How biotechnologies facilitate the production of insulin

While the biotech industry has seen enormous growth and diversification, even more because of the pandemic, the largest corporations in the industry still make most of their profits through the manufacturing of a single, small protein, insulin. More than 5% and an ever increasing amount of the world population is dependent on the availability of this pharmaceutical drug, and we found it important to mention its significance. As it is a product of biotechnology, we iGEM teams think highly of what has been achieved with the manufacturing of insulin, but we also here intend to highlight current problems with how insulin is currently manufactured, and some promising solutions. 

What is insulin? 

Insulin is a hormone that regulates glucose, lipid, and protein metabolism. It is called a “storage hormone” because glycolysis leads to storage of energy as triglycerides.

This hormone is produced by the pancreas, a little organ behind the stomach. Pancreas is composed of specialized areas called islets of Langerhans: these are made up of different types of cells, as α and β-cells, that synthesize, store and release insulin. 

This is a polypeptide structure; thus, it is produced by transcription and translation. First, insulin mRNA is translated into preproinsulin. Then, when its signal peptide is removed during insertion into the endoplasmic reticulum, it generates proinsulin. Lastly, proinsulin is exposed to many endopeptidases in the endoplasmic reticulum to form the mature insulin.

Insulin is an antagonist of glucagon, the “reserve mobilization hormone”, also produced by the pancreas. As insulin is produced by β-cells of pancreas, glucagon is produced by α-cells.

Insulin is secreted into the sanguine circulation when the glycemia (i.e concentration of sugar in the blood) is high, for example after a meal.    
Both insulin and glucagon bind to G protein-coupled receptors. Signal transduction leads to the activation of phosphatase proteins that dephosphorylate other proteins, causing their activation or inhibition. It results in :

• An augmentation of glycolysis (catabolism of glucose) and a diminution of gluconeogenesis (generation of glucose from non-carbohydrates substrates) by the activation of two enzymes:  phosphofructokinase-2 and pyruvate kinase,  and by the inactivation of fructose bisphosphatase enzyme.

The activation of glycolysis allows the conversion of pyruvate into acetyl-CoA. A part of acetyl-CoA is used for energy generation by Krebs cycle. The remaining part is stored in the form of glycogen in muscle and liver or used to produce triglycerides that will be stored into adipose tissue.

• This storage of glycogen is allowed by an augmentation of Glycogenesis by the activation of glycogen synthase, allowing the polymerization of Glucose into Glycogen and a diminution of glycogenolysis by the inactivation of phosphorylase kinase, the enzyme that activates glycogen phosphorylase which catabolize glycogen polymer.

• This production of triglycerides is permitted by the fact that insulin causes an augmentation of fatty acids synthesis and a diminution of lipolysis

After the insulin acts on its receptor site, it may be released back into the extracellular environment, or it may be degraded by the cell.

We will now speak about diabetes, which is a disease that requires most of the time the administration of synthetic insulin.

Diabetes: disease caused due to insulin deficiency 

In January 2020, about 415 million people worldwide are living with diabetes.               37.3 million Americans – about 1 in 10 – have diabetes and 96 million American adults – more than 1 in 3 – have prediabetes (i.e an hyperglycemia but not sufficient to diagnose a diabete).
There are two types of diabetes :

•  In type 1 diabetes, lymphocytes T identify 𝛽-pancreas cells as foreign and attack them, leading to a lack of insulin production. This diabetes is lethal if the patient doesn’t take insulin every day. Indeed, untreated hyperglycemia for long periods of time may damage nerves, blood vessels, tissues and organs. Damage to blood vessels can increase the risk of heart attack and stroke, and nerve damage may also lead to eye damage, kidney damage and non-healing wounds.

•  In type 2 diabetes, the body is unable to make enough insulin or the body doesn’t use it well. This diabetes can be prevented by having a healthy life : it can be managed by medication, exercise, diet but also prescription of insulin.

What are the current solutions to produce insulin?

Already since the 1980s, Escherischa Coli have been genetically modified to produce insulin for human use. This makes it the first protein to be produced with a pharmaceutical purpose, made thanks to genetic engineering. How this is made can be described by the figure below. 

What is problematic, is that insulin that we need, is not just in one gene, but rather the gene will have the cell create the non-active proinsulin. In order to solve this problem, there was a need to further modify the human gene to only include only the needed parts. Having successfully transformed the bacteria and having them produce insulin, the insulin is then harvested from the cells and purified. 

How biotechnology will improve the production of insulin

This described process has remained similar until this day, but diabetes is affecting more and more people, meaning that the production has to scale up drastically to keep up with demands. Sadly scaling up is not always straightforward, as biological processes are not as predictable as, for example, chemical ones. This means that new processes need to be developed to ensure the survival of diabetics around the world. Some promising methods are listed below and described in short.

1. Making E.Coli a more stable expression host.
   When modifying E.coli , the plasmid usually remains not more than for 100 generations until it is either lost from the organism or the plasmid function is altered. A solution to this problem is to engineer a dependency on the created product, explained in greater detail in the article of Rugbjerg P et al. Synthetic addiction extends the productive life time of engineered Escherichia coli populations. 
   
2. Using a different expression host that is more stable. 
   Different expression hosts such as even plants are suggested by Baeshen, N. A et al. in Cell factories for insulin production. Plants are there said to have the potential of high yields and being cost effective, while also producing the insulin in more stable containers, such as the seeds and leaves of the plant
   
3. Revitalizing the body’s own ways of producing insulin through gene therapy.
   There are hopes to restore the functions of damaged or destroyed islets of Langerhans, or create new insulin-creating cells through editing the human genome. Possibilities of such therapies and their current limitations are discussed by Chellappan DK et al. in Gene therapy and type 1 diabetes mellitus , who for the moment concludes that still more research is needed. However, in future we might see the complete eradication of diabetes type 1 through methods like these. 

Conclusion

This concludes this week’s blog post and we hope you’ve again learnt something new and exciting, or gained appreciation for what amazing ideas are currently being researched! 

Sources

Reiff, N. R. (2020, June 30). 10 biggest biotechnology companies. Investopedia. Retrieved August 30, 2022, from https://www.investopedia.com/articles/markets/122215/worlds-top-10-biotechnology-companies-jnj-rogvx.asp

A.M. (2019, June 4). Insulin synthesis. News-Medical.Net. Retrieved August 29, 2022, from https://www.news-medical.net/health/Insulin-Synthesis.aspx#:~:text=Insulin%20is%20synthesized%20in%20significant,acid%20chains%20or%20polypeptide%20chains

Centers for Disease Control and Prevention. (2020, January 2). CDC Global Health – Infographics – World Diabetes Day. Retrieved August 29, 2022, from https://www.cdc.gov/globalhealth/infographics/diabetes/world-diabetes-day.html

Centers for Disease Control and Prevention. (2022, January 24). The Facts, Stats, and Impacts of Diabetes. Retrieved August 29, 2022, from https://www.cdc.gov/diabetes/library/spotlights/diabetes-facts-stats.html

Inserm. (2017, July 11). Diabète de type 1 ⋅ Inserm, La science pour la santé. Retrieved August 28, 2022, from https://www.inserm.fr/dossier/diabete-type-1/

P.S. (2021). Aspects biochimiques du métabolisme. Pascal Schneider.

S.W. (2021, March 7). How insulin works. Webmd.Com. Retrieved August 29, 2022, from https://www.webmd.com/diabetes/insulin-explained#:~:text=High%20blood%20sugar%20stimulates%20clusters,more%20insulin%20your%20pancreas%20releases

The cell biology of systemic insulin function. (2018, July). Researchgate.net. Retrieved August 28, 2022, from https://www.researchgate.net/figure/Insulin-biosynthesis-and-secretion-A-Insulin-maturation-along-the-granule-secretory_fig2_324253965

Baeshen, N. A., Baeshen, M. N., Sheikh, A., Bora, R. S., Ahmed, M. M., Ramadan, H. A., Saini, K. S., & Redwan, E. M. (2014, October 2). Cell factories for insulin production. Pubmed. Retrieved August 30, 2022, from https://pubmed.ncbi.nlm.nih.gov/25270715/ 

Rugbjerg P, Sarup-Lytzen K, Nagy M, Sommer MOA. Synthetic addiction extends the productive life time of engineered Escherichia coli populations. Proc Natl Acad Sci U S A. 2018 Mar 6;115(10):2347-2352. doi: 10.1073/pnas.1718622115. Epub 2018 Feb 20. PMID: 29463739; PMCID: PMC5877936. from https://pubmed.ncbi.nlm.nih.gov/29463739/ 

Chellappan DK, Sivam NS, Teoh KX, Leong WP, Fui TZ, Chooi K, Khoo N, Yi FJ, Chellian J, Cheng LL, Dahiya R, Gupta G, Singhvi G, Nammi S, Hansbro PM, Dua K. Gene therapy and type 1 diabetes mellitus. Biomed Pharmacother. 2018 Dec; From https://pubmed.ncbi.nlm.nih.gov/30372820/