Team:CPU CHINA/HP/Gold Integrated

Intergrated and Gold HP

Summary


Our team aims to fully exploit our university’s advantage in pharmaceutical science to design an effective treatment of a disease. By comparing the data collected, we chose rheumatoid arthritis and other refractory autoimmune diseases for our project. Conducting thorough field studies and understanding the current clinical status of rheumatoid arthritis were very important to our design and optimization.
Through field studies, we knew that Syn-Notch CAR-Treg cell therapy satisfies the patient's desire for new therapies, and through a questionnaire survey of physicians, we concluded that Syn-Notch CAR -Treg cell therapy just hold the advantages of the minor side effects and lower recurrence rate compared to existing drug treatment. In designing the therapeutic method, we were also fully dependent on the patient's request for new therapies to meet current clinical needs.
We actively design, perfect and verify our own project. In the meantime, it is an opportunity for every single person to understand the competition, together with a new frontier in life science. What’s more, we communicate with other iGEM teams and help each other.In addition, all of the members actively organized various academic activities, and actively participated in disparate meetings inside and outside school to promote the majority of teachers and students to comprehend the field of life science, as well as to improve the international influence of iGEM. At the same time, it also made our team members accumulate cutting-edge knowledge, and provided a strong guarantee for the improvement of the subject

Survey data


Rheumatoid arthritis (RA) is a chronic inflammatory disease that may exhibit chronic synovitis, synovial hyperplasia and fibrosis, mononuclear cell infiltration, synovial fluid T, B lymphocytes and plasma cells, increased collagenase, etc. It’s a chronic disease with high morbidity. There are several new therapies that have been used for the treatment of RA, but due to low awareness and low recovery rate, many patients fail to receive a proper treatment in RA’s preliminary stage which not only afflicts the patients, but also brings a heavy financial burden to their families.
In China, the prevalence of rheumatoid arthritis is 0.2% -0.4%, so with a population of 1.3 billion, China's rheumatoid arthritis patients are expected to be around 5 million. If they cannot be given effective treatment, their mobility will be limited due to joint pain, and will eventually lead to disability. Statistically, disability rate in patients with two years of rheumatoid arthritis is up to 50% while those with three years can be 70%.

Field Studies


To further understand a patient's conditions and the clinical status of treatment, we decided to go to Nanjing Gulou Hospital and Nanjing First Hospital to perform a questionnaire survey in the department of rheumatology immunology after beforehand investigation and contact. First is our preparation:
Respondents: patients with rheumatoid arthritis at inpatient and outpatient in rheumatology department
Sample Size: 80 people
Procedure:
1. The production of questionnaire (Avoid sensitive issues and patients’ personal information, and respect patients for their personal privacy).
2. Assign teammates with different tasks.
3. Determine the procedure and the time.
With our planned scheduled, we went to the hospital for a two-day survey (Figure 1). We ensured the authenticity of the answers to our questionnaires. For patients with reduced mobility, our members completed the questionnaire according to their oral narration (Figure 2). We tried to gain more insights in the treatment they expected to receive, as well as the treatment they received in real life, which enabled us to feel more about the pain and suffering this disease could bring.

Figure 1. Conducting a questionnaire in the Nanjing First Hospital

Figure 2. Our teamer was talking with the patient carefully

Questionnaire analysis


The following conclusions can be drawn by analyzing the data collected from the questionnaire statistically (Figure 3-5 are part of the questionnaire data) together with some supporting materials:
Rheumatoid arthritis is a disease most commonly seen in women aged 40-60 years old, so it is a typical middle-aged and old-aged disease, which seriously affects patients' life quality. Most patients choose drug treatments to alleviate the pain, but in actual clinical practice all currently available drugs fail to show any very significant therapeutic effect, which is why patients crave for new treatment methods. They also hope that the new method can have less side effects and lower recurrence rate, thereby reducing their pain. In addition, rheumatoid arthritis can also increase the burden on the national economy. The United States has once carried out an epidemiological survey, showing that annual cost on rheumatoid arthritis and rheumatoid-related treatment is more than 1% of its GDP. Though there is no such exact data in China, it is estimated that the financial loss caused by this disease is not less than the United States. As the aging problem becomes increasingly prominent, this national economic burden will become more and more heavy. If a new treatment can be designed to cure this disease radically, it will be of great significance in promoting public health and people’s health conditions.

Figure 3. The treatment effect and gender of patients

Figure 4. The impact of patients’ life and work after the illness

Figure 5. The existing treatment methods

Figure 6. Chimeric Antigen Receptor T-Cell Immunotherapy

Finding new treatments


CAR-T, short for Chimeric Antigen Receptor T-Cell Immunotherapy, is a chimeric antigen receptor T cell immunotherapy. This is a new cytotherapy that has existed for many years but hasn’t been used clinically until some recent improvement. Similar to other immunotherapies, its basic principle is to use the patient's own immune cells to specifically remove cancer cells. As shown in Figure 6, T cells are differentiated and enriched by in vivo injection of cytokines, and then the target cells are separated by cell surface markers. Next, the cells are engineered in vitro with CAR (Chimeric Antigen Receptor), and are amplified by cell culture to a certain number before returned to the patient's body. In this way, the engineered T cells can kill the target cells specifically in vivo.

Syn-Notch CAR-Treg cytotherapy


We discussed how to design a treatment for rheumatoid arthritis during our group meeting (Figure 7) and with our advisor in depth. Eventually, based on traditional CAR-T, we chose Treg cells as engineering targets in the purpose of curing rheumatoid arthritis, and we innovatively introduced a Syn-Notch system to enhance the stability of the engineered Treg cells in vivo. We came to form a new Syn-Notch CAR-Treg system (the overall idea shown in Figure 8) after some optimization on our design.

Figure 7. The daily group meeting

Figure 8. The global design of Syn-Notch CAR-Treg system

Retrospection of Survey


With a deeper research on treatment methods, the feasibility was verified at experimental level. But it still had a long way to go before it can be applied clinically. With our first survey results and some questions, we went to the hospital again, and conducted another questionnaire survey and interviewed some residents (Figure 9).
Preparation work:
Respondents: patients and doctors with rheumatoid arthritis at inpatient and outpatient of rheumatology department
Sample size: 30 doctors and 80 patients
Procedure:
1. List a questionnaire for patients and doctors (Avoid patients’ and doctors’ personal information).
2. Assign teammates with different tasks.
3. Contact relevant departments of the hospital.
4. Determine the process and the time.

Figure 9. Go to the hospital for in-depth investigation again


According to our plan, we came to Nanjing First Hospital and Nanjing Gulou Hospital for a questionnaire survey. The majority of patients showed great interests in our innovative therapy and explored the specific way of using this therapy after a conversation with them. Some patients were confident in our treatment and encouraged us to make more efforts in getting this therapy into clinical as soon as possible (Video 1). According to the survey, most of them have suffered from RA for more than 5 years. It was gratifying to note that more than half of the patients said they were willing to try Syn-Notch CAR-Treg cell therapy while others showed some uncertainty (Figure 10). But we believe this therapy will be more recognized with more future research.

Figure 10. The personal willingness of patients to try syn-Notch CAR-T cell therapy

Video 1. We have a deep conversation with patient


As for our survey with the doctors, most of the them have worked for more than 5 years (Figure 11), and they believed that the current drug treatment had shortcomings like severe side effects, a high recurrence rate and a chronic medication use (Figure 12), while Syn- Notch CAR Treg cell therapy had a more specific target and may show less side effects (Figure 13), which surpassed the existing treatment. Its safety was also further recognized by healthcare professionals. However, there are many problems remaining to be solved before it can actually applied clinically(Figure14), such as a limitation on large-scale production, quality control standards, etc.

Figure 11. Doctor working years

Figure 12. The shortcomings of existing treatment(Multiple choice)

Figure 13. The advantages of CAR Treg cell therapy (multiple choice)

Figure 14. The CAR Treg cell therapy will face the challenges for applying to clinic (multiple choice)

Depth Interviews


In order to further validate the safety and clinical recognition of our program, we conducted a thorough interview with several physicians and discussed the reliability and effectiveness of Syn-Notch CAR-Treg cell therapy with dialectical thinking.
In June 2017, we interviewed Ai Zhang, a senior physician who specialized in rheumatoid arthritis at the department of rheumatology at the first affiliated hospital of Zhengzhou university, who was currently studying at Gulou Hospital in Nanjing (Figure 15). She was well aware of CAR-T and made a lot of suggestions to us.She pointed out the problems in actual operation which we needed to pay attention to and all of these are very instructive, such as Treg in the separation of the choice of flow sorting or magnetic bead sorting problems, Treg training problems, etc. We also had to further develop analysis and optimization of specific experimental programs. Plus, she suggested that we would focus on foreign pharmaceutical giants in CAR-T and Treg and other clinical trials of clinical therapy, and learned from some of the lessons learned.

Figure 15. We were interviewing Ai Zhang


Fortunately, we also interviewed Minning Shen, who is the associate chief physician of Nanjing first hospital in , Department of Rheumatology (Video 2). She was very concerned about this area and she gave a systematic evaluation within her point of view of the existing treatment and Syn-Notch CAR-Treg cell therapy. Her views were consistent with the conclusions we had made in the survey, and again verified the security and reliability of our program from an authority expert. She also made a general prediction of the clinical application of Syn-Notch CAR-Treg cell therapy. From the pre-basic research to the real application, Novartis's CAR-T pathways were significant for our application. Optimistically, Syn-Notch CAR-Treg cell therapy in the treatment of rheumatoid arthritis and other diseases will have a good effect.
Through the interview with two physicians, we received a lot of suggestions and opinions, and more of which was the recognition of Syn-Notch CAR-Treg cell therapy. And they were also considered Syn -Notch CAR-Treg cell therapy was highly viable.

Video 2. We communicate with doctor

Education and public engagement

CPU BioX Club Academic Association


In the joint efforts of all CPU_CHINA iGEMers, we established the first life science academic association in China Pharmaceutical University---- CPU BioX Club (Figure 16). The daily activities of CPU BioX Club include:

Figure 16. The speech meeting for establishing the club


1. Journal Club
It is held mainly by students in CPU BioX Club. We invite a reporter to make the presentation of the latest discoveries every week. The reporter will introduce the new discovery of medicine, synthetic biology and so forth. Our team members have absorbed a lot of knowledge, which widens our horizons and provides us with novel ideas (Figure 17).

Figure 17. The journal club


2. Academic debate contest
We encourage all students to participate in the debate contest. In the debate contest, we argued about some scientific problems such as whether tumorigenesis was determined by congenital genes. After a series of debate contests, all of team members have developed a critical thinking and judged scientific problems more dialectic (Figure 18).

Figure 18. Students in the debate contest


3. Thematic academic reports
According to the content which students were interested in, we invited professors of the relevant research areas to come to have the academic reports on their research areas to explain for the students’ questions (figure 19).

Figure 19. The teacher having the academic reports on their research areas

Teaching in high school


Invited by the biology teacher of the senior secondary school affiliated to the Bohai University in Jinzhou, Liaoning Province (Figure 20), our team members went to the high school for the students who were interested in biology to introduce the progress of synthetic biology and immunotherapy research. It actively promoted iGEM, and guided more people to join us in the development of synthetic biology to contribute their own strength.

Figure 20. One of our team members who is giving a lesson

Communicating with other teams


We held academic exchange activities with the other two iGEM teams, NJU-CHINA and NAU-CHINA.

1. Asking the NJU_CHINA team for experience exchange


At the Life Science Building Conference room in Nanjing University, the two leaders talked about the progress on their own subjects and the status quo of the team. Yu Zhou, the leader of Nanjing University team, told us about their recent sessions of the iGEM, and explained some important time node, like human practice attention points, the contents of the two co-operation and other issues. Also, he showed our team members around their labs. After talking with them, we have gained some knowledge of iGEM competition, which gave us a lot of help in participating in iGEM competition. (figure21).

Figure 21. Our team is listening to the introduction of NJU iGEMer

2. Inviting NJU-CHINA to attend academic debate


In order to enhance the exchange between two teams, we invited the iGEM team of Nanjing University to attend the academic debate (Figure22). Two teams conducted a debate on whether changes in epigenetic modification can determine tumorigenesis at the China Pharmaceutical University Conference Center:


Proposition: CPU_CHINA


Attitude: changes in epigenetic modifications can determine tumorigenesis


Counter: NJU-CHINA


Attitude: changes in epigenetic modifications cannot determine tumorigenesis


The two sides held different arguments, and cited frontier knowledge of self-argument and deduction to express their views. Also, two teams obtained a lot of gains during the debate, and made a further exchange on the issue of respective topics.

Figure 22. Inviting the iGEM team of Nanjing University to attend the academic debate

3. Participating in experience sharing meeting at Nanjing Agricultural University


Invited by the iGEM team of Nanjing Agricultural University, we participated in experience exchanging meeting (Figure 23). The meeting invited Haoqian Zhang, the predecessor of Peking University iGEM team, to share his experience. According to his previous experience, all the notes on the formation of the team and participation in the Giant Jamboree have been described in details and shared with us about the personal experience of participating in the iGEM competition. Besides, we actively communicated with other teams to share different ideas and projects processing. It inspired us a lot and we avoided a lot of problems that may happen in our experiment.

Figure 23. Participating in experience sharing meeting at NAU

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