Abstract
What is Exocrine Pancreatic Insufficiency?
Exocrine pancreatic insufficiency (EPI) is a condition in which the pancreas fails to produce sufficient amounts of functional digestive enzymes, including lipases, proteases, and amylases[1]. These enzymes hydrolyze food particles, allowing for nutrient absorption by the intestines[2]. In the absence of these enzymes, food passes through the body undigested, resulting in malnutrition, nausea, and abdominal pain[3]. If left untreated, the chronic undernutrition caused by EPI eventually leads to osteoporosis and decreased immune function, and severe cases can result in death. Statistically, the two leading causes of EPI are chronic pancreatitis in adults and cystic fibrosis in children[4].
10-20% of the global population is affected by EPI[5]
If left untreated, EPI can cause severe malnutrition leading to...
Arrhythmia
Anemia
Osteoporosis
The Role of Enzymes in Digestion
Digestive enzymes are necessary the body to digest and process food into nutrients. Pancreatic enzymes are described in three main categories: lipolytic enzymes (lipase), amylolytic enzymes (amylase), and proteolytic enzymes (protease)[7]. These different groups of enzymes process different components of food, including fats, starches, and proteins.
Lipolytic enzymes, such as pancreatic lipase, digest dietary lipids for absorption in the small intestine[8]. Pancreatic lipase is the most crucial gastrointestinal enzyme involved in the hydrolysis of lipid ester bonds, resulting in the breakdown of lipids and ease of uptake by absorptive cells in the intestine[9]. A lipase deficiency or insufficient lipase activity can prevent the body from absorbing fat and fat-soluble vitamins (A, D, E, and K)[10].
In the presence of amphiphiles such as phosphates and bile salts (which are prevalent in the digestive system), lipase requires a cofactor to bind to and catabolize lipid macromolecule structures[11]. Colipase is a necessary protein cofactor that stabilizes the attachment of lipase to its triglyceride substrate in the presence of bile salts[12]. Colipase is initially secreted as its precursor, procolipase, which is then cleaved in the intestine at the Arg5-Gly6 bond by the enzyme trypsin, a serine protease secreted by pancreatic acinar cells[13,14]. After procolipase is transformed into colipase, the lipase-colipase enzyme complex is formed and can then digest fats.
Amylolytic enzymes, such as pancreatic amylase, catalyze the hydrolysis of consumed starches (long-chain carbohydrates) into sugar, which is used as an energy source[10]. Insufficient amylase in the body can lead to diarrhea from undigested carbohydrates.
Proteolytic enzymes, such as protease, hydrolyze proteins into their constituent parts, including peptides and amino acids, which can be absorbed by cells in the intestine[15]. Proteases also protect the body from harmful yeast and bacteria in the intestines[16]. Undigested proteins can lead to allergic reactions or stomach pain in individuals.
Why Are Current Solutions Not Enough?
Currently, the standard treatment for EPI is pancreatic enzyme replacement therapy (PERT) with pancrelipase or other oral drugs containing the digestive enzymes pancreatic lipase, amylase, and protease16. These enzymes supplement those deficient in the body naturally, and thus permit the breakdown and digestion of food into nutrients. However, these medications are currently derived from porcine pancreases and are thus unsuitable for those who avoid pork for religious or dietary reasons[17]. Animal derived pharmaceuticals also have other ethical and environmental dilemmas. The extraction of these enzymes from pigs is often associated with inhumane treatment of pigs (poor feeding/living conditions and eventual slaughter) and large amounts of waste[18]. Each farmed pig consumes many resources and produces over 10 pounds of manure per day, leading to air and groundwater pollution[19]. In China, pig production accounts for roughly 42% of the nitrogen and 90% of the phosphorus flowing into the South China Sea. Additionally, while the possibility of harmful disease transmission via animal-derived products is low, porcine pancreatic enzymes are theorized to carry a pig virus, analogous to mad cow disease[20]. There are currently no FDA-approved, synthetically-produced PERT medications, highlighting the urgent need for a more sustainable and accessible treatment for EPI.
Our Solution
Our goal is to develop a probiotic that produces pancreatic lipases, procolipases, amylases, and proteases using engineered Bacillus subtilis. Probiotics are live microorganisms, such as bacteria or yeast, that have beneficial effects on the body when ingested. The benefits include boosting the immune system, producing nutrients, or preventing pathogens from colonizing the digestive track[21,22].
Scientists have successfully engineered the P. pastoris GS115 yeast and E. coli DH5α competent cells to produce the necessary digestive enzymes[23,24]. While both organisms are considered to be recombinant protein “workhorses”, the implementation of these specific strains into probiotics has yet to be extensively studied[25,26]. Disarmed strains of these organisms have already been used for their probiotic properties, including E. coli Nissle 1917 and P. pastoris X-33, but have not been proven in literature to have protective effects against pancreatic inflammation[27].
B. subtilis is a probiotic found naturally in the human gut and is often used in fermented foods such as yogurt. B. subtilis is beneficial for reducing pancreatitis-associated complications, including allergic responses and poor intestinal barrier integrity[30]. Additionally, B. subtilis naturally produces amylase and protease, making our engineered version that also produces pancreatic lipase and colipase a convenient, all-in-one option that replaces pancrelipase[31,32]. Our solution also embeds a glucose-switch mechanism, allowing the enzymes to be synthesized and naturally secreted once food (glucose) is consumed and detected by the device. Though B. subtilis CFR5 has the greatest benefits, we have chosen to establish a proof-of-concept in the more easily-obtainable B. subtilis 168, which posseses the same benefits with lower efficiency.
Looking ahead, we will place our probiotic in yogurt so it can be safely (and deliciously!) consumed as a probiotic. Our device will thus serve as a more accessible option for those who prefer a treatment free from porcine-derived enzymes.
Our Inspiration
While brainstorming project ideas, our team identified a lack of inclusive treatment options available for EPI patients. Existing FDA-approved medications, derived from porcine pancreas, present challenges for individuals with constraints against consuming pig products. Also, these pancreatic enzyme replacement therapies must be taken with every meal or snack, complicating daily management for the patients.
We are also concerned about the ethical and environmental implications associated with the use of porcine enzymes in pharmaceuticals. The inhumane treatment of pigs and environmental impact of pig farming (resource consumption and pollution) prompted us to seek a more sustainable solution.
Recognizing these shortcomings, we propose a synthetic alternative to porcine pancreatic enzymes—a solution that still produces the necessary enzymes, is safe to consume, and ameliorates the condition of the human gut biome. This innovation sets a new standard for inclusivity and safety, paving the way for future advancements in healthcare.
Ultimately, BLISS will revolutionize EPI treatment, offering patients a solution that aligns with their values and makes managing their symptoms a blissful experience.
References
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