Team:Manchester-Graz/Practices/Experts
Expert Discussions
Interview with Prof. Dr. Joachim Reidl
Due to our special interest in microbial behavior of our system in the human guts, we talked to University Professor Joachim Reidl from the Karl Franzens University, Graz about our project. Professor Reidl is working at the Institute of Molecular Biosciences and his research field especially includes Medical Microbiology.
Regarding our project, he told us, that it will be very interesting to observe the differences in terms of regulation of your quorum sense based system in E. coli Nissle 1917, due to the big difference to ordinary lab strains like E. coli BL-21. He also mentioned that it would be interesting to find out to what extend the intestinal produced dopamine really goes into the blood stream and subsequently how much of it reaches the brain. A final comparison of how much dopamine is actually produced in vivo in comparison to a standard lab reactor experiment has to be drawn to.
Furthermore, this would be especially important to discriminate the exact amount of bacteria that has to be brought and cultivated in the human gut. According to web resources (1), medication such as “Levodopa” is usually administrated to patients in doses from 500 mg to 6000 mg a day. In previous work done on L-DOPA metabolic engineering, researchers developed E. coli strains that were able to produce up to 1.51 g/l of L-DOPA in 50 hours in batch reactors (2), meaning that we would have to improve these results significantly to actually develop a sufficient production strain that can be administrated directly to the human guts.
Another interesting aspect is the molecular mechanism of how bacteria would keep the information of Dopamine/L-DOPA synthesis. In the intestine, different to the fermenter where an antibiotic can be added, selection pressure would not be present. The latter would mean that the engineered bacteria eliminate their plasmids. To counteract this problem, integration into the bacterial chromosome could be approached or selection pressure with knocked out essential E. coli genes could be applied. Still, this is no guarantee for the integrated DNA or the plasmid to be kept intact by the bacteria because of possible cut out or mutation of the foreign DNA.
The main point we were able to take back to the lab from this discussion was that we will most probably not have to worry about our developed strain overgrowing the human gut composition. Actually there is a possibility that the strain may have difficulties reaching the required L-DOPA levels, this is a problem we will have to work on in the future.
2) Ana Joyce Muñoz · et al (2011) Metabolic engineering of Escherichia coli for improving L-3,4-dihydroxyphenylalanine (L-DOPA) synthesis from glucose
Interview with Prof. Dr. Christoph Högenauer
In order to find out more about human gut bacteria composition we met Prof. Dr. Christoph Högenauer from the Institute of Gastroenterology and Hepatology at the Medical University of Graz.
We found out that composition of gastrointestinal colonization is not only different from person to person but also depends on geographic location, genetic disposition, possible disease and current eating habits. Quantification of these bacteria strains is quite difficult as some might only be abundant in very low concentrations. For actual quantification, q-PCR is the method of choice. However, Escherichia coli makes up 0,1-1% off all gut bacteria but up to 50% during enteritis (1).
Our selected probiotic E. coli strain Nissle1917 is currently sold under the trademark Mutaflor ®. If applied to the patient only once it is observed that the strain does not stay in the gut permanently but has a lifetime of only a few days and is not detectable after a week. Regular administration is still required.
Interestingly during our interview, Dr Högenauer also told us that he was also working with Parkinson’s patients, preparing them for a therapy form called Duodopa ® from Abbott (2). This therapy especially serves patients that cannot take L-DOPA pills for varying reasons (intolerance or too high tolerance to oral L-DOPA). It is a special way for administration of L-DOPA (Levodopa ®) directly into the human jejunum through a PEG-PEJ tube. This approach allows for a continuous and stable dosage of L-DOPA over the whole day. However some major problems of this approach are high costs (around 700€ per week plus costs for placement of the tube and aftercare) and wear out of the tube every 2 years. Also the system needs to be rinsed on a daily basis and during the night medication has to be taken oral anyways. This ads up to 100.000 € per year per patient.
Obviously this is exactly where our project comes into play! Our idea allows for continuous administration of L-DOPA straight into the gastrointestinal trace which does not only get rid of tubes going into your body but also allows for a much smaller concentration of L-DOPA administration. Levodopa is currently applied through Duodopa® in concentrations of 20-200mg per hour, in contrast to previously mentioned pills in the multi gram range. Gastrointestinal production of L-DOPA would not only allow for a better lifestyle for patients but also discharge health insurance funds.
2) Wenzel, K. et al. (2014) P-aktuell: Aktuelles zur Pumpentherapie mit intrajejunalem Levodopa-Gel
