Paroxysmal nocturnal hemoglobinuria (PNH) is a stem cell disorder caused by a mutation in a gene called phosphatidylinositol glycan anchor biosynthesis class A gene (PIGA). PIGA produces a protein important in the formation of a group of proteins called glycophosphatidylinositol anchored proteins (GPI-AP). GPI-AP are attached to the cell membrane via a small anchor. Among them CD59 and CD55 are major targets for therapy.
Due to PIGA mutation, PNH cells fail to synthesize a correct anchor and thereby lack of proper GPI-AP. To date, the drug developments for PNH have focused on supportive therapies to improve hemoglobin levels due to hemolysis or prevent thrombotic complications. Most successful to date were complement blockers designed for alleviation of hemolysis due to deficiency in complement. These therapies work by deactivating GPI-AP (CD55 and CD59). Our proposal focuses on the possibility to test agents selectively reducing the fraction of GPI-AP deficient cells and consequently improving the growth of GPI-AP normal cells.
We identified an agent (CGS-15943) selectively arresting the growth of GPI-AP deficient cells. CGS-15943 has a potential for translation into PNH precision medicine and might represent an alternative curative strategy for PNH.
Paroxysmal nocturnal hemoglobinuria (PNH) is caused by defects to DNA called mutations which occur in a gene called phosphatidylinositol glycan anchor biosynthesis class A (PIGA). These genetic defects happen very early in the formation of bone marrow stem cells (cells that give rise to every other cell in the blood). Cells with PIGA mutations lack a vast class of proteins which act to protect blood cells from being destroyed and fight infection. Clinical problems experienced by patients with PNH are the result of cells lacking those proteins. These cells proliferate over time and can eventually out-number the cells with normal function. Cells with PIGA mutations are also frequently found in patients with aplastic anemia, another life-threatening bone marrow disorder. To date, medicines available to treat patients with PNH are very limited. Our proposal is based on the possibility to pre-clinically test drugs improving the function of normal cells and decreasing the fractions of cells with PIGA mutations. We identified an agent (CGS-15943) that has a potential for translation into PNH precision medicine and might represent an alternative curative strategy for PNH.
Paroxysmal nocturnal hemoglobinuria (PNH) is caused by defects to DNA called mutations which occur in a gene called phosphatidylinositol glycan anchor biosynthesis class A (PIGA). These genetic defects happen in early stages of the formation of bone marrow stem cells: cells that give rise to every other cell in the blood including red blood cells, white blood cells and platelets. All cells of the progeny having PIGA mutations lack a vast class of proteins which have different functions in the cells and act to protect blood cells from being destroyed and fight infection. Clinical problems experienced by patients with PNH are the result of cells lacking these proteins. In fact patients with PNH have more risk to form blood clots and have organ damage. PNH cells with improper functions can proliferate over time and can take advantage over cells with normal function. Cells with PIGA mutations are also frequently found in patients with aplastic anemia (AA), another fatal disease in which the bone marrow cannot produce enough blood cells in the body.
To date, medicines available to treat patients with PNH are very limited. Recently we have studied the effects of many medicines possibly improving the function of normal cells in patients with PNH. Our study is based on the identification of new medicines with less toxicity against normal cells and our experimental approach is directed towards conditions in which a minor fraction of PNH cells are detected at a very early stage of the disease (especially in AA). This approach is intended to slow down the later complications due to the expansion of PNH cells and not to eradicate PNH cells in conditions in which these cells are the ones sustaining blood cell production.
Using a large screening of chemicals and natural compounds already approved by the U.S. Food and Drug Administration or tested in previous studies we identified one chemical substance called CGS- 15943 which has an effect in improving the proliferation of normal cells by decreasing the number of PNH cells. This medicine is known to improve the formation of bone marrow stem cells and regulate the differentiation of these cells in other blood cells with different functions (red cells, white cells, etc.). These experiments were conducted using cell models without and with the presence of key proteins working in preventing an abnormal activation of the complement system, such as CD55 and CD59. If these proteins are excessively activated they are unable to eliminate abnormal cells. We used cell models characterized by different types of cells such as lymphocytes called T-cells and bone marrow cells. These cellular models have been maintained in our laboratory or derived from culturing cells of patients with PNH. These cells were kept in contact with CGS 15943 at different doses for three days and the dose which had no toxic effects against normal cells was selected for further studies. Even in experiments in which we mixed PNH and normal cells we found that CGS 15943 was able to selectively induce a decrease in PNH cells while potentiating the proliferation of normal cells. While testing the effects of the medicine on cell proliferation we found that CGS 15943 not only increased the number of cells with normal function but also changed the levels of many proteins important for the function of blood cells. In fact CGS 15943 doubled the quantity of several factors driving the maturation and specialization of blood cells (RUNX1 and CD11b) and increased the quantity of factors involved in the formation of bone marrow stem cells (for instance ADA) suggesting that treatment with this medicine could help the production of blood cells in patients with PNH. This study also opens the possibility to develop new medicines using the characteristics of CGS 15943 given the fact that part of the shape of this substance might have crucial functions towards PNH cells.