Engineering Escherichia coli to cure IBD in comprehensive ways

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Our project aims to designs an engineered type of Escherichia coli which could perform multiple functions to alleviate intestinal inflammation and repress hazardous bacteria in intestine, in the hope that our experiments in mice could suggest its potential for treatment in human bodies. After careful comparison and evaluation, we have planned to utilize Escherichia coli as our primary research organism, based on previous studies indicating Escherichia coli's higher colonization capability in intestine compared to yeast.

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Background

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Our team is dedicated to addressing the challenges posed by inflammatory bowel diseases (IBD), affecting millions worldwide. These diseases are characterized by chronic inflammation of the digestive tract, significantly impacting patients' quality of life. Current treatments often target at symptoms rather than underlying causes and aim at only one of the factors causing this disease. The lack of universality in medical treatments across different patient types remains a challenge for scientists and a source of suffering for patients, which motivates our team to seek novel and comprehensive therapeutic strategies.

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Project Goals

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In this competition, we aim to optimize engineered Escherichia coli strains capable of interacting with gut microorganism, attaching to the intestinal surface, and restraining harmful bacteria. Our primary objectives include enhancing probiotic colonization in the intestines and utilizing anti-inflammatory mechanisms to alleviate intestinal inflammation.

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Project Inspiration

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We selected IBD as our study model because it is one of the most commonly observed diseases, and everyone around us, or even we ourselves, could potentially be victims. around us and even we ourselves could be potential victims of. Even if it is cured once, patients are still faced with high risks of its recurrence. We conceive that the tortured victims of IBD could be better saved from pain using the constantly developing and maturing biological tool of genetic engineering. Additionally, academic knowledge from microorganism and biomaterial courses provided us with new perspectives that our engineered E.coli could possibly express a certain kind of bioactive material adhesive to intestine surface, so that it could stay there longer for treatment. We were also inspired by the cocktail therapy initially for AIDS and extended this concept, believing that our engineered E.coli could perform various functions in treating IBD, targeted at different factors causing the inflammation, in other words, one bacterium serving as multiple sorts of medicine.

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Methodology

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We plan to design a plasmid consisting of different functioning components, including a ROS-responsive promoter and other sequences which encode anti-inflammatory and adhesion factors or materials. Nevertheless, before that we have to determine the best of these factors by in vitro and in vivo experiments. For the in vitro part, we have used macrophages to stimulate the intestinal microenvironment because studies have indicated that one of the main causes of inflammation in IBD is the response of these cells in the face of pathogenic bacteria. We induced macrophages into an inflammatory state using S.Tm, added different kinds of polypeptides and evaluated their performances in various dimensions, including cell survivals, the number of intracellular bacteria and ROS level. The results provide a preliminary reflection of the curative capabilities of all these factors. We then plan to test them in IBD-affected mice. Those in vivo experiments include creating the IBD model in mice, delivering of the anti-inflammatory factors, and testing indicators such as Disease Activity Index level, colon length, bacterial diversity and the transcription level of inflammatory factors in stool, etc. PCR-based and bioinformatics methods are expected to be largely used in our research. After seeking out all the best factors and figuring out the sequences encoding them, we will assemble all the elements into a plasmid and deliver it into E.coli, which is expected to be a effective medicine combating IBD.

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Innovative Strategy

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Our innovative engineered E.coli has a huge difference with the traditional IBD medicine, given that it is organisms-based, responsive and interactive, performing more than one functions by combing multiple functioning elements to achieve ideal goals.

To avoid the side effects of overexpressing anti-inflammatory factors, we will design a responsive promoter in the plasmid to ensure that our medicine provides stable, appropriate curative effects.We also designed a safety plasmid to ensure that the bacteria undergo self-destruction if they fail to function.

Furthermore, we would like to introduce an artificially designed adhesion protein to our engineered bacteria, extending its functioning time and targeted efficiency.The design of this adhesion factor is based on a glucan-binding domain, which uses its glucan-binding properties to act as a bridge between the intestine and engineering bacteria.

We also made particularly interesting modifications to the anti-inflammatory factors by using a linker to connect two bee venom peptide monomers, forming a hairpin-like structure that reduces cytotoxicity and enhances efficacy.