Overview

In our project, we followed a clear progression from problem identification → solution design → technical implementation → market application. We realized that lab research alone cannot fully address the complexities of clinical and industrial challenges. Therefore, at each stage, we engaged key stakeholders to refine our project direction.

Through social surveys and literature review, we found that cardiovascular and cerebrovascular diseases are highly prevalent, with patient numbers increasing steadily.

​ ● Pharmacology experts and clinicians highlighted the therapeutic potential of leonurine for cardiovascular health.

​ ● Plant synthetic biology and metabolic engineering specialists guided our biosynthetic design, enabling a shift from plant-based to microbial chassis systems.

​ ● Pharmaceutical directors and industry partners provided insights on scale-up production and market readiness.

​ ● Patients and high-risk groups shared perspectives on efficacy, safety, price, and acceptability.

​ ● TCM practitioners and regulatory stakeholders ensured scientific credibility and cultural acceptability.

This stakeholder-driven approach ensures our project is technically feasible, practically translatable, and socially responsible, bridging innovation, industrial value, and public trust. All these practices are goal-oriented and effectively implemented, making our project not only a scientific exploration but also a direct response to real clinical and societal needs, allowing synthetic biology to better fulfill its mission of improving cardiovascular health.

Highlights

Human practices ran through our entire project cycle, encompassing problem identification, solution design, engineering, application scenario recommendations, industrial production chains, as well as the integration and potential conflicts of patient needs and traditional cultural values, making our project more likely to be practically implementable.

Stakeholder mapping

Guided by market-driven design principles, we classified the stakeholders of the Leonurine project using Mendelow’s Matrix, considering both “interest level” and “influence.” Groups with high interest and high influence, such as cardiologists, pharmacology experts, and pharmaceutical company leaders, were regarded as core participants; groups with high influence but relatively low interest, such as traditional Chinese medicine practitioners, need to be kept satisfied; cardiovascular patients and high-risk populations, although having limited influence, were placed in the key-informed category due to their strong concern for efficacy and safety; the general public was monitored overall. This classification helps us focus on critical feedback and optimize design and dissemination strategies.

Problem identification

Cardiovascular disease (CVD) is the leading cause of death globally. According to the World Health Organization (WHO), an estimated 19.8 million people died from CVDs in 2022, accounting for 32% of all global deaths[1]. Among these, 85% of deaths were due to heart attack and stroke, and over three-quarters of CVD deaths occurred in low- and middle-income countries[1]. In China, against the backdrop of a progressively aging population, CVD remains the primary cause of mortality among both urban and rural residents. In 2021, it was responsible for 48.98% and 47.35% of all deaths in rural and urban areas, respectively — meaning approximately 2 out of every 5 deaths were attributed to CVD[2].

Data from the Global Burden of Disease (GBD) study show that between 1990 and 2019, the number of incident CVD cases in China increased from 5.30 million to 12.34 million, a rise of 132.82%, while deaths from CVD increased from 2.42 million to 4.58 million, an increase of 89.12%[3].

Given this severe disease burden, we conducted a questionnaire survey to further investigate the prevalence of CVD among people in our community and their satisfaction with currently available medications.

Proportion of Major Causes of Death Among Rural and Urban Residents in China, 2021(Source: Chinese Circulation Journal, June 2025, Vol. 40, No. 6 (Serial No. 324))

Public survey

To understand the real-world prevalence of cardiovascular and cerebrovascular diseases in our community, we conducted a questionnaire survey with 804 participants. The results revealed that 52.24% of respondents either suffered from or had family members affected by cardiovascular diseases, including hypertension, hyperlipidemia, coronary heart disease, or stroke. This figure is slightly higher than the data reported by authoritative institutions (compared to 48.98% and 47.35%), which we hypothesize may be attributed to the limited sample size or a further increase in the prevalence of these diseases in recent years.

We further investigated the satisfaction level of this population with current drug treatments and found that 50.82% of them were not very satisfied. Among the reasons for dissatisfaction, significant side effects and unstable efficacy of medications were the two most prominent concerns. Therefore, given the large and continuously growing number of affected individuals, coupled with the limitations of existing therapies, we aim to employ synthetic biology approaches to develop an auxiliary treatment strategy for cardiovascular and cerebrovascular diseases.

Project inspiration

From the very beginning, since our team was formed at the end of 2024, the development of novel therapeutics for cardiovascular and cerebrovascular diseases has remained at the forefront of our minds. In April this year, our school organized a study tour to Chenshan Botanical Garden—an institution that also served as the guiding partner for our school’s NAIS team last year.

During this visit, the researchers responsible for germplasm conservation and medicinal plant development introduced us to the garden’s remarkable biodiversity and to the cultivation and pharmacological properties of various traditional Chinese medicinal herbs. It was here that we first encountered Leonurus japonicus, known in Chinese as “yi mu cao”. For over two thousand years, this herb has been widely used in traditional medicine, particularly for gynecological disorders, due to its well-recognized efficacy in promoting blood circulation and removing blood stasis. This experience sparked our attention toward its key bioactive component, leonurine.

Anchoring Direction

a. Pharmacology Expert Interview

Interviewee: Dr. Yizhun Zhu

Institution: Dean of the Faculty of Pharmacy, Macau University of Science and Technology, and Chair Professor at Fudan University

Field of research: Molecular pharmacology, development of active components in traditional Chinese medicine, and basic/clinical research on cardiovascular & cerebrovascular diseases.

Inspiration

Through interactions with patients, we identified a core issue: although current cardiovascular drugs can delay disease progression, they often cause side effects or provide insufficient efficacy, leading to poor long-term adherence. Therefore, from a pharmacological perspective, we aim to explore novel auxiliary treatment strategies and seek guidance from leading experts in this field.

Intercommunication

We contacted Professor Yi-Zhun Zhu, who has long been engaged in pharmacological research on natural products. During our discussions, Professor Zhu proposed a direction worthy of in-depth exploration: the potential application of leonurine in the treatment of cardiovascular and cerebrovascular diseases. He not only shared relevant experimental data but also recommended published literature from his research team, providing us with a solid scientific foundation for our work.

Investigation

Professor Zhu elaborated on the molecular mechanisms of leonurine in cardiovascular and cerebrovascular diseases:

1. Anti-platelet aggregation: Improves hemorheological properties by inhibiting thromboxane A₂ (TXA₂) production and upregulating prostacyclin (PGI₂) levels

2. Antioxidant and anti-inflammatory effects: Modulates reactive oxygen species (ROS) levels, suppresses the NF-κB pathway, and reduces the release of inflammatory factors

3. Protection of endothelial function: Promotes nitric oxide (NO) synthesis and slows the progression of atherosclerosis

He emphasized that while these mechanisms have been validated in animal studies and in vitro models, randomized controlled clinical trials remain essential for future translation. Furthermore, he highlighted that if leonurine can be produced with high purity in a scalable and controllable manner through synthetic biology approaches, it holds significant potential as an adjunctive therapy for cardiovascular diseases.

Implementation

Based on Professor Zhu’s recommendations, we have incorporated a focus on validating the molecular mechanisms of leonurine into our project design, while clearly prioritizing stability and controllability as core criteria for the synthetic strategy. This approach not only aligns with the key pharmacological principle of transitioning "from bench to bedside," but also enables us to establish a more coherent and targeted technical pathway.

Impact

Through the interview with Professor Zhu, we gained a novel research direction: exploring leonurine as a potential candidate for adjunctive therapy in cardiovascular and cerebrovascular diseases. His research findings and insights have not only strengthened the scientific foundation and applicability of our project, but also provided a robust basis for subsequent communication with healthcare professionals, pharmaceutical companies, and the public.

b. Clinical Physician Interview

Interviewee: Dr. Huanchun Ni

Institution: Fudan University Huashan Hospital

Field of research: Cardiology

Inspiration

Following Professor Zhu Yizhun's proposal of leonurine as a novel therapeutic approach, we sought to further evaluate its clinical feasibility and therefore consulted with Chief Physician Ni Huanchun, who has extensive experience in diagnosing and treating cardiovascular diseases.

Intercommunication

Dr. Ni pointed out that cardiovascular patients often rely on long-term multi-drug regimens—such as antihypertensive, anticoagulant, and lipid-lowering medications—which frequently led to side effects and poor adherence. As a result, there is a clear and urgent clinical need for new auxiliary treatment options.

Investigation

He particularly emphasized that if leonurine can demonstrate high safety, minimal drug interactions, and tangible benefits in improving blood circulation or reducing vascular events, it would hold significant appeal for patients. However, he also cautioned that its efficacy must be substantiated through rigorous clinical trials.

Implementation

In response to Dr. Ni’s advice, we have incorporated drug interaction studies and side effect assessments into our project design. This ensures that synthetic leonurine is not only feasible at the experimental stage, but also aligned with the practical needs of both clinicians and patients.

Impact

Dr. Ni’s feedback made us realize that the true value of the synthetic leonurine project lies not only in its scientific innovation, but also in its ability to address genuine clinical challenges. His insights have provided our research with a clearer clinical orientation.

Exploring strategies

a. Research Progress on the Biosynthetic Pathway of Leonurine

Interviewee: Dr. Ping Xu

Institution: Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences; Chenshan Botanical Garden Science Center

Field of research: Specializes in plant natural products, with a focus on discovering medicinally valuable plants and elucidating their biosynthetic pathways.

Inspiration

After deciding to focus on leonurine as a candidate compound for developing new therapeutics for cardiovascular and cerebrovascular diseases, we returned to the institution where we had conducted our study tour in April: Chenshan Botanical Garden. We aimed to consult with experts who were directly engaged in research on leonurine biosynthesis.

Intercommunication

We interviewed Professor Ping Xu, who has been deeply involved in studying natural products in medicinal plants. She explained that, to date, there have been no international reports on the synthetic biology-based production of leonurine. The only progress so far was published by her lab in 2024, in the article “Multiomics analyses of two Leonurus species illuminate leonurine biosynthesis and its evolution” (Li et al., 2024).

Investigation

Professor Xu’s team successfully identified the two key enzymes catalyzing the final steps of leonurine biosynthesis, LjUGT5 and LjSCPL12, and validated their catalytic functions in tobacco leaves and yeast cells. However, the yields obtained were extremely low—approximately 0.14 µg/mL in yeast, and even lower in transient tobacco expression. She emphasized that these levels are far from industrial feasibility (≥60 mg/200 mL), highlighting the challenges of producing leonurine in heterologous systems.

Implementation

In her guidance, Professor Xu pointed out that to pursue large-scale production, one should consider more mature commercial chassis systems, such as hairy roots, which already have precedents for scale-up using large fermenters. This insight redirected our focus: instead of attempting to engineer leonurine production in less efficient systems, we should prioritize platforms with proven scalability.

Impact

Through this interview, we gained a realistic understanding of the current research frontier in leonurine biosynthesis and the significant gap between academic validation and industrial application. Professor Xu’s expertise provided us with not only the latest knowledge on pathway elucidation but also practical advice on feasible production strategies. Her guidance reinforced the importance of integrating cutting-edge plant science with industrial biotechnology, which shaped the foundation of our subsequent chassis selection and project design.

b. Plant Synthetic Biologist Interview

Interviewee: Prof. Qing Zhao

Institution: Shanghai Chenshan Plant Science Research Center, Chinese Academy of Sciences

Field of research: Plant Biotechnology and Plant Chassis-based Synthetic Biology

Inspiration

After confirming the pharmacological potential of leonurine, we needed to urgently assess the feasibility of its biosynthesis in plant systems. Whether Leonurus japonicus — the natural source of leonurine — could serve directly as a production chassis was our initial question.

Intercommunication

In discussions with Dr. Zhao Qing, he affirmed the scientific value of producing leonurine via plant cell factories and indicated that a hairy root culture system of Leonurus japonicus could, in theory, generate the target compound. However, he also pointed out practical limitations such as lengthy culture cycles, difficulties in genetic manipulation, and low yield, which may hinder scalable production.

Investigation

Regarding our question about choosing between Leonurus and tobacco hairy roots, Dr. Zhao clearly recommended:

While Leonurus hairy roots can produce leonurine, their low genetic manipulation efficiency and limited yield (due to low accumulation in root tissues) make them impractical for efficient production.

In contrast, tobacco hairy roots, as a versatile synthetic biology chassis, offer easier genetic transformation, rapid proliferation, and high biomass accumulation, making them better suited for high-efficiency leonurine synthesis.

He advice prompted a critical pivot in our chassis selection.

Implementation

Dr. Zhao further emphasized that there is no inherent conflict between synthetic biology and the modernization of traditional Chinese medicine. Citing the artemisinin case, he noted that if the final molecular structure is identical to the natural compound, regulatory approval does not differ significantly between microbial and plant-based production systems — alleviating concerns about “naturalness.” This perspective provides important support for future public communication and compliance evaluation.

On specific strategies, he suggested employing an inducible expression system to avoid metabolic burden. He recommended initially testing whether the target molecule can be produced — an approach well-suited to the iGEM timeline.

Impact

Dr. Zhao’s guidance helped us avoid a “theoretically feasible yet practically challenging” route early on. He recommendations enabled our project to strike a balance between scientific rigor and practical operability, resulting in a clearer technical trajectory. Moreover, he clarification on regulatory compliance and naturalness not only enhanced the project’s feasibility and industrial potential but also laid a solid foundation for future discussions with metabolic engineering experts on the complementary use of microbial and plant systems.

c. Metabolic Engineering Expert Interview

Interviewee: Prof. Menghao Cai

Institution: School of Biotechnology, East China University of Science and Technology

Field of research: Microbial Synthetic Biology

Inspiration

In our initial synthesis strategy for leonurine, we primarily focused on plant hairy root systems. However, from an industrial perspective, relying solely on plant chassis may present challenges related to limited scalability and higher production costs. Therefore, we needed to re-evaluate the potential of using Escherichia coli as an alternative microbial production chassis.

Intercommunication

Dr. Cai Menghao emphasized that E. coli offers inherent advantages in synthetic biology and metabolic engineering due to its rapid growth, ease of genetic manipulation, and well-established engineering tools, making it particularly suitable for industrial scaling. While he acknowledged that E. coli might face challenges in maintaining the activity of certain enzymes, its superior controllability and cost-effectiveness in large-scale production make it a more promising option compared to plant-based systems.

Investigation

To address the complexity of leonurine synthesis, Dr. Cai proposed two key strategies:

​ ● Exogenous Feeding: Supplementing the E. coli system with inexpensive and readily available precursors or intermediates to significantly enhance synthesis efficiency and reduce metabolic burden.

​ ● In Vitro Enzymatic Reactions: Employing purified enzymes to catalyze specific "bottleneck reactions" outside the cell, thereby bypassing cellular metabolic constraints and improving yield and process control.

These insights provide practical pathways for our future exploration of microbial production systems.

Implementation

He advised that, within the timeframe of the iGEM competition, we should first focus on functional validation of the core synthetic genes in E. coli, and investigate whether adding exogenous precursors can facilitate product formation. For the in vitro enzymatic approach, he suggested designing a preliminary proof-of-concept experiment—especially under limited experimental conditions—without initially aiming for a fully optimized process. This implementation route lowers the barrier to experimentation while accumulating experience for future industrial applications.

Impact

Dr. Cai’s recommendations prompted a pivotal shift in our project direction: from plant chassis to exploring microbial alternatives. Moreover, his emphasis on the industrial potential of exogenous substrate feeding and in vitro enzymatic strategies not only refined our understanding of scalable leonurine production but also provided a strategic foundation for future hybrid approaches combining microbial fermentation and cell-free enzymatic catalysis.

Implementing

a. Industrial chain

Interviewee: Dr. Lu Jin

Institution: Corporate Leader, Shanghai Zensun Pharmaceutical Co., Ltd.

Core Company Drug Product: An innovative medication for the treatment of heart failure.

Inspiration

In exploring the industrial prospects of leonurine, we recognized that confining the research to academic laboratories would be insufficient to advance it into clinical application. To better understand the requirements of pharmaceutical companies regarding API supply, process stability, and market development, we consulted Dr. Jin Lu, a pharmaceutical plant director.

Intercommunication

Dr. Jin first acknowledged the potential of Leonurine in the cardiovascular field and noted that achieving a stable supply via synthetic biology would greatly alleviate current issues caused by reliance on natural herbs, such as inconsistent yields and significant quality variations. She emphasized that, as downstream users, pharmaceutical manufacturers are primarily concerned with the standardization, purity, and batch-to-batch consistency of raw materials, as these factors directly determine the feasibility of drug development and formulation.

Investigation

Regarding future technical pathways, Dr. Jin highlighted:

​ ● Cost and Scale: To enter the pharmaceutical market, the synthetic route must be cost-competitive or superior to existing extraction processes.

​ ● Process Compliance: Industrial production must adhere to GMP standards and consistency evaluation required by drug regulatory authorities, imposing stringent requirements on API manufacturing.

​ ● Collaboration Model: She specifically pointed out that if academic teams can supply high-purity, standardized leonurine API, pharmaceutical companies would be highly interested in subsequent formulation development and commercialization.

Implementation

In response to her feedback, we have further refined our design strategy to prioritize purity, stability, and scalability in both metabolic engineering and in vitro enzymatic processes. Moreover, we have repositioned our laboratory output as API supply for pharmaceutical manufacturers rather than direct entry into the retail market, establishing a clearer business-to-business (B2B) orientation.

Impact

Dr. Jin’s advice helped clarify the industrial endpoint of our project: the value of synthetic leonurine lies not only in scientific innovation but also in providing pharmaceutical companies with stable and standardized raw materials. This has sharpened our industrial logic and laid a foundation for future collaboration with drug manufacturers. Furthermore, her feedback underscored that bridging the gap between technology and the market requires a focus on cost control and regulatory compliance, which will serve as critical guiding principles in our ongoing design optimization.

b. Traditional Chinese Medicine (TCM) Practitioners as Channel Partners

Interviewee: Mr. Zhou

Field of research: Chinese herbal medicine cultivators and gatherers, Traditional Chinese Medicine (TCM) Practitioner

Inspiration

Following discussions with pharmaceutical manufacturers regarding industrial pathways, we recognized that, in addition to industrial demand, acceptance and promotion of leonurine would also be influenced by attitudes within traditional Chinese medicine clinical practice. TCM physicians, who have long relied on herbal medicines, might express skepticism toward drugs derived from novel synthetic biology approaches. We therefore sought to gain deeper insight into their perspectives.

Intercommunication

During interviews, TCM physicians first affirmed the traditional value of Leonurus japonicus in gynecological and cardiovascular health, and also acknowledged the pharmacological effects of leonurine as a single compound. However, they pointed out that if the active ingredient were produced via synthetic biology rather than extracted from natural medicinal materials, it might encounter resistance from both patients and some practitioners due to perceptual differences.

Investigation

The physicians highlighted three key concerns:

​ ● Cultural and Cognitive Differences: Some patients exhibit a preference for “natural herbs” and are skeptical of “synthetically produced” compounds;

​ ● Need for Clinical Evidence: Widespread adoption of leonurine would require robust evidence-based medical data demonstrating its safety and efficacy;

​ ● Integration of Tradition and Modernity: They suggested positioning synthetic leonurine as “a TCM active ingredient backed by modern pharmaceutical technology” to alleviate ideological conflicts.

Implementation

Based on this feedback, we plan to incorporate the following into our subsequent human practices activities:

​ ● Develop public science communication initiatives emphasizing that “synthetic does not mean unsafe,” while highlighting advantages in quality stability and control ability;

​ ● Collaborate with pharmaceutical partners to generate evidence-based clinical data during formulation development to gain acceptance among physicians;

​ ● In messaging, frame the product as a modern upgrade of TCM rather than a rejection of tradition.

Impact

The feedback from TCM practitioners made us realize that market acceptance depends not only on industrial demand but also on a shift in perceptions among patients and doctors. Their insights helped us identify potential cultural challenges in future promotion efforts and guided us to proactively plan strategies for science communication and market education. This approach will support smoother integration of synthetic biology-derived products into the existing TCM system and help prevent barriers to practical application.

c. Patients and High-Risk Groups (End Consumers)

Inspiration

After receiving positive feedback from experts and industry partners, we recognized that the true test of our project’s relevance lies with end users—patients and high-risk groups. Their awareness, needs, and concerns ultimately determine whether leonurine can successfully reach the market.

Intercommunication

We conducted interviews with a 75-year-old cardiovascular patient and several elderly high-risk individuals. The patient expressed frustration with the burden and side effects of long-term multi-drug therapy, showing both hope and concern about new treatments. The high-risk group reported obtaining health information primarily through community channels and doctors, actively monitoring their health indicators, and expressing a preference for traditional Chinese medicine (TCM)-based approaches due to their perceived milder effects.

Investigation

Key insights from the interviews include:

Patients:

​ ● Fatigue with complex multi-drug regimens and noticeable side effects (e.g., dizziness, gastrointestinal discomfort, fatigue);

​ ● Initial unfamiliarity with leonurine, but openness to trying it after explanation—especially when framed as “naturally-derived”;

​ ● High price sensitivity, with an acceptable range of ¥50–100;

​ ● Concerns about drug interactions and additive side effects.

High-Risk Groups:

​ ● Preference for TCM, viewed as “gentle and suitable for long-term 调理”;

​ ● Value most: efficacy, natural origin, and affordability when choosing products;

​ ● Initial skepticism toward synthetic biology, turning to cautious optimism after education—expressed willingness to adopt “if proven effective and affordable”;

​ ● Emphasized that “minimal side effects and low cost” are the strongest motivators.

Implementation

Based on this feedback, we have adjusted our design and outreach strategy:

​ ● Communication: Highlight leonurine’s natural origins combined with efficiency gains from synthetic biology to alleviate concerns about “artificial modification” among elderly users;

​ ● Preclinical Focus: Prioritize drug interaction and safety studies to address primary patient concerns;

​ ● Market Positioning: Develop “health/prevention-grade” To C products tailored to the needs of high-risk groups;

​ ● Cost Control: Collaborate with process experts and manufacturers to keep prices within the widely acceptable range indicated.

Impact

These interactions underscored that although different groups occupy varying positions on the disease spectrum, they uniformly emphasize safety, natural perception, and affordability. Patient interviews helped us better grasp real-world treatment challenges, while input from at-risk groups revealed the significant potential of our project in prevention and health management. This not only validates the practical relevance of synthetic leonurine but also broadens potential product development pathways.

Summary

Overall, our Human Practices journey for the Leonurine project reflects our continuous reflection and dedicated exploration of the relationship between humans, medicine, and health. By actively engaging with diverse stakeholders—from cardiologists and pharmacology experts to pharmaceutical company leaders and high-risk patient populations—we incorporated a broad spectrum of human needs into the design and implementation of our project. Through addressing societal gaps, including raising awareness of cardiovascular health and introducing modernized approaches to traditional medicine via synthetic biology, we endeavored to improve human well-being in a practical and meaningful way.

Furthermore, by persistently exploring both scientific and industrial fronts—from optimizing microbial and plant-based production platforms to considering regulatory pathways and patient preferences—we strived to balance innovation, safety, and accessibility. This project not only advances the application of synthetic biology in cardiovascular health but also highlights how human-centered design can bridge science, society, and culture. Our journey does not end here; with the continuous curiosity and commitment of our team, we will further pursue strategies that harmonize cutting-edge biotechnology with real-world medical and societal needs.

Reference

1. WHO. Cardiovascular Diseases (CVDs) Factsheet, 2021. https://www.who.int/zh/news-room/fact-sheets/detail/cardiovascular-diseases-(cvds).

2. National Center for Cardiovascular Diseases & The Writing Committee of the Report on Cardiovascular Health and Diseases in China. (2025). 中国心血管健康与疾病报告 2024 概要. 中国循环杂志, 40 (06), 521-559.

3. Institute for Health Metrics and Evaluation (lHME). Global Burden of Disease Study 2021 (GBD 2021) results[EB/OL].(2022-11-07) [2025-4-26]. https://vizhub.healthdata.org/gbd-results/.

4. Peng Li, Mengxiao Yan , Pan Liu , Danjie Yang , Zekun He, Yun Gao, Yan Jiang, Yu Kong, Xin Zhong, Sheng Wu, Jun Yang, Hongxia Wang, Yanbo Huang, Le Wang, Xiaoya Chen, Yonghong Hu, Qing Zhao, Ping Xu*. Multiomics analyses of two Leonurus species illuminate leonurine biosynthesis and its evolution, Molecular Plant, 2024, 17(1): 158-177.