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620 A Novel Mouse Model of Hemoglobin-SC Disease: Bridging the Knowledge Gap in Sickle Cell Disease

Program: Oral and Poster Abstracts
Type: Oral
Session: 113. Sickle Cell Disease, Sickle Cell Trait, and Other Hemoglobinopathies, Excluding Thalassemias: Basic and Translational: Attenuating Sickle Cell Disease Complications – Lessons from Pre-clinical Models
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Genetic Disorders, Diseases
Sunday, December 8, 2024: 4:45 PM

Tahereh Setayesh1*, Mengna Chi, PhD1*, Zachery Oestreicher, PhD2*, Masahide Sakabe, PhD1*, Xin Mei, PhD1*, Katherine Vandenheuvel, MD3*, Tim M. Townes, PhD4*, Yueh-Chiang Hu, PhD5* and Punam Malik, MD3,6

1Cincinnati Children's Hospital Medical Center (CCHMC), Cincinnati, OH
2Center for Advanced Microscopy and Imaging, University of Miami, Oxford, Ohio, United States, Oxford, OH
3Cincinnati Children's Hospital Medical Center, Cincinnati, OH
4Univ. of AL at Birmingham, Birmingham, AL
5Transgenic Animal and Genome Editing Core, Cincinnati Children's Hospital, Cincinnati, OH
6Cincinnati Children's Hospital Medical Center, Cincinnati

Introduction: Hemoglobin-SC (HbSC) disease occurs from compound heterozygosity of hemoglobin S (HbS) and hemoglobin C (HbC), affecting 30% of patients with sickle cell disease (SCD). While it has a milder phenotype relative to homozygous HbSS SCD, HbSC disease leads to significant morbidity, early mortality, and severe complications, some occurring at a higher incidence than HbSS disease, such as proliferative sickle retinopathy and avascular necrosis, from the increased blood viscosity. While extensive research has focused on HbSS disease to develop treatment strategies, HbSC disease is highly understudied, leading to a lack of therapies for this genotype of SCD. HbSC patients are deprived of therapies like gene therapy due to the lack of preclinical models or clinical testing, as they represent a minority SCD population.

Methods: We used CRISPR editing to modify humanized Townes HbSS (HBBE6V/E6V) mice, converting one of the HBBE6V alleles to HBBE6K, generating HbSC (HBBE6V/E6K) mice. We then compared the phenotype of HbSC mice directly with HbSS mice, which are otherwise genetically identical except for the HBBE6K mutation on one allele, and to the Townes HbAA (HBBE6/E6) controls. The hematological, and specifically RBC parameters, RBC morphology, rheology, hydration, sickling kinetics, whole blood viscosity, and organ pathology, including retinal vasculature, were analyzed. Mice were also subjected to hypoxia/reoxygenation to mimic vaso-occlusive crisis.

Results: The HbSC mice exhibited hematological characteristics like HbSC patients with anemia but higher Hb and RBC counts, and lower reticulocytes than HbSS mice. Moreover, HbSC mouse RBCs showed features distinct to HbSC disease, with significant RBC xerocytosis reflected in higher mean corpuscular hemoglobin concentration, lower hematocrits, and higher whole blood viscosity, as compared to HbSS mouse RBC. Scanning electron microscopy showed the classic membrane folding with pita-shaped RBC and target cells in HbSC mice. This xerocytosis was also present in HbSC reticulocytes. Osmoscan ektacytometry analysis confirmed severe dehydration in HbSC RBCs, akin to human HbSC RBCs and sickling kinetics mimicked those seen in human HbSC RBCs. The HbSC mice showed longer RBC half-life, less hemolysis and inflammation than HbSS mice (albeit significantly different from HbAA mice), but had significant organ damage, including sickle retinopathy. Additionally, we observed significant hypoxia-induced membrane damage, specifically in HbSC RBCs, although RBC dehydration occurred universally in all genotypes.

Conclusion: The HbSC mouse model successfully mimics human HbSC disease, providing a critical tool for studying HbSC disease pathophysiology and testing novel therapies, which can be specifically tailored to HbSC disease. The development of this model bridges a significant knowledge gap that will enhance treatment strategies and help in improving the quality of life of the hitherto neglected HbSC subset of SCD patients.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH