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Notebook 2025

Overview

This year, our journey was meticulously documented, capturing every triumph, every setback, and every pivotal moment across our wet lab experiments, computational modeling, software development, and crucial stakeholder engagements, including our extensive troubleshooting efforts

January
Week 1 (18th of January to 24th of January) Dry lab
Our efforts focused on a comprehensive analysis of previous iGEM projects and extensive reading of synthetic biology literature.
Week 2 (25th of January to 31st of January) Dry lab
We identified asthma as a significant global health issue, noting the pressing unmet needs in treatment adherence and the management of severe type 2 asthma.
Human Practice
We began designing application forms for the recruitment of new team members.
February
Week 3 (1st of February to 7th of February) Dry lab
We continued to review previous iGEM projects, shifting our focus towards primary synthetic biology literature to develop a therapeutic approach for asthma.
Human Practice
We gathered opinions from over 15 asthmatic patients, helping us understand the challenges they face in daily life and with medication adherence, which was instrumental in shaping our solution to patient needs.
Integrated Human Practice
Our team visited Al-Abassya Chest Hospital, where we held a productive meeting with Dr. Sameh Ahmed. This discussion provided valuable insights into the pathogenesis, symptoms, triggers, and current treatment modalities of asthma.
Week 4 (8th of February to 14th of February) Dry lab
The entire team engaged in extensive discussions, brainstorming numerous ideas for the development of a potential approach to asthma treatment.
Human Practice
The recruitment of new team members was successfully completed.
Week 5 (15th of February to 21st of February) Human Practice
An urgent team meeting was convened to make a final decision on the core technology to be used in our approach.
Dry lab
We settled on developing a therapeutic approach specifically targeting IL-13.The recruitment of new team members was successfully completed.
Week 6 (22nd of February to 28th of February) Dry lab
We deepened our literature analysis of IL-13, and meticulously reviewed previous iGEM projects related to this cytokine.
After understanding asthma pathogenesis, we shifted our focus to TSLP. Notably, TSLP is an upstream cytokine that drives both type 2 and non-type 2 inflammatory pathways, thus offering a broader therapeutic target.
March
Week 7 (1st of March to 7th of March) Human Practice
Our team attended the Pulmo-Port Said Conference, where we actively explored the competitor landscape and identified potential partnership opportunities.
Integrated Human Practice
We organized a meeting with Dr. Ahmed Beshir at the conference to delve into patient adherence problems, side effects of current treatment, and emerging therapeutic approaches such as TSLP inhibitors.
Week 8 (8th of March to 14th of March) Dry lab
Our team intensified its literature analysis concerning TSLP and began brainstorming various technologies that could be incorporated into our approach.
Integrated Human Practice
We arranged another meeting with Dr. Ahmed Bashir to discuss the latest advancement in asthma treatment, particularly anti-TSLP therapy. Our goal was to explore solutions that could reduce medication frequency, minimize side effects, and improve the quality of life for patients.
Week 9 (15th of March to 21st of March) Dry lab
After careful consideration, we decided to utilize Antisense Oligonucleotides (ASOs) to knock down the TSLP activity at the transcriptional level.
We designed specific ASO sequences engineered to target and bind to particular mRNA sequences (TSLP mRNA), thereby inhibiting its expression.
Week 10 (22nd of March to 28th of March) Modelling
A comprehensive ASO model was developed and performed to validate its ability to effectively degrade TSLP mRNA, primarily through RNase H-dependent mechanisms.
Integrated Human Practice
Our team attended a meeting with Dr. Mohamed Hussein, who clarified the drawbacks of ASO, emphasizing the need for chemical modification and potential conflict with philosophical principles. These insights prompted us to explore alternative targeting of TSLP.
Week 11 (29th of March to 4th of April) Dry lab
We embarked on a renewed review of synthetic biology literature and sought expert opinions. We reaffirmed our commitment to developing a therapeutic approach targeting TSLP.
We shifted our focus toward CO-BERA, recognizing its advantage in not requiring chemical modification to achieve its function and its ability to be expressed endogenously from bacteria.
Human Practice
Our innovative approach was named “PRESS”.
April
Week 12 (5th of April to 11th of April) Modelling
The CO-BERA model was completed, which validated its ability to degrade TSLP mRNA effectively with the aid of essential enzymes that exist in human cells. The result of this model significantly outperformed those of the ASO model.
Software
Our team initiated a molecular dynamics simulation by understanding our project parameters and identifying the most suitable tool for simulating our complexes. This involved preparing DNA sequences for each protein within the complex and determining additional parameters required for accurate simulation.
Integrated Human Practice
We joined a meeting with DR. Atef, who validated siRNA as an effective silencing mechanism and supported using the Golden Gate method at Assembly.
Week 13 (12th of April to 18th of April) Dry lab
We investigated compatible technologies for CO-BERA delivery to the distal airway.
We decided to utilize liposomal vesicles for direct CO-BERA delivery into the distal airway tract.
Integrated Human Practice
We joined a meeting with Dr. Ragda to validate the process of CO-BERA-siRNA interactions, aiming to reduce off-targeting probabilities.
Week 14 (19th of April to 25th of April) Dry lab
We conducted a detailed literature analysis of liposomal vesicles, which revealed their short half-life in vivo and the necessity for repeated administration.
Integrated Human Practice
We organized a meeting with Dr.Hazman, who expressed reservations about using liposomes. He reassured us about the potential of bacterial vehicles and clarified potential side effects and sources for purchasing suitable bacterial strains.
Week 15 (25th of April to 2nd of May) Dry lab
Our delivery system was revised to explore the use of Corynebacterium species.
Human Practice
We attended World Asthma Day, gathering crucial feedback on asthma prevalence, patient perspectives, and suitable inhaler designs to further refine our “PRESS” approach.
May
Week 16 (3rd of May to 9th of May) Dry lab
We transitioned from Corynebacterium abundance to a respiratory microbiome called Lactobacillus plantarum.
We thought about the ideal device for engineered Lactobacillus plantarum delivery.
Human Practice
We attended the 5th Arab Sustainable Development Week, which encouraged us to align PRESS to serve the SDGs goals.
Software
We prepared our parameters and topology files using the CHARMM-GUI Package, and proceeded to simulate these files using Gromacs software.
Week 17 (10th of May to 16th of May) Dry lab
Our team settled on an aerosol device for the delivery of our engineered bacteria.
Software
Gromacs simulation progressed through eight distinct steps for each complex, comprising one minimization, six equilibration phases, and one production step.
Integrated Human Practice
Dr. Souzan Fouad joined us in a meeting to clarify the drawbacks of nebulizers and recommended using Dry Powder Inhalers (DPIs) as a superior delivery method.
We had a valuable meeting with Dr. Tarek Salim, who outlined the differences between inhaling devices, confirming DPIs as more patient-friendly and safer. He also recommended advanced technology, specifically an artificial lung, for preclinical testing.
SDGs
We focused on SDG 3 – Good Health and Well-being to reduce the worldwide burden of asthma, offering a treatment like PRESS.
Week 18 (17th of May to 23rd of May) Dry lab
Based on extensive literature review, experimental data, and expert opinions, our dry lab definitively settled on the usage of Dry Powder Inhalers (DPIs).
We began researching a spray drying method for bacterial drying.
Modelling
We developed a spray drying model using MATLAB to predict bacterial viability, which unfortunately indicated low bacterial viability and poor storage features.
Week 19 (24th of May to 30th of May) Human Practice
To address the drying challenge, our team members visited EIPICO Pharmaceutical Factory, where discussions led to the selection of freeze-drying as the ideal drying method for preserving bacterial viability.
Integrated Human Practice
We attended a meeting with Dr. Wagida Anwar (IRB), who emphasized the critical importance of patient safety, biosecurity, and ethical standard considerations for PRESS research.
Week 20 (31st of May to 6th of June) Dry lab
Following a comprehensive review of expert opinions and experimental data, our dry lab confirmed the selection of the freeze-drying process.
Modelling
We developed a freeze-drying model using MATLAB to predict the powder potency by calculating the number of viable bacteria per milligram of powder, which was successfully completed.
Integrated Human Practice
We discussed with Mr. Yahia Badawy, who clarified Egyptian regulatory steps for releasing a new drug to market, starting from preclinical studies.
June
Week 21 (7th of June to 13th of June) Dry lab
We recognized that a small amount of CO-BERA could be efficiently loaded into membrane vesicles, so we began searching for suitable RNA-binding proteins (RBPs) for CO-BERA loading.
Integrated Human Practice
We joined a meeting with Mr. Rejeesh (FDA Consultant), who highlighted the importance of preclinical studies, cytotoxicity testing, and sustainable planning for FDA approval.
Education
We planned through our educational program to manage visits to all age groups and different occupations. Aiming to raise asthma awareness and introduce synthetic biology aligned with life skills.
Week 22 (14th of June to 20th of June) Dry lab
We initially settled on Cold Shock Proteins (CSPs) for RNA loading into the membrane vesicles.
After further literature analysis and review, we shifted our focus from CSPs toward synthetic TMP–linker–RBP.
Software
Molecular dynamics simulation for CO-BERA-CD box complex with a protein consisting of RNA binding protein and linker (L7Ae) and transmembrane binding protein (Dough).
We tested the membrane orientation and stability of synthetic TMP–linker–RBP fusions through in-silico studies with SWISS-Model, CHARMM-GUI, and GROMACS simulations.
Week 23 (21st of June to 27th of June) Dry lab
Our team aimed to target NO for the conditioned expression of CO-BERA, ensuring its release only in the context of asthma.
Modelling
NO conditioning model developed using CO-LAB, demonstrated the possibility of CO-BERA release in the presence of NO, a known biomarker of asthma. However, it also revealed the impossibility of using this switch in Lactobacillus plantarum (LB) due to the absence of necessary activator factors.
Integrated Human Practice
We met with Dr. Mohamed Anany to identify the strength of PRESS in solving adherence problems; he also highlighted its potentially high cost and safety concerns.
Education
We conveyed to New Academy kindergarten to teach kids there the proper inhaler technique through songs and educate them about asthma triggers using activities and our children's book. Additionally, we emphasized the emergency steps for asthma attacks.
Week 24 (28th June to 4th July) Dry lab
We shifted our focus toward utilizing H₂O₂ to stimulate CO-BERA expression, as H₂O₂ levels are known to increase in the inflammatory response of asthma.
Modelling
H2O2 conditioning model developed using CO-LAB, proved the high efficiency of CO-BERA under increased H2O2 conditions, which is specific to airway inflammation.
Software
We further tested H₂O₂ conditioning through in-silico studies, confirming that H₂O₂ successfully achieved spatial and temporal control of gene expression in stressed bacterial environments.
Integrated Human Practice
We held a meeting with Mr. Mohamed Mahrous (AstraZeneca) to demonstrate the viability of PRESS’s and explore a possible future collaboration for global market entry.
Education
We organized a visit to Al-Salam preparatory school, where we detailed the improper inhaler technique side effects, and then we clarified the proper inhaler technique.
July
Week 25 (5th of July to 11th of July) Dry lab
We discovered that CO-BERA experienced significant degradation before achieving its intended function, prompting us to research methods for enhancing endosomal escape.
Modelling
We developed an H2O2 DIFFUSION model in the bacteria using CO-LAB that confirmed an increase in the diffusion rate of H₂O₂ into the bacteria, reaching above the threshold of pKat promoter.
SDGs
We committed to SDG 13 – Climate Action to reduce emissions harming our planet’s climate from normal inhalers.
Week 26 (12th of July to 18th of July) Dry lab
Following a thorough literature review and analysis, we settled on Listerolysin O (LLO), recognizing its ability to disrupt endosomal membranes and enhance endosomal escape.
Wet lab
Concurrently, the wet lab team initiated comprehensive training sessions to enhance technical proficiency, thereby preparing actively for subsequent scientific discussions and strategic planning meetings.
Integrated Human Practice
We organized a meeting with Dr. Asmaa Mohamed, who expressed reassurance about PRESS’s potential to reduce psychological burdens and enhance patients’ well-being.
We had a meeting with Engineers El Sebay & Mohammed, who provided valuable tips regarding our asthma-friendly city design, including air filter placement and renewable energy usage.
Human Practice
Our team visited El-Zagazig University Hospital to collect feedback from doctors and patients, which identified concerns about fear of bacterial use, emphasizing the need for robust safety measures and effective educational awareness campaigns.
Education
We made specific materials for family members and used them through our visits, such as a child's book, a parent's side book, and a song and game about asthma management.
Week 27 (19th of July to 25th of July) Dry lab
We decided to utilize a mutated form of LLO, as this variant is considered safer and prevents the pathogenicity of normal LLO.
Our overarching goal remains to develop an approach with the highest safety level, leading us to investigate technologies that could further enhance the safety of our project.
Software
We conducted simulations for Listeriolysin O under two different pH levels within the lipid membrane of phagosomes in the lungs of human cells.
An in-silico protein model was also employed to validate the lower toxicity and sustained structural stability of both LLO and its mutated form.
Education
In the New Vision School secondary school, we explained asthma pathophysiology.
SDGs
We supported SDG 11 – Sustainable Cities and Communities to design a more convenient and healthier environment for asthmatic patients.
Week 28 (26th of July to 1st of August) Dry lab
Our team finalized the decision to incorporate a toxin-antitoxin system to enhance the biosafety and controllability of our probiotic platform.
Software
We awaited the LLO simulation results to confirm its stability at various PH levels.
Wet lab
Our wet lab team thoroughly discussed the proposed methodologies with experts, leading to the finalization of a comprehensive laboratory plan.
Education
Through our visit to STEM Gharbiya, we discussed the differences between current asthma medications and the pros and cons of various inhaler types.
SDGs
We pursued SDG 7 – Affordable and Clean Energy to lower energy pollution related to fossil fuel consumption, which is a main trigger for asthma.
August
Week 29 (2nd of August to 8th of August) Modelling
The double conditioning model for CO-BERA expression confirmed that the CO-BERA concentration exceeded 8 Micromole and PH below 7. This highly specific activation profile is ideal for our target population, suffering from severe uncontrolled asthma
SDGs
We contributed to SDG 15 – Life on Land to protect green landscapes, which provide clean air that reduces asthma prevalence worldwide.
Education
During our meeting with Public & Geriatrics, we learned more about the struggles that asthma patients face in their lives and learned the current treatments' limitations and side effects. Additionally, we introduced asthma to the public and heard their thoughts about it.
Week 30 (9th of August to 15th of August) Wet lab
Detailed lab protocols were drafted in collaboration with Dr. Atef and Dr. Abdullah, and team members received ongoing training in fundamental lab techniques.
Education
We visited many asthma patients across different high-risk asthma occupations such as farming, bakeries, brick factories, spinning and weaving factories, painters, cleaners, wood factories, pet shops, and sand quarries. Through these visits, we stressed wearing PPE and educated them about their occupations’ triggers.
Week 31 (16th of August to 22nd of August) Wet lab
All necessary fragments, materials, and primers required for the experiments were ordered from IDT and Benchling.
Education
We visited pharmacies, the main suppliers, to present our therapeutic approach & receive feedback to further tweek our project.The purpose was also to incrase public engagement further.
SDGs
We embraced SDG 4 – Quality Education to help children with asthma have a normal education in life with PRESS.
Week 32 (23rd of August to 29th of August) Education
We visited Future Blossoms Kindergarten, where we educated the children on human body structures and parts, and some life skills, such as teamwork and respect for nature.
SDGs
We aligned with SDG 5 – Gender Equality to relieve the burden on the shoulders of women, who are primary caregivers, allowing them to concentrate on their career growth.
Week 33 (30th of August to 5th of September) Wet lab
Initial lab procedures commenced with the resuspension of DNA fragments 1, 2, and 3, followed by their amplification using Polymerase Chain Reaction (PCR).
The size verification of all amplified fragments was performed via gel electrophoresis, after which these fragments were transformed into E. coli JM109 competent cells.
Education
At Future Leaders Primary School, we explained asthma, its symptoms, and lungs using models and gave an intro about DNA and microbes. Additionally, they developed critical thinking skills and learned how to solve problems.
Human Practice
Pharmaconex Event: selecting HPMC as the ideal DPI capsule, compared to alternatives.
September
Week 34 (6th of September to 12th of September) Wet lab
PCR amplification of Fragments 1 and 2 was repeated to achieve optimal yields. Concurrently, Fragment 3's PCR amplification was optimized to determine the ideal annealing temperature. Additionally, M13 primer variants were tested for functionality through PCR amplification of the 1-D5 plasmid.
Transformed bacteria were cryopreserved by creating glycerol stocks for long-term storage.
Education
In our visit to a local educational center, we briefly explained DNA, the central dogma, and genes. Not to forget, we detailed for them the scientific research method and cycle & the difference between observation and inference.
Week 35 (13th of September to 19th of September) Wet lab
Stored bacterial stocks were subsequently revived and cultured to expand pure populations containing the desired recombinant plasmid.
Both the target DNA fragments and the destination vector were isolated and then digested with compatible restriction enzymes. The digested fragments and vectors were subsequently ligated to form a Level 0 assembly, which was then transformed into E. coli JM109 competent cells.
Education
During our visit to Green Heights School, we discussed with students synthetic biology, gene expression, and siRNA. In addition, we introduced them to safety and research ethics.
Week 36 (20th of September to 26th of September) Wet lab
The transformation of the Level 0 ligated parts into E. coli JM109 competent cells was repeated due to initial unreliable results.
The assembled fragments were isolated via PCR amplification, and their correct size was verified by gel electrophoresis in preparation for the next assembly level.
Education
Through our Zoom meeting with ASU students, we talked about synthetic biology applications, plasmid design, and genetic circuits. Also, we explained to them the importance of dry lab precautions and ethical validation.
Week 37 (27th of September to 3rd of October) Wet lab
Following size verification, these ligated parts and the pJUMP23-1A plasmid were digested with the same restriction enzymes. A subsequent ligation reaction was performed, combining the digested components with ligase buffer, T4 DNA ligase, and ATP to create the Level 1 assembly. These ligated constructs were then purified and transformed into E. coli JM109 competent cells.
Education
By gathering with the public at a local society club, we were able to introduce the concept of using synthetic biology in therapeutics. Through our discussion session, we identified their doubts and corrected their misconceptions.
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