications.

Through this very high level of expression can be achieved, which is good for protein production and purification (used in biopharmaceuticals and structural studies).
However, when this is performed, the post-translational modifications are not the same as in mammals and the proteins are often incorrectly folded.

Xenopus Oocytes

The second main reason to express a protein is that you might want to do experiments with them, or screen drugs.

In the past it was very common to use Xenopus oocytes for this purpose. These are unfertilized eggs from the Xenopus laevis (African clawed frog) and are amongst the largest of all eukaryotic cells.

Buy the full version of these notes or essay plans and more in our Drug Development (BIOL10822) Notes.