Human Practices

A Way out of the Ivory Tower

We, young guys buried in piles of schoolwork, have been in the ivory tower for so long, with everyday tasks in front of the laptop and after-class scientific training inside the lab. Before starting iGEM project, seldom had we truly stepped outside the lab to know about the realistic demands of the world and never had we understood the distance between what we were and what we would be. However, what is always branded on our mind is that human practice bridges the gap among us young researchers, experts and stakeholders. It is the bridge that facilitates us with insights into step-by-step adjustment of our project. Therefore, although in the special time of pandemic, we elaborately designed our online and offline human practice plans and started the people-oriented journey in a systematic manner.

This is where our dream begins.

Summary

In ZJU iGEM’s project this year, our product MagHER2some, a novel contrast agent for MRI breast cancer examination, is tightly related to clinical treatment. Developing the effective product in the lab is a demanding job, whereas it is more of a challenge to optimize the product that fits in the clinical use. To validate our plan, we first made a framework of whole process of regular breast cancer management and according to the framework, we went on practice from downstream to upstream step by step to explore and shape our product design.

Figure 1. The framework of our integrated human practice. According to the process of breast cancer management, we carried out our human practice from the downstream back to the upstream.

On the one hand, what we are working on is derived from what we learn from the reality, ranging from investigating background, designing scheme, to closing the loop, through dialogues with patients, medical specialists in breast cancer, radiologist, researchers in pharmocochemistry, etc. On the other hand, we interviewed company researchers working on the field of molecular contrast agents and referred to relevant regulations to learn on how our product will influence the world in consideration of all aspects, including effects, manufacturing, safety, ethics, etc. We also did some preliminary work by consulting medicine company for the establishment of our future platform of alternative targets.

Video 1. Constructive insights from all people concerning our project, from downstream to the upstream of the breast cancer management.

Based on the two-way interactive mode of human practice, in this part, we summarized the most critical insights from the people we met and how we response to the feedback by making impressive changes in our design and experiments.

Inspiration from the Screen

When we were watching the TV program, The Reader, we were deeply moved by words from Zhang Jiamin, who had survived form the breast cancer and now dedicated to breast cancer rehabilitation voluntary jobs. She found that many breast cancer patients faced physical disability and great pain from the fear of recurrence. Therefore, she opened a 24-hour hotline for breast cancer patients and helped them nearly 10,000 times in more than 20 years, bringing courage and hope to many people. Here's a photo of her on the show, with a pink ribbon on her chest.

Figure 2. Zhang Jiamin in the TV program, The Reader.

We were deeply moved when she said that she hoped that "Chinese women would not be plagued by breast cancer any more", so we hoped to carry forward this belief and make some contribution to breast cancer patients in China by applying synthetic biology methods. Further information at Inspiration page.

Explore Breast Cancer Issue

We aimed to develop a functional contrast agent with the innovative design of magnetosome. The target clients of the product are doctors and breast cancer patients. We cared about the feasibility to make a difference in the diagnosis and therapy of breast cancer with synthetic biology.

Inner voice of breast cancer patient

When we had an interview with a female breast cancer patient with fair prognosis, she told us the feeling she had after diagnosis of the disease. (Figure 3.) She told us her struggle with breast cancer treatment. "Breast cancer has long been far away from our real life," she said, "until one day I happened to have examination with my friend." We were informed of the treatment methods available, the treatment methods she finally chose and her personal experience of MRI examination. Her concerns about treatment choices guided us to think towards where we could improve.

Figure 3. Interview with a breast cancer patient. (NOTE: photo taken and published with interviewee’s consent, and photo taken from backwards was to protect their privacy and information)

Interview with Dr. Fu

In order to shape our basic biological design, we are required to have a specific knowledge of the breast cancer from the angle of medical specialists in breast cancer. We met Dr. Fu in her office in The First Affiliated Hospital, Zhejiang University School of Medicine. Dr. Fu first generally introduced the issue of breast cancer and then in further details, she went on in its standard examination modalities, molecular subtypes and existing therapy. We also asked about trends in breast cancer management and reshaped the role that MRI played in the breast cancer therapy. Different from what we used to learn from the articles, in China, MRI was used more often in the real-time evaluation in response to therapy than in the diagnosis. This was partly related to its high cost. Additionally, given that each particular molecular subtypes of breast cancer (e.g. HER2 positive) accounts for a relatively small part of the total cases, engineering contrast agents targeting molecule may fail to fulfill their potential in the diagnosis when facing the patients of unknown subtypes. Yet there have been little broad-spectrum molecular biomarkers for all subtypes. Still, efforts to endow the MRI method with molecular specificity in evaluation process are rewarding, Dr. Fu said.

It was after the deep talk with Dr. Fu that we fortunately learned the significance of approaching local medical specialist in breast cancer, who can unveil the local problem clearly and sharply. Accordingly, we shifted our orientation from the diagnosis of the disease to the evaluation of post-therapy responses and chose to target the HER2 positive breast cancer, the subtype with poor prognosis and malignant rate, in our following works. (Click HERE to see the interview record)

Know about MRI Method

MRI was an interdisciplinary technology of great complexity, so it’s necessary to view the project from the angle of radiologist. During the whole MRI examination, doctors from radiology department are responsible for operating the examination and analyzing images, thus playing an indispensable role in assisting medical specialist in breast cancer to make judgements. In other word, they are the direct users of our products who inject the contrast agent into person. We were fully convinced that their experience in MRI can offer us resourceful insights into our design. Then we consulted Prof. Chen from The First Affiliated Hospital, Zhejiang University School of Medicine to clear our confusion in MRI.

Prof. Chen found it exciting to learn that we focused on the improvement of contrast agents and was interested in the novel design of magnetosomes. For one thing, he agreed with our ideas that the magnetosome enclosing iron mineral as active constituent, had lower cellular toxicity and less side effects than traditional agents. He also pointed out the lack of specificity in MRI examination could be the bottleneck of its further application, because much diagnosis today had still been relied on the empirical judgements due to the unspecific imaging. For another, we should consider the size and the metabolic model of magnetosomes, for inappropriate size may lead to phagocytosis in liver or fast removal by blood flow. Coincidently, both of us favored the idea of integration of evaluation and treatment by enriching the functions of the mono-function contrast agent.
(Click here to see other functions in Future work page)

The dialogue with Prof. Chen contributed to our further consideration into the genetic design to control the structure of magnetosomes and their dynamics in modelling process. Meanwhile, when we heard the inner voice of doctors from radiology department, we solidated determination to keep working hard in saving their time in analyzing images.
(Click HERE to see the interview record)

We paid a visit to the MRI room in the hospital under the guidance of Prof. Chen. He introduced the basic steps of MRI examination to us as a frontline radiologist was performing the procedures, including preparing the testing bed, injecting contrast agent, receiving signals and analyzing imaging. Besides, Prof. Chen showed us the testing device for mice and taught us how to use it, helping us know about the device if we progressed into the animal test stage.
(Photo of the MRI room taken and published with doctors’ and patients’ consent)

Improve Current Contrast Agent

Select Target Device

A functional contrast agent with high specificity should be armed with a potent target device. After going through the articles, we were interested in several kinds target probes and decided to load the magnetosome with both the HER2-antibody and aptamers (nucleic acid antibody). We also wanted to have a try on the widely-used small peptide target molecules. Then we noted that our product was going to be qualified in manufacturing and finally came up in the market, so we had an online discussion with Prof. Zhou expert in nano-drug industry, who offered insightful suggestions on the target mechanism from the angle of chemical industry.He had been working on the research of tumor micro environment and molecular target drug for many years.

In the meeting, we had a broad review of the mechanism of target system, including the biomarker, target probe, connector and reporting molecules. Prof. Zhou pointed that to validate the high efficiency of targeting, we were required to conduct paralleled experiments among different target probes and chose the best one to link with magnetosome, because multi-target device may face the competition for target sites and result in competitive inhibition. As a result, we chose anti-HER2 as the target probe according to advice from Pro.Zhu. Besides, sensitivity of the contrast agent was also a limitation in current use, which was we can improve, he said.

Having chosen HER2 as biomarker, we finally narrowed down the focus of our design into finding appropriate HER2 antibodies and suspended our multi-target device. We then chose single chain antibody fragment (scFv) with a smaller size and higher affinity and designed a self-assembly path as its connecting method. This was a typical synthetic biological design without using the traditional chemical cross-linking reaction and was less energy-consuming in manufacturing.
(Click HERE to see the discussion record)

Cultivatate Magnetotactic Bacteria

Besides caring about the biological design, we started shifting the focus from what we will do to how we will do. Magnetotactic bacteria were known to form magnetosome which was inhibited by high levels of oxygen and occurred only under microaerobic or anaerobic conditions, but its growth correlates with dissolved oxygen. Therefore, it is critical to control dissolved oxygen concentration (DO) at an optimal level to improve magnetosome yield.

We consulted Prof. Lv, an expert in environmental microbiology and she provided unique points on how to cultivate microaerobic bacteria. She offered to use the well-designed anaerobic devices to control DO along with particular indicator of oxygen. Also we discussed three plans of achieving the goal of producing magnetosomes together and finally decided to devide the in vitro assembly of magnetosome into two processes of both magnetotactic bacteria and E.coli. Through the talk, we studied the relationships among the cell growth, magnetosome formation, DO, and the ability to supply oxygen to the cells and conceived a method for cultivation.

Safety

Safety in iGEM Project

In order to tackle with our concern about safety in our project, we referred to a considerable number of rules and policies, and took prudent attitude towards what we were working on and what might pose risk to the world.

On 23^rd, June, we took part in the special meeting, Risk Identification Workshop-iGEM2020 online, held by iGEM foundation. All of our team members cautiously thought about the examples and principles given in the meeting. In the Q&A part, we had a one-to-one communication with Dorothy Zhang, from iGEM safety committee. We briefly introduced our project design and mentioned possible risks to her. We also talked about how we planned to carry out our animal experiments and solve the safety problem when using it. (Due to limited time, we didn’t progress into animal test stage.) This meeting helped us trouble shoot problems in our safety form and drove us to think further in safety problems. Additionally, the meeting with Genescript Biotechnology Company offered us insights into the notes in the biosafety and bioethics of heated gene engineering techs, including gene editing with CRISPR, RNA interference, stem cell therapy et al.

During our interview, we also attached great importance to obtaining informed consent, protecting privacy and concerning vulnerable population. Prior to any interview, we have our interviewee informed the data we wanted to collect and the way we collecting data. When we visited the MRI room in the hospital, we especially asked for the permission and obeyed rules.

Safety in Product

In terms of our product, although we focused on the research stage of it, further procedures were also considered if it was to come out of market. In that contrast agent was a kind of biological agents, distinct from traditional chemical medicine, we went through the website of Zhejiang Medical Products Administration and referred to relevant rules and regulations in China. We had a better understanding of the procedures of releasing new medicine on the market through analyzing the Provisions for Drug Registration and Measures for the Supervision over and Administration of Pharmaceutical Production. Some circular telling the revelation of the side effects of contrast agents also caught our attention. We would strictly follow the regulations and completing all the testing stages before registration. This was tightly related to being responsible for our entrepreneurship.

Entrepreneurship

MagHER2some is a functional MRI contrast agent for breast cancer. We skillfully combined the magnetosome from magnetotactic bacteria with anti-HER2 scFv by synthetic biological assembly to design a specific contrast agent, which could realize precise MRI method. Our target clients were radiologists from hospitals and medical institutes, whereas medical specialist in breast cancer, patients were also concerned about our products. The relatively narrow range of oriented populations of MagHER2some decided that we had to have core competitiveness and comprehensive service to earn an edge in the vigorous market. Under the circumstances, we had an online interview with Dr.Yu, a researcher of a molecular contrast agent company and hoped to learn the market and competition of our product.

In the interview, we learned that the specificity of the MagHER2some showed privilege in filling the market niche of targeted contrast agents. For one thing, to have a cutting edge over regular gadolinium based contrast agents, we should work towards magnifying the strengths of the coral technology in our production in our evolving design, for example the higher biocompatibility causing less side effects, the low-carbon biological manufacturing. For another, there were targeted molecular contrast agent in the lab stages using small peptides. Despite the higher affinity and specificity of the scFv we used, how we can make sure the relatively lower cost in manufacturing remained critical. Additionally, he introduced to us the basic steps of the whole process for medicine authorization and establishment of trade standard of this emerging product may also deserve further discussion. (Click HERE to see the interview record.)

To validate our strategic concept of building platform with alternative targeted diseases, we also consulted Genechem Company, working on the targeted medicine of tumors. We discussed the possible paths of the screening and verification of biomarkers and the differences between producing contrast agent and medicine with targeted function.

Although the past few decades has witnessed great advance in China’s pharmaceutical industry, there is still a lack of domestic products innovated by Chinese researchers, especially anti-cancer ones. The high level of importing dependency hindered our development in disease treatment, thus inspiring us to strive for a better future in anti-cancer entrpreneurship. Further information at Implementation page.

All above contributed to our project up to date and also facilitated us with preliminary work for our strategic concept. (Click to see our Future)

Team:ZJU-China/Human Practices