Aurin Tricarboxylic Acid Protects against Red Blood Cell Hemolysis in Patients with Paroxysmal Nocturnal Hemoglobinemia

Moonhee Lee,Patrick L Mcgeer,Edith G Mcgeer,Sujaatha Narayanan

doi:10.1371/journal.pone.0087316

Moonhee Lee, Patrick L Mcgeer + Show 2 more

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https://doi.org/10.1371/journal.pone.0087316

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Abstract

ObjectivesParoxysmal nocturnal hemoglobinemia (PNH) is a rare but serious condition characterized by complement-mediated red blood cell (RBC) hemolysis and episodic thrombotic attack. It results from decay accelerating factor (CD55), and protectin (CD59), becoming attached to RBC and other cell surfaces. Absence of these protective proteins leaves such cells vulnerable to self attack at the C3 convertase and membrane attack complex (MAC) stages of complement activation. We have previously reported that aurin tricarboxylic acid (ATA) is an orally effective agent that selectively blocks complement activation at the C3 convertase stage as well as MAC formation at the C9 insertion stage.Design and MethodsWe used a CH50 assay method and western blot analysis to investigate the vulnerability to complement attack of PNH RBCs compared with normal RBCs. Zymosan was used as the activator of normal serum and PNH serum. ATA was added to the sera to determine the concentration necessary to protect the RBCs from lysis by the zymosan-activated sera.ResultsWe found that erythrocytes from PNH patients on long term treatment with eculizumab were twice as vulnerable as normal erythrocytes to lysis induced by complement activated serum. Western blot data showed the presence of both C3 and C5 convertases on the PNH patient erythrocyte membranes. These data indicate persistent vulnerability of PNH erythrocytes to complement attack due to deficiencies in CD55 and CD59. ATA, when added to serum in vitro, protected PNH erythrocytes from complement attack, restoring their resistance to that of normal erythrocytes.ConclusionsWe conclude that ATA, by protecting PNH erythrocytes from their decay accelerating factor (CD55) and protectin (CD59) deficiencies, may be an effective oral treatment in this disorder.

Highlights

Paroxysmal nocturnal hemoglobinemia (PNH) is an episodic disorder involving complement-mediated hemolytic anemia, with an accompanying risk of thrombosis [1,2]
We found that erythrocytes from PNH patients on long term treatment with eculizumab were twice as vulnerable as normal erythrocytes to lysis induced by complement activated serum
Western blot data showed the presence of both C3 and C5 convertases on the PNH patient erythrocyte membranes

Summary

Introduction

Paroxysmal nocturnal hemoglobinemia (PNH) is an episodic disorder involving complement-mediated hemolytic anemia, with an accompanying risk of thrombosis [1,2]. PNH is a rare disease that was first recognized in the second half of the nineteenth century. It was not properly understood until investigators discovered that PNH patients develop stem cell clones in their marrow that have a deletion of glycosyl phosphoinositol (GPI)anchored proteins (GPI-APs) [3]. Genetic studies have identified the cause to be somatic mutations in the gene phosphatidylinositol glycan class A (PIG-A) [4,5]. The gene encodes enzymes catalysing the first step of GPI-anchor-biosynthesis, in which there is a transfer of N-acetylglucosamine to phosphatidylinositol in hematopoietic stem cells [4,5]. The proteins which fail to become anchored, and are non-functional, include decayaccelerating factor (DAF, CD55), an inhibitor of alternative pathway C3 convertase, and protectin (CD59), an inhibitor of membrane attack complex (MAC) formation [6,7,8]

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