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Background: Apohemoglobin-haptoglobin (apoHb-Hp) complex represents a bioengineered complex designed to scavenge hemoglobin and heme. These substances are released into the intravascular space during hemolysis, leading to vasoconstriction, inflammation, and disease progression. The use of apoHb-Hp complex has already demonstrated promising results in sickle cell and beta-thalassemia mouse models, highlighting its potential as a therapeutic agent. However, there are still crucial aspects that remain poorly understood, particularly concerning the biodistribution and pharmacokinetics associated during chronic administration. Understanding how the drug is distributed throughout the body is essential for optimizing its therapeutic potential, ensuring patient safety, and guiding both drug development and clinical use.

Introduction

Introduction:In conditions with significant hemolysis, like sickle cell and thalassemia, red blood cells break down, leading to higher levels of cell-free hemoglobin (Hb) in the bloodstream. When this cell-free Hb is oxidized into methemoglobin, it releases heme, a crucial iron-protoporphyrin complex essential for oxygen transport. However, excessive free heme can become problematic, since plasma concentrations of the body protective proteins, haptoglobin and hemopexin, to scavenge and clear hemoglobin and heme is very limited. Thus, these protective mechanisms can get overwhelmed by high levels of free Hb or heme in the blood. Additionally, heme can interact with the coagulation cascade, potentially causing harmful pro-coagulant effects in the body. To address this, we introduce a new synthetic scavenger protein called apohemoglobin-haptoglobin (apoHb+Hp). It mimics hemopexin's function and binds to free heme with even greater affinity. Moreover, it utilizes haptoglobin's clearance mechanism for efficient removal from the plasma. Our aim is for apoHb+Hp to effectively clear free heme from the plasma, thereby alleviating pro-coagulatory markers induced by heme. This novel approach may enhance the body's natural defense against heme, leading to potential therapeutic benefits.

The constant ischemia-reperfusion injury and the release of hemoglobin and heme in patients with sickle cell disease (SCD) can affect the heart, lungs, and vasculature. While treatments have improved survival, cardiovascular complications are becoming more prevalent, including pulmonary hypertension and left ventricular diastolic dysfunction. Pulmonary hypertension is associated with increased mortality risk and linked to intravascular hemolysis, leg ulcers, renal insufficiency, iron overload, and liver dysfunction. Chronic anemia leads to heart chamber dilation and left ventricular mass increase, often accompanied by left ventricular diastolic dysfunction, both impacting exercise capacity. Additionally, it can cause Acute Chest Syndrome (ACS), which is characterized by inflammation and blockage of small blood vessels in the lungs. In some cases, the heart may enlarge (cardiomegaly) due to the increased workload it faces in pumping blood through narrowed or obstructed blood vessels. Sudden death is also emerging as a concern, necessitating further cardiac investigations and risk assessment for effective management. These issues are speculated to result from the high amount of hemoglobin and heme released into circulation.

Pathophysiological conditions characterized by chronic hemolysis, such as genetic and acquired hemolytic diseases, lead to increased levels of circulatory cell-free hemoglobin (fHb), free heme, and iron. The toxicity of free acellular hemoglobin has been a subject of intense study to understand its impact on the human body. fHb's small size allows it to extravasate into various organs, causing damage. The kidney is particularly vulnerable to Hb toxicity as it can filtrate through the kidneys, affecting glomerular filtration rate and leading to kidney failure.

Thalassemia is a blood disorder that affects the production of hemoglobin (Hb), the crucial protein responsible for carrying and delivering oxygen to tissues. It is caused by mutations in the genes that control the production of hemoglobin, leading to decreased or absent production of one of the globin chains (alpha or beta) that make up hemoglobin. Individuals with thalassemia often need regular blood transfusions to maintain adequate Hb levels. However, frequent transfusions can cause iron overload in the body, necessitating chelation therapy to remove excess iron and prevent organ damage. The iron overload also results in higher levels of hemolysis, leading to the release of free acellular Hb and heme, which the body's natural scavenging proteins (hemopexin and haptoglobin) cannot effectively handle.

Sickle Cell Disease (SCD) is characterized by hemolytic anemia, vaso-occlusion, and progressive end-organ damage. The underlying mechanism of SCD is the polymerization of sickle hemoglobin (HbS) that occurs when sickle erythrocytes (SS RBCs) are partially deoxygenated in microcirculation, leading to SCD pathophysiologic features. One of the most devastating complications of SCD occurs in the central nervous system (CNS), where overt stroke or repeated silent cerebral infarcts lead to significant physical and neurocognitive consequences. In SCD, the brain's response to insufficient oxygen (O2) delivery is balanced by increased blood flow to preserve O2supply. However, the systemic endothelial dysfunction in SCD limits the capacity for vascular regulatory and compensatory changes to preserve appropriate tissue oxygenation, especially in tissues with high O2demand like the brain. Low hemoglobin (Hb) levels and increased cerebral blood flow (CBF) are associated with increased stroke risk, suggesting that anemia-induced tissue hypoxia is an important factor contributing to subsequent morbidity in SCD patients.