Calcium-oxidative stress signaling axis and casein kinase 1α mediate eryptosis and hemolysis elicited by novel p53 agonist inauhzin

Abstract

Inauhzin (INZ) is a novel p53 agonist with antitumor activity. Anemia is a common side effect of chemotherapy and may arise from red blood cell (RBC) hemolysis or eryptosis. In this study, we investigate the mechanisms of INZ toxicity in human RBCs. RBCs were isolated from healthy donors and treated with antitumor concentrations of INZ (5–500 μM) for 24 h at 37 °C. Hemoglobin was photometrically measured, and cells were stained with Annexin-V-FITC for phosphatidylserine (PS), Fluo4/AM for calcium, and 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFDA) for oxidative stress. INZ caused significant dose-responsive, calcium-dependent hemolysis starting at 40 μM. Furthermore, INZ significantly increased Annexin-positive cells and Fluo4 and DCF fluorescence. The cytotoxicity of INZ was also significantly mitigated in presence of D4476. INZ possesses hemolytic and eryptotic potential characterized by cell membrane scrambling, intracellular calcium overload, cell shrinkage, and oxidative stress secondary to calcium influx from the extracellular space.

Keywords:

• Inauhzin
• hemolysis
• eryptosis
• calcium
• oxidative stress
• casein kinase
• anticancer

Acknowledgment

The authors extend their appreciation to the Deanship of Scientific Research at King Saud University for funding this work through research group no. RG-1441-335.

Disclosure statement

No potential conflict of interest was reported by the authors.

Ethics approval and consent to participate

This study was approved by the Ethics Committee of the College of Medicine, King Saud University (Project No. E-20-4544). All participants have provided written informed consent to participate in this study.

Author contributions

M.A.A.: conceptualization; funding acquisition; data curation; formal analysis; investigation; methodology; project administration; software; supervision; writing-original draft; and writing review and editing. E.H.A.: data curation; formal analysis; investigation; and methodology. A.H.A.: project administration; resources; and supervision. J.A.: conceptualization; data curation; writing-original draft; and writing review and editing. M.A.: data curation; formal analysis; and resources. A.M.B.: data curation; formal analysis; and resources.

Data availability statement

All data in this study are available and included in this published article.