Thinker-ShenZhen  ChDeHome

Before the project began, several members of our team had a casual conversation about the common fungal skin disease and the fact that it was a problem for people around us. We had also heard lots of complaints about those disease and the medication. We then distributed a questionnaire to investigate the public's feelings on fungal skin diseases and found that the disease afflicted many people.

However, common chemical drugs had side effects and tended to lose their effectiveness. With this in mind, we decided to design and produce our own innovative medicine to treat fungal skin diseases and address the pain points of the general public. After our efforts, we finally contacted a teacher at Lanzhou University and had an initial exchange of ideas, identifying the titin in the fungal cell wall as the target and using chitinase to attack and degrade it from the outside. Later in an interview with the dermatologist, we presented our first experimental data to her and received feedback that the degradation efficiency was hardly efficient in inhibiting the fungus. We then read a lot of literature and contacted the dermatologist again to redesign the product by adding a tributylin-binding structural domain at the nitrogen and carbon ends of the tributylase respectively.

In a second interview with a medical professional, she concluded that the data from the second experiment were sufficient for efficient fungal inhibition and treatment of fungal skin diseases. We sent out a questionnaire to the general public about how to use the product and found that people were resistant and averse to E. coli and we were aware that direct application of E. coli to the skin could easily cause genetic leakage and pose biosafety issues, so we took advice and decided to use our chitinase as a pure enzyme product. In the follow-up questionnaire, we received feedback that the general public accepted the use of pure chitinase drugs.

The next question we considered was how to extract the E.coli BL21 chitinase in the plant. Our team read a number of papers and production reports on the subject and found that an efficient lysis system could be used to disrupt the cell wall of E.coli BL21 to release the intracellular chitinase. The initial idea was established and we encountered difficulties in implementation.

Fortunately, we discovered in a later project exchange meeting with Thinker-China that they had designed a successful lysis system of their own, and with their help we were able to get our own cracking system. We took our cracking system to our teachers and they thought it was feasible, so we started validation experiments on the cracking system and successfully proved that it was feasible. We also interviewed the people in charge of the drug companies to find out about drug design, packaging, price and safety. Similarly, we again distributed questionnaires to collect consumers' thoughts on what they expected our drugs to cost, design style and so on.

All in all, Human Practices was used throughout our project to help us gather feedback, iterate and improve.

Before the project started, a few of us talked about how many people around us have fungal skin diseases, such as foot fungus, ringworm and candidiasis, and how they cover a wide range of ages, from elderly grandparents to middle-aged parents or relatives to young people around them.

With the return of two hundred valid questionnaires, we found that more than half of the respondents reported that they or someone close to them had struggled or were currently struggling with fungal skin disease. The sheer number of people suffering from fungal skin disease makes us realise that this problem cannot be ignored

More than half of the respondents said that the drugs they were using had declined in effectiveness to varying degrees, to the extent that they needed to increase their dose or even choose a new drug. It is therefore important to develop a new medicine.

We have reviewed a number of chemicals used in the treatment of fungal dermatoses and found that some lyse the fungus by inhibiting the synthesis of ergocalciferol, such as triazoles, while others interfere with cell wall synthesis by inhibiting the synthesis of chitin, such as Nikkomycin./p>

Our team had learned about Chitin and the fungal cell wall and thought we could take the idea of Nikkomycin and target Chitin, but in a different way, instead of inhibiting synthesis from the inside, we could attack and degrade Chitin from the outside.

We purchased a commercial chitinase and put it into experiments to measure its ability to bind chitin, degrade it and inhibit the fungus, and obtained first experimental data.

We interviewed a doctor specialising in dermatology and presented our data to her. After looking at it, she concluded that the current degradation capacity of chitinase was not sufficient to inhibit fungal growth well enough. It needs further enhancement.

After receiving this evaluation, our team was confused and don't know where to start to improve the degradation capacity of the enzyme. Did the enzyme concentration need to be increased? Or were the optimum conditions not met?

We re-examined the fungal cell wall structure and had a second conversation with the faculty at Lanzhou University to analyse how we should improve the degradation of the tributylase enzyme. In the end, we found that the low degradation efficiency was due to the fact that fungal titin is located deep in the cell wall, which is generally difficult for titin to bind and degrade directly from the outside, so we planned to bind fungal titin by adding a titin-binding structural domain to the nitrogen and carbon ends of the titinase respectively to better inhibit the fungus.

We conducted a second interview with the same doctor after the second experiment. We have tested the binding capacity, degradation capacity and bacterial inhibition capacity of the modified chitinase. After presenting the data from our second experiment, she concluded that the enhanced degradation capacity was sufficient to meet the criteria for bacterial inhibition, and she also made a number of valuable suggestions.

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Fungal dermatoses are usually treated with antifungal medication, and there are many drugs that can be derived from different targets. However, most of them do not address the issue of fungal resistance, and the effect of the drugs will diminish to varying degrees during use. The usual option chosen by doctors for this problem is to follow up with a drug sensitivity test. Most dermatological treatments are longer than a six-week course of treatment. Topical and oral medications are used in the treatment of fungal dermatoses, for example, oral medications are used to prevent deep fungal infections that can affect the liver and kidney function. Most of the medications are inexpensive, costing no more than $50. One of the most common topical medications is Dakine, although due to abuse, fungal resistance can easily develop resulting in ineffectiveness. When it comes to the standard procedure for treating skin diseases, it is usually a case of going to the hospital for an allergy investigation and scraping some samples from the affected area for a fungal test to determine the cause. We have been unsure whether the incidence of skin disease is related to age and gender, to which we have been given the answer that genes determine fungal susceptibility and also the external environment, which can cause relapses. On the question of self-testing for pathogens at home, the doctor replied that a prick test is now available for patients to self-test at home, although the results are not accurate due to individual variability and it is recommended to go to hospital for a more accurate diagnosis. If you must test at home, the test reagents can be used repeatedly at a cost of nearly $100 per test. If left untreated, a common fungal skin disease can become a deep infection, which can worsen and endanger health.

In relation to our project to increase the antifungal capacity of chitinase to treat fungal dermatoses, the doctors made several suggestions.

1. translational medicine already has relevant pathways for fungal agent customisation, e.g. Nicomycin-based drugs can inhibit titin synthesis to reduce fungal presence

2. the cost and conservation of biologic enzymes is relatively high

3. The cost and yield of Ecoli-generated chitinase is uncertain and it is difficult to ensure homogeneity of the active substance in the cream

4. The direct use of E. coli could easily lead to gene leakage and affect biosafety

(For the above suggestions, our team will then meet to discuss improvements.)

Once again, we have sent out an online questionnaire to the public to find out what people think about our products

On the question of whether they would accept the use of E.coli BL21 directly on human skin, 89.86% of respondents said no. They felt that E.coli is, after all, a bacterium extracted from the large intestine and that there would be a psychological aversion and resistance to the E.coli drug. The doctors also felt that if BL21 was used as the main ingredient, we would not be able to guarantee the same production of titinase for each bacterium, which could lead to mediocre results. In addition, the doctor also warned us that the direct use of BL21 could easily cause gene leakage during fragmentation, posing a biosafety issue. Taking these factors into account, we have made a further improvement: we grow large quantities of BL21 directly in the plant and break the cell wall at a specific time to release large quantities of intracellular chitinase.

We reviewed a large amount of literature on how to break the E.coli cell wall and found that an efficient cracking system could be designed to lyse the cell wall. After talking to teachers from SMS, our idea was to use an arabinose cracking system, but we encountered difficulties in the specific design and operation.

Later, during a project exchange with Think-China, we discovered that they had a proven cracking system of their own, so we asked them to help us design a cracking system together.

After showing the cracking system to our teacher, he thought it was theoretically feasible and we carried out verification experiments, which proved that our cracking system worked properly.

Pharmarcy Enterprise

Our team interviewed Si Shufeng - the director of the CKJY Institute of Microecology and Ecology - about some drug production, cycle times, raw materials and safety, in the hope that we can learn how to plan our business and package our goods.

After the interview, our team members who are responsible for business planning can do a better job of merchandise packaging, audience targeting and promotion.

Consumers:

We interviewed people with fungal skin diseases and pet owners to ask about skin diseases.

Interviews with people with skin diseases

The number of people suffering from skin diseases in China is extremely large, with a wide range of causes, common types and treatment options. Our project also had to incorporate patient ideas, meet needs and overcome pain points. In order to gain first-hand knowledge of patients' dermatological conditions to improve our project, we found several typical dermatological patients and conducted valuable interviews over the summer.

We interviewed mainly two lady patients at the hospital's dermatology department as follows.

Pet owner interview

Skin diseases can affect not only humans, but also pets, such as dogs and cats, who also need to be treated for skin diseases, both because their health is affected and because pets can transmit skin diseases to their owners. In order to make our titinase programme more universally available to treat both humans and pets, our team conducted online interviews with a number of pet owners, as follows.

Nearly three quarters of the pet owners we interviewed had dogs and the remaining quarter of the pets were cats. The common skin conditions were some form of ringworm, which was generally treated by taking them for a medicated bath and spraying them with antifungal medication; the medicated bath was between $100-200, twice as expensive compared to a typical bath; the antifungal medication was also mostly nearly $100, with one representative being Pitocin (50mL, $90). If left untreated, the pet's localised itchy skin will then scratch unconsciously, producing a dandruff-like substance to the point where the owner will feel uncomfortable during stroking; the pet will also lose hair and become ugly in appearance, causing the owner to feel psychologically distant. When it comes to the final advice session, the main pain points are the high price of treatment - medicated baths are twice as expensive, and the small amount of antifungal medication that usually needs to be used consistently is expensive, which is a burden for pet owners who are not financially well off; and the inability to solve the problem themselves - skin diseases in general We wish we could tell at home whether the skin disease is fungal or not, but it is too much trouble and we don't know much about it to check it out at home, so we have to spend time and effort to go to the hospital for biopsy.

In response to the above, we are acutely aware that it is a very important and urgent matter to be able to quickly detect the cause of skin diseases at home, saving a lot of time and shortening the treatment time, so that discomfort can be investigated and treated quickly at home, eliminating the trouble caused by some minor diseases.

After two in-depth interviews we discovered that our medication can treat fungal skin diseases not only in humans but also in pets. As fungal skin diseases can affect a wide range of people, and children of a younger age are also at risk of infection, we decided to design different products for different groups. For children, our packaging would be cute and not easy to use, in order to prevent misuse by children; for adults, our packaging would be simple and easy to carry and use without delaying the commute. And in the follow-up questionnaire, we found that people preferred to use sprays and gels. Taking into account storage issues and the state of the presence of the enzyme titinase, we decided to use gels as the main form of the medicine, which is easy to store and use.

All in all, Human Practices has been used throughout our project, and we have been able to reap the benefits of the world helping us, as well as improving on the suggestions, iterating and improving.

The world affects our work, we affect the world.