This section describes the design of the three main components conforming Hormonic, together with a description of our proof of concept for our system validation.
The heart of Hormonic lies in the ability to quantify the levels of T3 hormone in vivo. That's why our amazing team, conformed by multidisciplinary scientists, successfully came up with an innovative and completely new way to carry out these measurements: an intein mediated T3 biosensor engineered using synthetic biology. But Hormonic goes beyond that, it is designed to restablish the hormonal feedback system thanks to an external software coupled to our modified bacteria to achieve a groundbreaking therapy approach for hormonal disbalances.
The following block explains the three main components of Hormonic:
How can we measure the amount of T3 hormone in the patient?
Bearing in mind that the design of a robust T3 hormone biosensor is a key component of Hormonic, the first thing we need is a biological agent capable of translating the biological information into measurable values, that will be interpreted by the external controller previously explained.
For this we used E. coli bacteria, and to be able to sense T3 , we decided to engineer a protein called intein. Inteins are a naturally occurring type of protein-segment that, at a post-translational level, are able to remove themselves from the peptide-chain, joining the two remaining portions at both sides.
Since the main characteristic of inteins is their ability to reconstruct virtually any split protein, we thought that, if split correctly, we could use a Fluorescent Protein. So, what we did was to split different GFPs, and couple them to the intein.
Using light as a reporter we can quantitatively and continuously know the amount of hormone. This way, when T3 is present in the media, fluorescent proteins are formed and the light signal can be detected by our computational unit.
How can we regulate the right dose of levothyroxine to supply?
The majority of control systems use PID controllers: regulation mechanisms that aim to keep a system in a steady state by the use of feedback loops. To do so, they continuously compute the difference between the desired hormonal level and the measured one and calculate the proper action to compensate for the possible deviations (see Figure 3). In our case, this action would be the injection of levothyroxine.
Moreover, the levothyroxine’s half-life is quite large and its metabolization into T3 slow, generating a delay on the response that should be borne in mind when defining the periodicity of the measurements and the subsequent injections. If this was not considered, it could derive into the uncontrolled injection of diluted levothyroxine, which would be risky for our patient. Therefore, this was all included in our PID software.
How the drug will be subministrated?
Hormonic is able to sense the amount of T3 and calculate a compensation for the system, so it also needs a way to respond. That’s what we call the effector. This way, we are closing the loop by subministratrating small quantities of a drug that will gradually bring the hormone levels to the desired concentration.
To treat hypothyroidism, the drug that’s used it's levothyroxine, which increases the T3 hormone levels. Its administration would be done by the release of several microdoses into the system until the desired levels are reached. This will be possible thanks to small pumps that the PID controller will activate depending on the amount of drug needed at every moment.
The future of Hormonic goes beyond this, the biosensor, PID and effector would be merged into a compact device. Our design is based on a skin patch with an array of microneedles, which will mediate both: the liquid sampling and the release of Levothyroxine. Moreover, the device would be connected to a computer, integrating all the data collected from the biosensor and establishing a connection with the clinician to remotely know the hormone concentration profiles.
Lactone Proof of concept
How can we prove that Hormonic would correctly restablish the feedback loop?
Knowing the 3 components that create Hormonic, it is necessary to know if the whole regulation mechanism works. The endocrine system is very complex: the triple feedback that rules the hypothalamic-pituitary-thyroid axis makes the thyroid hormonal levels not constant, thus possibly producing unknown temporal cycles. The fact that we could not know the effect of levothyroxine in our system (the release drug would not be metabolized into T3 hormone, because we are not working with human cells) and the uncertainties related to thyroid behaviour made it impossible to close the loop.
Consequently, we had to design a closed mockup system to test the ability of our PID to maintain the stability of the feedback. A minimal hormonal regulation model was created with lactone as the target hormone of the system. Different cellular types have been used to create a whole genetic circuit with Lactone Producer Cells (acting as the Thyroid gland), Lactone Sensing Cells (acting as our Biosensor) and two different effector molecules, that will act upon the Lactone Production to increase or decrease it. In this way, we are able to test and prove the ability of our PID to regulate hormonal levels, while guaranteeing the health and security of the patients.
But the most interesting thing of being a general circuit is the fact that if Hormonic can regulate an hormone cycle, it can be extrapolated and used to regulate any other hormonal feedback.