A large group of scientists from the USA and China have published exciting new observations about the sickling process in The Journal of Clinical Investigation, which may well have important treatment implications for patients with sickle cell disease.
The fundamental problem in sickle cell disease is the coming together of sickle haemoglobin molecules, within red blood cells, to form tactoids. These long, rigid, crystal-like structures distort the shape of the red cell, result in obstruction to blood flow or vaso-occlusion and cause anaemia, due to a shortened red cell lifespan. The process of sickling, what initiates it and what controls it, is poorly understood. In a previous blog (03/06/14: Update on Clinical Trials in Sickle Cell Disease) I discussed a drug called Aes-103, currently in clinical trial, which interferes with the sickling process by stabilising sickle haemoglobin in a high oxygen affinity state. Whether this drug will be effective in clinical practice remains to be seen; all previous attempts to use drugs to block the sickling process have failed. This current research identifies a new, previously unknown, biochemical pathway, which promotes sickling and sickle cell related tissue damage, and which may be amenable to therapeutic manipulation.
The researchers used a process called metabolomic screening to look at thousands of chemicals within red cells from patients with sickle cell disease, and then compared them with the same chemicals from individuals with normal blood. They focused on a particular chemical called sphingosine-1-phosphate (S1P), which they found was present in high concentrations in all red cells, but which was increased to very high levels in the red cells and blood of patients with sickle cell disease. What all of this S1P inside red cells is doing is not clear but, in other parts of the body, it is known to be an important chemical involved in a variety of biological processes, including inflammation, blood vessel growth, damage to the lining of blood vessels and blood clotting. Inside red cells S1P is produced by an enzyme called sphingosine kinase 1 (SPHK1); the researchers found that the activity of this enzyme is greatly increased in sickle red cells accounting for the very high levels of S1P. They were able to manipulate the levels of S1P using a new drug, which is a potent inhibitor of the enzyme SPHK1, called PF-543.
Using a strain of mice, which have been genetically engineered so that they have sickle cell disease, the scientists found that treatment of the mice with PF-543 significantly reduced levels of S1P and, to their surprise, they found that this was also associated with an improvement in the appearance and shape of the red cells in the blood of the mice. Further work showed that treatment with PF-543 resulted in reduced red cell sickling, reduced haemolysis with improved anaemia and a reduced white cell count, reflecting an anti-inflammatory effect. As a result of these changes the sickle cell mice were healthier, there was less tissue and organ damage and they survived longer, even if exposed to low oxygen levels, which would normally cause extensive, fatal sickling. The scientists confirmed these unexpected findings using a different technique, in which they de-activated, or knocked-out, the gene making the enzyme SPHK1, in chimeric mice with sickle cell disease, using a lentivirus. These knock-out mice were also healthier with less sickle cell related tissue and organ damage.
But is this relevant to patients with sickle cell disease? There is no reason to think that human sickle cell disease is any different to the condition in the mice. The researchers found that S1P and SPHK1 are also present at very high concentrations in patients with sickle cell disease and, in the test tube at least, treatment of human sickle red cells with the enzyme inhibitor, PF-543, reduced the amount of sickling which occurred if the red cells were exposed to low levels of oxygen, results similar to those found in sickle cell mice.
On the basis of these results the authors think that S1P is a key, previously unknown, chemical, which promotes sickling by acting within the red cell on sickle haemoglobin. The high levels of S1P are due to activation of the enzyme, SPHK1, probably by low levels of oxygen, which are commonly found in patients with sickle cell disease, explaining the very high levels found in the disease. Blocking the activity of this enzyme has a dramatic effect, it reduces the level of S1P and alleviates many of the consequences of red cell sickling. These observations open up the possibility of potential treatments for sickle cell disease in the future, by manipulating this new biochemical pathway. It is important now to understand how exactly S1P interacts with sickle haemoglobin to cause increased sickling and whether the enzyme inhibitor PF-543 can safely and effectively be given to patients with sickle cell.
Elevated sphingosine-1-phosphate promotes sickling and sickle cell disease progression. Journal of Clinical Investigation 2014; volume 124, pages 2750-2761.