Overview

Our project, IRIS, owes its current form to expert guidance.

During our brainstorming, designing and lab work days, we were fortunate to collaborate with experts of various fields. People from Greece, the United Kingdom and Stockholm, from several different fields, such as Biology, Chemistry and Medicine, professors, doctoral students and independent researchers, contributed to IRIS’ creation.

We would like to express our gratitude by briefly describing what each of them did for us.

Our Integrated Human Practices can be splitted in three categories: The ones that happened during our brainstorming sessions and helped us reach to the main idea of our project, the ones that helped us out while conducting our experiments in the lab and finally the ones that gave us advice on the safe implementation of IRIS in the real world.

Step 1: Where it all began

iGEM Ioannina 2021

Mr Kolletas, Mrs Eugenia Roupakia and Mrs Simone with members of iGEM Ioannina wet lab

Photodynamic Therapy

Step 2: Lab advice

Tarvi Teder- Low copy-number plasmids and Drug Delivery Systems

Tarvi Teder, a PostDoctoral researcher in Biophysics at Karolinska Institutet, provided us with precious advice on how to isolate low-copy plasmids, as we encountered issues while isolating pBBR1-MCS2. After explaining our problem, he was confident that our failed attempts of isolating the plasmid had something to do with the plasmid’s antibiotic resistance and advised us on checking if we indeed had the right plasmid to start with. His advice was precious and led us to start a troubleshooting procedure for the isolation of pBBR1-MCS2.

You can find the troubleshooting procedure HERE

He also found the direct injection of active bacteria in the human body risky as directly injecting bacteria in a person’s body can trigger endogenous immune responses. The bacteria are likely to be recognized as threats, and that could lead to immune activation (leukocytes, such as neutrophils and macrophages are recruited to the site of infection to destroy the bacteria) or an inflammatory response (swelling, redness, heat and pain) at the injection site.

A possible solution to that issue would be the utilization of a drug delivery system. Liposomes, for example, can be used to deliver IRIS directly to the target tumor site, where a triggered release of the bacteria could take place. In that way, the bacteria are not going to directly interact with the patient and possible immune responses can be avoided.

Step 3: Iris as a cancer treatment

Barry Campbell- targeted therapy

Barry Cambell, professor in Infection Biology and Microbiomes at University of Liverpool, had a conversation with one of our members about the role of adhesive proteins and how we can incorporate them to our bacteria for a more targeted and safe therapy. Adhesive proteins can bind to glycosylated receptors that exist specifically in cancer cell membranes. This binding would lead to the closer secretion of our bacteria’s factors directly into the cancer cell, and would therefore minimize the possibilities of untargeted secretions.

He also suggested possible methods that can be employed for inserting bacteria into a cancer cell, in order to minimize the loss of light emission and important molecules in the intercellular space and therefore increase the project's effectiveness. His suggestions are valuable for the future development of IRIS.

Department of Medicine × iGEM Ioannina

Mr Kolletas, Mrs Eugenia Roupakia and Mrs Simone with members of iGEM Ioannina wet lab

In September, we had an insightful conversation with Mr Kolletas, Biology professor at the Department of Medicine of University of Ioannina, Mrs Eugenia Roupakia, Doctoral Student in Molecular Cellular Biology at the Department of Medicine of University of Ioannina, and Mrs Simone Besta, PhD candidate in Molecular Cellular Biology at School of Medicine of University of Ioannina.

We presented our project to them and we had a big conversation about the safety of our project. They agreed that an effective way to a more targeted therapy is including adhesive proteins and even suggested several other molecules that we can employ to increase the target specificity of our mechanism.

We also talked about possible kill-switch mechanisms to further increase our project’s safety. Mr. Kolletas, specifically, had many suggestions about this topic, including a kill switch mechanism inducible by our bacteria population, or by an σ³⁸ rpoS factor, which regulates gene expression in E. coli.

Another outcome from that meeting was the highlighting of the importance of knowing methoxy e-coelenterazine’s half-life, as it would better regulate our therapy and its future implementation. The idea of co-culturing our engineered bacteria with breast cancer cells to test the proof of our concept seemed to puzzle Mr. Kolletas, who noted that some subtypes of breast cancer have mechanisms for countering oxidative stress. He proposed that we possibly need to make our therapy more specialized to a certain type of solid tumor cell line to achieve better results.

Mrs Kallergi G. - stages of drug development

Mrs Kallergi, and members of iGEM Ioannina wet lab

In October, we had a meeting with Mrs Kallergi, Assistant Biochemistry Professor at the University of Patras, a researcher at the European Liquid Biopsy Society (ELBS) and a former supervisor of the team iGEM Patras Med 2022. Due to her experience with iGEM and her expertise in the field of liquid biopsy for cancer diagnostics, we were happy to engage in a conversation with her.

Commenting on our idea to co-culture the engineered bacteria with cancer cells, she suggested the possible co-culture of our transformed bacteria with healthy human cell lines in order to test possible side-effects of our project.

She explained to us the whole process of drug approval and she mentioned the significance of a precise and clear step-by-step plan for the possible in vivo application of our project in the real world. A logical workflow of in vivo experiments would start by testing on cancer cell cultures, then move on to animal models and after many stages of approvals our experiments could finally continue on clinical trials on humans.

All these steps should be taken very carefully in order to minimize possible risks and ensure that possible positive effects are indeed our project’s effect, rather than an artifact. Finally, she agreed that our therapy must be very well regulated in order to avoid immune responses. Especially with cancer patients who have already received treatment and are under immunosuppression, the possibility of risks occurring is greater, and for that reason, she noted that we should consider even more parameters that would make our therapeutic approach safe.

Step 4: Mentorship Program

A fourth, special category dedicated to our mentor, as she contributed to all the categories above. During our journey, we were lucky to have an amazing mentor and friend by our side, Sofia Oikonomou.

Sofia Oikonomou

Sofia, currently a master’s student at University of Copenhagen, a graduate from the Department of Biology, University of Patras, a former member of the Enzyme and Synthetic Biotechnology Laboratory at the National Hellenic Research Foundation and a team member of iGEM Athens 2022, has all the necessary knowledge and experience to help us improve our project.