The results of a study in mouse models by scientists at the University of Geneva (UNIGE) have provided proof of principle that a protein known as S100A9 can significantly improve metabolism in insulin deficiency (ID), and may point to the future development of new treatments for diabetes. The researchers have for years been working on an alternative to insulin therapy, based on the S100A9 protein. They say that by deciphering biological mechanisms at work, their research has also identified a previously unknown anti-inflammatory effect that could prove key well beyond diabetes.

The newly reported study, headed by Roberto Coppari, PhD, a professor in the Department of Cell Physiology and Metabolism and Coordinator of the Diabetes Centre of UNIGE Faculty of Medicine, is described in a paper in Nature Communications, titled “Hepatic non-parenchymal S100A9-TLR4-mTORC1 axis normalizes diabetic ketogenesis,” in which the team concluded “Thus, S100A9 is a realistic next-generation biological agent for treatment of ID, a disorder that is at present pandemic.”

Patients with a severe form of diabetes in which pancreatic beta cells produce insufficient, or even no insulin have must inject themselves regularly with artificial insulin to survive. Insulin therapy, which celebrated its 100th anniversary in 2021, has probably saved the lives of hundreds of millions of people with either type 1 diabetes (T1D) or with severe forms of type 2 diabetes (T2D). However, insulin therapy is not without risks if the doses are too high or too low, and in the long term, it can also lead to serious metabolic and cardiovascular problems, and even directly be responsible for some potentially fatal conditions.

Consequently, the life expectancy of individuals with insulin-dependent diabetes is reduced by about 10–15 years compared with the norm. “Life-threatening hypoglycaemia, negative impact on fat metabolism and increased cholesterol: these are some severe side effects of insulin,” Coppari said. “This is why we are looking to develop complementary or alternative treatments that are more effective and less dangerous.”

Despite continuing advances in treatment for insulin deficiency, current methods are “suboptimal” in achieving adequate metabolic control in patients with diabetes, the team further pointed out. “This shortcoming might be due to the fact that the current approach to diabetes treatment is chiefly glucose-centric, with virtually all available, and investigational, adjunctive therapies aiming at improving hyperglycemia.” There is, the scientists noted, “the need to extend treatment approaches beyond correcting hyperglycemia.”

In diabetic people, insulin deficiency can cause a sudden increase in ketones and acidification of the blood, a mechanism called diabetic ketoacidosis. This kind of pathogenic ketogenesis can be fatal. “… diabetic hyperketonemia develops when circulating insulin levels are insufficient to suppress ketogenesis hence leading to uncontrolled ketone body production,” the investigators explained in their paper. “If severe, this defect brings about diabetic ketoacidosis (DKA; that is blood acidification owing to the low acid dissociation constant of ketone bodies) that can be fatal; therefore, DKA needs emergency medical attention.”

In 2019, Professor Coppari’s team identified a protein called S100A9 that regulates blood glucose, lipids and ketones (a product of fatty acidic oxidation in the liver when the body no longer has enough glucose to function), without the side effects of insulin. “To develop a drug, however, we had to understand how this protein works precisely and demonstrate its effectiveness in animal models,” explained Girorgio Ramadori, PhD, a research associate in Professor Coppari’s lab and lead author of this study.

The team first set out to decipher the mode of action of S100A9 in mouse models of diabetes. “It turns out that this protein acts in the liver,” said Gloria Ursino, PhD, a first author of the study and post-doctoral fellow in the research team. “It activates the TLR4 receptor, which is located on the membrane of certain cells, but not on the hepatocytes, which are the main functional cells of the liver.” This finding is great news from a pharmacological point of view, because it means that S100A9 does not need to enter the liver cells to act and should allow for a simple injection mode of administration.

Diabetic ketoacidosis is life-threatening emergency that affects 2–4% of people with type 1 diabetes every year. “TLR4 activation in the liver controls the production of ketones,” explained Ursino. “But this activation process does not trigger inflammation, whereas TLR4 is usually pro-inflammatory. The S100A9-TLR4 dialogue therefore seems to act as a totally unexpected anti-inflammatory drug.” The authors further explained, “Mechanistically, S100A9 acts extracellularly to activate the mechanistic target of rapamycin complex 1 (mTORC1) in a TLR4-dependent manner … Therapeutically, recombinant S100A9 administration restrains ketogenesis and improves hyperglycemia without causing hypoglycemia in diabetic mice.”

The scientists in addition examined blood of diabetic people arriving at the emergency room with severe insulin deficiency. “A slight but insufficient natural increase in S100A9 is detected,” said Ramadori. “Therefore, additional administration of S100A9 is expected to enhance this compensatory mechanism.” As the team noted in the published paper, “… circulating S100A9 in patients with ketoacidosis is only marginally increased hence unveiling a window of opportunity to pharmacologically augment S100A9 for preventing unrestrained ketogenesis.”

In their discussion, the authors concluded, “In this study, by generating and assessing diverse genetically engineered animal models of diabetic hyperketonemia we determined the site (i.e., liver) and key molecular and cellular components (i.e., TLR4 and mTORC1 in non-parenchymal hepatic cells) underlying the hyperketonemia-normalizing effect of S100A9. Furthermore, we show pre-clinical and observational clinical data indicating that recombinant S100A9 is a putative investigational drug displaying safety and efficacy profiles owning therapeutic potential.’

While the idea of a combination of drugs has already been explored for diabetes therapy, previous research has focused on strategies that would increase insulin sensitivity. “But this only leads to the same results with lower doses,” commented Coppari. “The side effects of insulin therapy remain the same. Here, we propose a radically different strategy with a drug that works independently of insulin and that can neither trigger hypoglycaemia nor disrupt fat metabolism.”

The scientists will initially aim to test their drug in conjunction with low doses of insulin, but they do not rule out the possibility of administering the S100A9 protein alone in the future, in specific conditions. To further develop the treatment Coppari and Ramadori have created a start-up company, Diatheris, supported by UNITEC, the UNIGE technology transfer office, and FONGIT, the main foundation supporting technological entrepreneurship in the canton of Geneva.

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