Glucocorticoids Reduce Expression Of p53 Target Genes and Reverse Anemia In a Mouse Model Of Diamond-Blackfan Anemia

Diamond-Blackfan anemia (DBA) is a rare congenital disease of macrocytic anemia, increased cancer predisposition and progressive bone marrow failure. Mutations in ribosomal protein genes are responsible for most cases and the most commonly mutated gene, RPS19 is affected in around 25% of DBA patients. Although glucocorticoid (GC) administration has been used to stimulate erythropoiesis in DBA patients for decades, the therapeutic mechanism of GC treatment is poorly understood. Since it has been shown that bone marrow failure in DBA is caused by p53 activation we hypothesized the therapeutic mechanism of GC involves reduced p53 activity. To assess possible connections between p53 activity and GC administration in DBA we used a doxycycline-inducible mouse model for RPS19-deficient DBA (Jaako et al, Blood 2011). In these mice, doxycycline administration leads to reduced Rps19 levels, defective ribosomal biogenesis, proliferative arrest, increased apoptosis of erythroid progenitors and p53-dependent bone marrow failure.

In this study we show that Rps19-deficient mice respond to treatment with the GC prednisolone, with a significant increase in red blood cell count as well as in hemoglobin concentration and hematocrit compared to Rps19-deficient controls (p-values<0.05). Also, prednisolone treated Rps19-deficient mice show a significant increase in the Ter119⁺ erythroid cell fraction in the bone marrow compared to Rps19-deficient controls. Gene expression analysis of c-Kit⁺ fetal liver cells from E14.5-15.5 of Rps19-deficient embryos cultured in vitro in the presence of doxycycline for 3 days, showed an increase in p53 responsive gene expression of multiple p53 targets such as p21, Bax, Ccng1, Phlda3 and Prl-3. However, upon administration of the synthetic GC dexamethasone all these genes significantly failed to up-regulate, even in the presence of doxycycline (p-values< 0.02). Also, dexamethasone treated RPS19-deficient c-Kit⁺ fetal liver cells show an increase in proliferation compared to RPS19-deficient controls (p-value< 0.05), that starts already after 2 days of in vitro culture, being more pronounced over time. These results indicate that administration of GC dampens p53 responsive gene expression of genes known to promote cell cycle arrest as well as apoptosis. Although GCs are known to regulate many functions within an organism, our results indicate that the therapeutic effect of GC in DBA patients at least in part is explained by inhibition of p53 responsive gene activation, leading to increased proliferation and survival of erythroid progenitors.