A Welbio team from the de Duve Institute has just published the results of a long-term research project that explains two congenital metabolic diseases characterised by a defect in neutrophils (white blood cells), and considers a new treatment.
Our cells are home to an impressive series of chemical reactions – intermediary metabolism – that take place mainly through the presence of enzyme catalysts. A precise set of reactions – a metabolic pathway – thus makes it possible to transform one molecule into another, to convert food into useful compounds. A well-known example of the metabolic pathway is glycolysis, a set of ten different enzymatic reactions that can extract the energy our body needs from glucose (sugar).
For most of us, this metabolism works well, but the machine can seize up. This is the case when a gene encoding an enzyme that intervenes in these reactions has mutated. The production line is then interrupted (the miscoded enzyme is inactive). Upstream from this weak link, substances – waste – accumulate, while downstream, there is a deficit in the final substance. These diseases, or innate errors of metabolism, as they are called, are the research focus of Maria Veiga da Cunha and Guido Bommer, FNRS research associates at the de Duve Institute, as well as Emile Van Schaftingen, an emeritus professor in the UCLouvain Faculty of Medicine, who works in the same institute. This research allowed them to explain in PNAS two congenital neutropenia diseases, which are characterised by defects in neutrophils, our main white blood cells, and to propose a possible treatment. Their results have recently been published in PNAS.
A longstanding tradition
This achievement is part of a longstanding research tradition in which Christian de Duve himself played an important role. One of these metabolic diseases, glycogenosis type 1, was described in the 1940s by two American researchers, Carl and Gerty Cori, joint winners of the 1947 Nobel Prize in Medicine. They showed that the disease is caused by a deficiency in a crucial enzyme in our metabolism that is responsible for the production of glucose: glucose-6-phosphatase (G6PC1). It is in a Université de Saint-Louis Bruxelles laboratory that a young researcher, Christian de Duve – who also won a Nobel in 1974 – pursued a postdoctorate and was eager to understand how insulin affects glucose metabolism.
Once back in Leuven, Christian de Duve and his young collaborator Géry Hers – who later became Emile Van Schaftingen’s boss – showed that the crucial enzyme is associated with the endoplasmic reticulum.
Two glycogenoses type 1 exist
Meanwhile, several researchers, including Géry Hers, found that there were two types of glycogenosis type I. Patients with the disease have very specific symptoms: hypoglycaemia, an overabundance of lactic acid, liver and kidney hypertrophy caused by their accumulation of glycogen – a stored carbohydrate that becomes glucose when the body needs it. Four times out of five patients have a glucose-6-phosphatase deficiency, but in about one in five glucose-6-phosphatase activity measured in liver biopsies is quite normal. Hence glycogenosis type Ia and type Ib, depending on whether glucose-6-phosphatase activity is deficient. But where is the problem in type 1b glycogenosis?
The endoplasmic reticulum (where G6PC1 is located) is surrounded by a membrane through which polar molecules cannot pass unless assisted by a suitable transport protein. This is the case here. The G6PC1 enzyme catalyses the glucose-6-phosphate hydrolysis reaction to D-glucose, but this reaction – crucial to our liver supplying the glucose our brain needs – takes place inside the endoplasmic reticulum, whereas glucose-6-phosphate is formed in the cell’s cytosol. For the conversion of glucose-6-phosphate to glucose to take place, a transporter must penetrate the glucose-6-phosphate in the reticulum. The transporter’s existence was postulated in the 1970s by an American group, but was identified only in 1997, by Emile Van Schaftingen and his collaborators. The UCLouvain researchers showed that the G6P transporter in patients with glycogenosis type Ib is mutated. The conversion reaction of glucose-6-phosphate into glucose cannot be triggered, exactly as if there was G6PC1 deficiency. The metabolic problems of patients with type Ib glycogenosis are thus easily explained.
The enigma of neutropenia…
But there was still a problem! In the 1960s, clinicians also identified another important difference between patients with type Ia or type Ib glycogenosis: the latter also suffer from neutropenia problems, i.e. a lack of neutrophils, the main white blood cells that defend us against bacteria. Hence, of course, the recurring infections. However, the neutropenia was not explained by the above-mentioned glucose-6-phosphate conversion problems in glucose – leaving a mystery that has endured to the present day.
… and three G6PC enzymes
Meanwhile, the sequencing of our genome has led to major advances, including the fact that our glucose-6-phosphatase is not unique and exists in three versions (or more precisely, three different genes that encode three similar proteins): G6PC1, G6PC2 and G6PC3. The last has interested our researchers because while it resembles G6PC1, its activity on glucose-6-phosphate is much lower. In addition, while G6PC1 is expressed in the liver and kidneys, G6PC3 is present in all tissues, as is the glucose-6-phosphate transporter. Especially interesting is that when this enzyme is deficient, there is no problem of metabolism (liver and kidney glycogen accumulation) but indeed a problem of neutropenia, as in glycogenosis type Ib.
This is what led the Duve Institute team to resume its research, in order to elucidate this phenomenon with the idea that G6PC3 and the glucose-6-phosphate transporter were probably used to destroy a still unknown molecule toxic to neutrophils. This unknown molecule would be transported within the endoplasmic reticulum by glucose-6-phosphate transporters and then hydrolysed by G6PC3.
Enzymes that clean house
But what was this molecule? Some detours can be fruitful. After its discoveries in 1997, the de Duve Institute team worked on other metabolic diseases. This helped lead to the surprising discovery of enzymes that ‘clean up’ during intermediary metabolism. We read in all the biochemistry books that enzymes of the major metabolic pathways are very specific in their purpose, that is, they produce only their physiological product, without waste. But the reality is different: all these enzymes generate small amounts of waste, some of which is toxic to the body. To avoid this toxicity – and this is an important contribution by our UCLouvain team – nature invented so-called metabolic repair enzymes that eliminate these toxic products by metabolising them. To date, the team has identified a dozen such enzymes, including G6PC3.
And the guilty party is …
But if G6PC3 is a repair enzyme, what does it repair? As observed, it hydrolyses glucose-6-phosphate poorly. Hence the idea of trying to find a substance that would be a good substrate for G6PC3 and could be transported by the glucose-6-phosphate transporter.
And this is what Maria Veiga da Cunha, Guido Bommer and Emile Van Schaftingen found: 1,5-anhydroglucitol-6-phosphate (1,5 AG6P), a derivative formed by the phosphorylation of 1,5-anhydroglucitol, a glucose analogue that we all have in the blood and is seemingly useless. Our three researchers have shown that if the G6PC3 repair enzyme does not play its role – does not clean – or if the G6PT transporter does not play its role, 1,5-anhydroglucitol-6-phosphate accumulates in neutrophils and causes them problems by blocking glucose metabolism.
Analyses of patients suffering from G6PC3 and G6PT deficiency thus showed that their neutrophils contain concentrations of 1,5AG6P at least 1,000 times higher than those found in healthy subjects.
A future treatment?
To treat patients, the concentration of 1,5AG6P in their cells should be lowered by decreasing the blood concentration of 1,5-anhydroglucitol, the molecule from which it is formed. Could this be achieved by removing 1,5-anhydroglucitol from the diet? Not easily, since it is in almost all foods. The solution is probably a drug used to fight diabetes that inhibits the transport of glucose from the kidney to the blood and causes a loss of glucose in the urine. This drug also causes a loss of 1,5-anhydroglucitol in the urine and therefore lowers its concentration in the blood. Our researchers have shown that it restores normal neutrophil levels in G6PC3-deficient mice. Of course clinical trials must be done before we can confirm the effectiveness of this treatment in humans. But if it is confirmed, it would have advantages over current treatments: less expensive, less frequently required, less painful, and probably with fewer side effects.
Henri Dupuis
A glance at the researchers' bios
Emile Van Schaftingen
Having been tempted to study physics, Emile Van Schaftingen ended up studying medicine, graduating in 1978 from UCLouvain, but for a specific purpose: research. He began as soon as the opportunity arose. On the team of Géry Hers, a student of Christian de Duve, he quickly became interested in the intermediary metabolism problems on which his mentor worked. A teacher training graduate in 1985, he directed the de Duve Institute until 2004 and taught biochemistry in the Faculty of Medicine until awarded emeritus status in 2018.
Maria Veiga da Cunha
After studying at the School of Agronomy of the Technical University of Lisbon, Maria Veiga da Cunha completed her agricultural engineering bachelor’s degree in 1986 at UCLouvain. In 1990, she graduated from the University of Oxford with a PhD in bacterial metabolism microbiology. She joined the de Duve Institute in 1992. Passionate about metabolism, she has worked in this field for more than 25 years in collaboration with Emile Van Schaftingen. Since 2000, she has been an FNRS research associate.
Guido Bommer
Guido Bommer completed his medical training at Ludwig Maximilian University in Munich (2001). Still in Munich, he specialised in gastroenterology, then conducted research at the University of Michigan (Ann Arbor, Michigan, USA). After a PhD in biomedical sciences at UCLouvain in 2009, he founded a team at the de Duve Institute to study aspects of metabolic control, particularly the role of microRNAs. An associate professor in the Department of Biochemistry, he is also an FNRS research associate. He recently received a European Research Council grant.