Making medicines in yeast
A significant proportion of the drugs in the average doctor’s bag have their roots in nature. Digoxin, artemisin, morphine and salicylates are all good examples. Another is forskolin. This is an incredibly useful therapeutic chemical but it’s also an almost impossibly difficult molecule to make, so doctors have been forced to rely, so far, on a natural source, limiting supply and driving up costs. Now Irini Pateraki has found a way to make yeast brew it up for her, as Chris Smith found out...
Irini - Forskolin is a compound that is produced only from one plant. The plant is called Coleus forskohlii, mainly growing in India, and south Asia. Forskolin is important because it has a lot of pharmaceutical properties that all of them depend on the ability that forskolin has to help in relaxing the vessels of our body. For example, forskolin helps treating hypertension, asthma, glaucoma in the eyes. Actually, it’s one of the very few drugs that are efficient in treating glaucoma. Today, we have in the market medicines that contain forskolin for that purposes.
Chris - But because it comes from one single plant source, we’re therefore dependent on that plant to extract the precursors and the chemical.
Irini - Exactly. Also, we have to have in mind, this plant produces forskolin in relatively low amounts. So, when we need forskolin to give it to the market, this source is not enough. And also, forskolin in that plant is produced in combination with many different metabolites. So this plant is producing something like 70 different metabolites, but quite similar with forskolin. So it’s very challenging to isolate and purify forskolin from this plant.
Chris - Why can't we just make forskolin artificially just in the test tube?
Irini - Because Forskolin is a quite complicated molecule so it’s very challenging chemically to produce it. Also, it will be very expensive. The amounts that you can achieve are very low... so the price and the efficiency is not really favourable.
Chris - On the other hand, if you can work out how the plant is doing it and then steal the genetic recipe then you could recreate the synthetic pathway in another organism and mass produce it.
Irini - This is exactly what we are doing. We went back to the plant, sequenced the transcriptome which means we sequence genes that they were expressed in the plant, and we found out the genes and enzymes responsible for the production of forskolin.
Chris - Did you focus on the part of the plant which makes the highest concentrations of forskolin so that you knew you had a reasonable chance of the genes which are switched on there are the ones which are directly linked to making those chemicals?
Irini - Exactly. This is what exactly we did. So we knew that forskolin generally is produced in the roots and we knew that from the tradition because I mean as I said, forskolin is used for many years. So people knew that they can find it in the roots. But when we started to work on that and we went closer, we found out that the forskolin is not generally produced in the whole root but is produced very specifically in the cork of the root. And then we found out that these cork cells, they contain some oil bodies where these oil bodies, they are able to store forskolin and also forskolin precursors. From this data, we realised that forskolin is also produced in these cells, not only accumulating but also produced. So we went back and we sequenced this very, very specific tissue, and this was a big help for the identification of these enzymes.
Chris - How many enzymes are involved in the whole pathway?
Irini - We found nine different enzymes that were necessary to produce this compound. The difficult part though is to make all these genes to express together and to work together in a different organism to produce what you want them to produce because the environment of a plant is very different than an environment of a yeast cell. A plant cell is much more complicated than a yeast cell so you have to make some adaptations and you have to be sure that all these enzymes that you move from the plant to the yeast cell, you can make them functional.
Chris - Does the yeast tolerate making forskolin okay or is it toxic?
Irini - You could say that it’s toxic but because it’s toxic, here, they have some nice machinery that excrete forskolin out of their cell to the medium.
Chris - How much forskolin does it make?
Irini - In this paper, we have said that it can produce up to 40 mg of forskolin per litre of yeast culture. Actually today, we have managed to get much higher titers.
Chris - How does that compare then in terms of the amount of plant matter you would need to extract the same quantity of the drug? Is this a viable and scalable way of producing forskolin?
Irini - This is totally scalable because of course, you can use as many yeast culture as you like. So you can have several bioreactors while for the plant, the plant needs a whole year to grow and then you have to chop it off to take the roots. When you rely on plants to produce forskolin, you have to depend on yearly growth.