Team:UCAS-China/Parkinson Disease

Parkinson's Disease

Overview

To prove the potential of microbial therapeutics and our platform’s universality and unique advantage on microbial therapeutics, we chose Parkinson's disease as an example of long-term medication diseases, which needs frequent and precise doses of L-dopa.

We firstly transferred the tyrosine hydroxylase(TyrOH) gene, which can catalyze tyrosine to L-dopa, into our engineered E.coli to provide a stable and sustainable concentration of DOPA in the patient’s blood. Compared with the traditional therapy by taking pills regularly for a long time, microbial therapeutics can achieve a constant and stable supply of DOPA.

Finally, combining biocontainment module of our platform with the circuit for releasing TyrOH can make this microbial therapeutic for Parkinson's disease safer and meet the requirements of clinical and commercial applications.

Background

Parkinson's disease is a common progressive bradykinetic disorder that can be accurately diagnosed. It is characterized by the presence of severe pars-compacta nigral-cell loss, and accumulation of aggregated α-synuclein in the specific brain stem, spinal cord, and cortical regions. The main known risk factor is age.

As this is a chronic disorder with a prolonged course, prevalence is much higher than incidence. Crude prevalence estimates vary from 18 per 100 000 persons in a population survey in Shanghai, China, to 328 per 100 000 in a door-to-door survey of the Parsi community in Bombay, India[1].

According to Figure 1 [2](Modified), whose data comes from Dorsey, E. R. et al., about half regions of the world keep the growth rate of more than 50% in the number of individuals with Parkinson's disease, which indicates that Parkinson's disease actually has severe threat worldwide.

Figure 1. Projected growth rates in the number of individuals over 50 with Parkinson's disease in the most populous nations in Western Europe and the world from 2005 to 2030

At the early and intermediate stage of Parkinson's disease, patients mainly depend on drug, levodopa, to delay the progression of motor complications. [1]

However, frequent and precise doses are needed for the sake of the harmful side-effects of levodopa with a high concentration in plasma, and the short-term spikes in the bloodstream caused by the drug.

At the same time, with the aggravation of the disease, the quality of life of patients with Parkinson's disease gradually declines.

What's worse, the degeneration of the nervous system in the brain makes them lose control of the body gradually, accompanied by sluggish movement and uncontrollable limb twitch.

Moreover, apart from the normal patients, or called "stable" patients, there is a group of patients, which is called "fluctuate" patients, showing functionally important “wearing-off" phenomena [3]. With the same doses intake, the length of clinical effect for "fluctuate" patients is much shorter than "stable" patients and their motor response (evaluated by tapping score) usually drops back to baseline in 2-3 hours. These patients generally have a longer disease duration and a worse clinical-stage, and have to be treated the levodopa with on average twice the frequency (on average 5 doses per day) while twice the size of every dose (on average 550mg per dose).

From all these facts listed above, we can make the conclusion that it is quite difficult for patients, especially for those "fluctuate" patients, to take medicine frequently for a long time. Therefore, we shall seek a new way of drug delivery which can provide a suitable and stable concentration of levodopa in plasma, to achieve a stable clinical effect while reducing the unwanted side-effects.

Microbial Therapeutics

Microbial therapeutics has unique advantages on this severe problem by providing a sustained, stable and trace supply of levodopa.

Figure 2. TyrOH(or TyrH, tyrosine hydroxylase) catalyzes tyrosin to levodopa

Figure 3. Gene of TyrH under a constitutive promotera

Firstly, we transferred the tyrosine hydroxylase(TyrH or TyrOH) gene into E.coli, so that the engineered bacteria can synthesize dopa in the presence of tyrosine. After that, dopa will enter the brain area through the blood to maintain a normal dopamine concentration.

To characterize the efficiency of TyrH, we conducted the fermentation experiment of levodopa, and the result, as shown in Figure 4, shows that the maximum concentration of dopa can reach more than 130 mg/L after 8 to 12 hours of fermentation. By the way, the subsequent decline maybe due to oxidation of dopa during fermentation, but this hardly occurs in the gut.

Figure 4. The fermentation curve of levodopa

The comparison of taking drugs by pill and microbial therapeutics

We employed a model to explain the advantage in dose controlling the drug.

We abstract the body into three compartments, absorption compartment –the intestinal tract, central compartment –blood circulation, and effect compartment, which is related to clinical effect. The rate of drug delivery is in direct proportion of the drug amount.

Then we consider two ways of drug delivery—by pills and by bacteria. Their difference origins in the absorption compartment. If by pills, some drugs are released initially, and then no drug is produced. If by bacteria, as we built a model to simulate the distribution of bacteria in the intestinal tract and found the distribution tends to be even, thus the drugs are produced and released continuously.

Then we made a comparison between bacterial delivery and oral delivery, by comparing the clinical effect versus time, and by comparing the drug concentration in plasma, which is related to possible side effects.

Figure 5. Drug Concentration Versus Time by Different Methods, for Fluctuate Patients

Figure 6. Clinical Effect Versus Time by Different Methods, for Fluctuate Patients

(The data of pills comes from our study of current drug use [3][4], while the data of microbial therapeutics comes from our fermentation experiment. Here we only showed the result of the fluctuate patients).

As shown in Figures 5 & 6, obviously, compared to oral delivery, the bacterial delivery generally has a high-and-stable clinical effect, while having a much lower drug concentration in plasma, thus will bring much fewer side effects. So the bacterial delivery can perfectly reduce the side effects while maintaining a good clinical effect, and with such method, patients no longer need to take pills frequently.

See more details about this model here.

The combination of microbial therapeutics and our platform

To make this microbial therapeutic for parkinson’s disease more safe and meet the requirements of clinical and commercial application, as shown in Figure 7, we combined the biocontainment module of our platform with the circuit for releasing TyrH to product the drug, levodopa.

In our biocontainment module, we use low-temperature inducible switch to control the expression of toxic proteins to prevent the engineered bacterial from escaping into the environment.

Figure 7. The combination of biocontainment module and therapeutic module

References

[1] WHO (World Health Organization) (2006). Neurological Disorders: Public Health Challenges.

[2] Dorsey, E. R., Constantinescu, R., Thompson, J. P., Biglan, K. M., Holloway, R. G., Kieburtz, K., . . . Tanner, C. M. (2007). Projected number of people with Parkinson disease in the most populous nations, 2005 through 2030. Neurology, 68(5), 384-386.

[3] Contin, M., Riva, R., Martinelli, P., Cortelli, P., Albani, F., & Baruzzi, A. (1993). Pharmacodynamic modeling of oral levodopa: clinical application in Parkinson's disease. Neurology, 43(2), 367-371.

[4] Contin, M., Riva, R., Martinelli, P., Albani, F., Avoni, P., & Baruzzi, A. (2001). Levodopa therapy monitoring in patients with Parkinson's disease: a kinetic-dynamic approach. Ther Drug Monit, 23(6), 621-629.