Todralazine Protects Zebrafish from Lethal Effects of Ionizing Radiation: Role of Hematopoietic Cell Expansion

The Johns Hopkins Clinical Compound Library (JHCCL), a collection of Food and Drug Administration (FDA)-approved small molecules (1400), was screened in silico for identification of novel beta(2)AR blockers and tested for hematopoietic stem cell (HSC) expansion and radioprotection in zebrafish embryos. Docking studies, followed by the capacity to hasten erythropoiesis, identified todralazine (Binding energy, -8.4 kcal/mol) as a potential HSC-modulating agent. Todralazine (5 mu M) significantly increased erythropoiesis in caudal hematopoietic tissue (CHT) in wild-type and anemic zebrafish embryos (2.33- and 1.44-folds, respectively) when compared with untreated and anemic control groups. Todralazine (5 mu M) treatment also led to an increased number of erythroid progenitors, as revealed from the increased expression of erythroid progenitor-specific genes in the CHT region. Consistent with these effects, zebrafish embryos, Tg(cmyb:gfp), treated with 5 mu M todralazine from 24 to 36 hours post fertilization (hpf) showed increased (approximately two-folds) number of HSCs at the aorta-gonad-mesonephros region (AGM). Similarly, expression of HSC marker genes, runx1 (3.3-folds), and cMyb (1.41-folds) also increased in case of todralazine-treated embryos, further supporting its HSC expansion potential. Metoprolol, a known beta blocker, also induced HSC expansion (1.36- and 1.48-fold increase in runx1 and cMyb, respectively). Todralazine (5 mu M) when added 30 min before 20 Gy gamma radiation, protected zebrafish from radiation-induced organ toxicity, apoptosis, and improved survival (80% survival advantage over 6 days). The 2-deoxyribose degradation test further suggested hydroxyl (OH) radical scavenging potential of todralazine, and the same is recapitulated in vivo. These results suggest that todralazine is a potential HSC expanding agent, which might be acting along with important functions, such as antioxidant and free radical scavenging, in manifesting radioprotection.