High-Throughput Assay to Screen Small Molecules for Their Ability to Prevent Sickling of Red Blood Cells,ACS Omega

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High-Throughput Assay to Screen Small Molecules for Their Ability to Prevent Sickling of Red Blood Cells
ACS Omega ( IF 3.7 ) Pub Date : 2022-04-15 , DOI: 10.1021/acsomega.2c00541
Akito Nakagawa ₁ , Marissa K Cooper ₁ , Maria Kost-Alimova ₂ , James Berstler ₂ , Binglan Yu ₁ , Lorenzo Berra ₁ , Elizabeth S Klings ₃ , Mary S Huang ₄ , Matthew M Heeney ₅ , Donald B Bloch _{1,

6} , Warren M Zapol ₁

Affiliation

Sickle cell disease (SCD) is an inherited disorder of hemoglobin (Hb); approximately 300,000 babies are born worldwide with SCD each year. In SCD, fibers of polymerized sickle Hb (HbS) form in red blood cells (RBCs), which cause RBCs to develop their characteristic “sickled” shape, resulting in hemolytic anemia and numerous vascular complications including vaso-occlusive crises. The development of novel antisickling compounds will provide new therapeutic options for patients with SCD. We developed a high-throughput “sickling assay” that is based on an automated high-content imaging system to quantify the effects of hypoxia on the shape and size of RBCs from HbSS SCD patients (SS RBCs). We used this assay to screen thousands of compounds for their ability to inhibit sickling. In the assay, voxelotor (an FDA-approved medication used to treat SCD) prevented sickling with a z′-factor > 0.4, suggesting that the assay is capable of identifying compounds that inhibit sickling. We screened the Broad Repurposing Library of 5393 compounds for their ability to prevent sickling in 4% oxygen/96% nitrogen. We identified two compounds, SNS-314 mesylate and voxelotor itself, that successfully prevented sickling. SNS-314 mesylate prevented sickling in the absence of oxygen, while voxelotor did not, suggesting that SNS-314 mesylate acts by a mechanism that is different from that of voxelotor. The sickling assay described in this study will permit the identification of additional, novel antisickling compounds, which will potentially expand the therapeutic options for SCD.

中文翻译：

高通量测定筛选小分子防止红细胞镰状化的能力

镰状细胞病 (SCD) 是一种血红蛋白 (Hb) 遗传性疾病；全球每年约有 300,000 名婴儿出生时患有 SCD。在 SCD 中，红细胞 (RBC) 中形成聚合镰状血红蛋白 (HbS) 纤维，导致红细胞形成其特有的“镰状”形状，导致溶血性贫血和包括血管闭塞危象在内的多种血管并发症。新型抗镰状化合物的开发将为SCD患者提供新的治疗选择。我们开发了一种基于自动化高内涵成像系统的高通量“镰状化测定”，用于量化缺氧对 HbSS SCD 患者红细胞 (SS RBC) 形状和大小的影响。我们使用这种测定法筛选了数千种化合物的抑制镰状化的能力。在该测定中，voxelotor（FDA 批准的用于治疗 SCD 的药物）通过z ' 因子 > 0.4 预防镰状化，表明该测定能够识别抑制镰状化的化合物。我们筛选了包含 5393 种化合物的广泛再利用库，了解它们在 4% 氧气/96% 氮气中防止镰状化的能力。我们鉴定出两种化合物，SNS-314 甲磺酸盐和 voxelotor 本身，可以成功防止镰状化。 SNS-314 甲磺酸盐可在缺氧条件下防止镰状化，而 voxelotor 则不能，这表明 SNS-314 甲磺酸盐的作用机制与 voxelotor 不同。本研究中描述的镰状化分析将允许鉴定额外的新型抗镰状化化合物，这将有可能扩大 SCD 的治疗选择。

更新日期：2022-04-15

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