Celiac is an autoimmune disease where the immune system mistakenly attacks and damages the small intestine after ingesting foods containing gluten. This immune response causes damage and inflammation to the villi, impairing the absorption of nutrients, which can result in a large range of symptoms, such as diarrhea, malnutrition, weight loss, and fatigue.

Our first step to approaching this problem is through researching and developing the background knowledge we would need to truly understand our topic. We knew that genetic editing is a very complex and advanced field, and a deep understanding is required to implicate it in our project properly. After studying celiac thouroughly, we were able to formulate our main research questions.

1. What are the best and most efficient ways to treat Celiac? 

2. How can implement these methods using CRISPR?

3. How do we insure that these methods are safe?

CELIAC

As mentioned in the beginning, celiac disease is an autoimmune disease in which the immune system damages and inflames the villi when gluten, especially gliadin, is ingested. The villi are hair-like structures inside the intestine that absorb nutrients into the bloodstream, and when impaired by diseases like celiac, they prevent the body from taking in the nutrients from food.

 

People with celiac will often have the genetic markers, HLA-DQ2 and HLA-DQ8. However in some cases, people with these genetic markers may not have celiac, and people that are diagnosed with celiac may not have these markers either. When the gliadin peptides in gluten break down in the digestive system, they will bind to the HLA-DQ2 or HLA-DQ8 molecules on the surface of antigen-presenting cells, which then triggers the activation of T-cells in the immune system, causing an inflammatory response in the small intestine, damaging the villi.

What is HLA-DQ2 and HLA-DQ8?

HLA-DQ2 and HLA-DQ8 are genetic markers strongly associated with celiac disease, as their presence significantly increases the risk of developing the condition, though it doesn't guarantee it. They are human leukocyte antigens; proteins, or markers, on the surface of most cells in the body that the immune system uses to differentiate between cells and foreign invaders. They can bind to gluten proteins.

CRISPR

Gene editing is a group of technologies that allow scientists to change an organism's DNA. CRISPR also known as clustered regularly interspaced short palindromic repeats, a more recent type of gene editing has been developed to be faster, cheaper, more accurate, and more efficient than other types of genetic editing methods. There are 3 classes of CRISPR, each with 6 types and many other subtypes. It is capable of performing gene insertion, deletion, and base editing in DNA or RNA. However more advancements in gene editing have shown more types and subtypes exist.

Background research: Celiac Genome Editing Research

There are 3 main methods that will be used to approach this problem.

1. Liver Regeneration

2. Editing Gluten

3. Notch Signaling

Aditionally, research on CRISPR will be linked below.
CRISPR Genome Editing

This is a research-based project.

Throughout our research, we have found three ways that celiac could be potentially treated in the future through the genome editing tool, CRISPR. CRISPR (clustered regularly interspaced short palindromic repeats) is a gene editing technology that is from the e-coli bacteria. It is considered faster and more inexpensive compared to old methods used such as ZFNs (zinc finger nucleases) or TALENs (transcription activator-like effector nucleases).
In liver regeneration, the idea at the beginning of research was to edit the hepatocytes. However, it will prove to create too many challenges, even if there are solutions to these problems. Instead, we could genetically modify the intestinal epithelial cells that form the protective layer on the villi to replicate the hepatocyte's proliferative abilities, allowing for an increased rate of stem and epithelial cell regeneration. By using CRISPRa to overexpress certain genes to activate signaling pathways such as Wnt and Notch, this approach could potentially strengthen the barrier to protect the villi, and treat celiac. Fortunately, this solution is much more applicable than the original idea, but it may not have a large enough influence to make an impact on the intestine epithelial layer. In the future, after going through rigorous experiments, this approach could be used as a "backup", and would be incredibly useful if used alongside other treatments. 
Throughout editing gluten, the large problem we faced was that editing out a-gliadin would completely change the molecular composition of gluten itself. The two ideas that we found were strengthening the existing gliadins (b,y, & w) and finding a replacement for a-gliadin (the main storage protein in gluten). Strengthening would be through CRISPRa implementing crosslinking which would strengthen the gluten network and disulfide bonds which would allow for the glutenins to interact more. Replacements we found were from the main storage proteins of rice, soy, pea, and whey, they would be knocked out using the most well-known type of CRISPR, Cas9. Editing gluten is also one of our most effective solutions, as we are directly editing the source of the problem.
Notch signaling was our last idea, the problem we faced with notch was a rather severe one; the risk of cancer, tumor formation, or unwanted cell growth. However, by inhibiting notch signaling, it would allow for the t-cells to become Tregs (regulatory t-cells) instead of pro-inflammatory t-cells (such as th1 and th17 the cause of the immune reaction). The second way to implement notch signaling would be to over express certain genes through CRISPRa for gut protection and to increase the number of Tregs in the system. These genes are RBPJ (recombination signal binding protein for immunoglobulin kappa J region) and HES1 & HES5 (hairy and enhancer of split-1 & 5). This approach would be incredibly helpful to strengthen the epithelial cells of the small intestinal villi. 

How will we make sure that CRISPR is safe? Gene editing has as many risks as there are lifesaving oppertunites, how we insure that these treatments are completely safe will be through trials, similar to the ones done today for cancer and sickle cell disease, regular screening and close monitoring. Another way we are able to make sure treatments and solutions are completely safe, is by using gene editing, gene editing is very versatile, it could allow us to correct potential probelms before risks such as unwanted cell growth develop.

References

(n.d.). Gliadins from wheat grain: an overview, from primary structure to nanostructures of aggregates. Retrieved March 5, 2025, from https://pmc.ncbi.nlm.nih.gov/articles/PMC5899726/

(2020, April 24). CRISPR/Cas9 Gene Editing of Gluten in Wheat to Reduce Gluten Content and Exposure—Reviewing Methods to Screen for Coeliac Safety. Retrieved March 5, 2025, from https://pmc.ncbi.nlm.nih.gov/articles/PMC7193451/

(2022, November 22). Insights on Chemical Crosslinking Strategies for Proteins. Retrieved March 5, 2025, from https://pmc.ncbi.nlm.nih.gov/articles/PMC9738610/

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We would first like to thank our science fair coordinators, Ms. Rheinstein and Mr. Lahoda for giving us this opportunity to participate in this year's science fair. We would also like to offer our genuine gratitude for Dr. Pierre Billon who helped us answer a few of our questions that helped steer us in the right direction for our project. A special thanks for Jimmy Xu, who played as an actor in our video! We also acknowledge that we are situated on the Treaty 7 region of Alberta. Lastly, a huge thanks to our family and friends who motivated and supported us!