Negative regulation of the osteoblast function in multiple myeloma through the repressor gene E4BP4 activated by malignant plasma cells.

PURPOSE

To explore the pathogenetic mechanisms that suppress the osteoblast function in multiple myeloma because osteogenesis results in defective new bone formation and repair.

EXPERIMENTAL DESIGN

Microarray gene analysis revealed the overexpression of E4BP4, a transcriptional repressor gene, in normal osteoblasts cocultured with myeloma cells that were releasing the parathyroid hormone-related protein (PTHrP). Thus, the effect of E4BP4 was assessed in PTHrP-stimulated osteoblasts by measuring the RNA levels of both Runx2 and Osterix as major osteoblast transcriptional activators. Because E4BP4 is a negative regulator of the cyclooxygenase-2 (COX-2) pathway that drives the expression of both Runx2 and Osterix, these factors were investigated after prostaglandin E(2) treatment to overcome the COX-2 defect as well as in E4BP4-silenced osteoblasts. Finally, E4BP4, PTHrP, Osterix, and osteocalcin levels were measured in vivo in patients with bone disease together with the E4BP4 protein in bone biopsies.

RESULTS

E4BP4 was specifically induced by PTHrP and inhibited both Runx2 and Osterix, whereas E4BP4-silenced osteoblasts expressed functional levels of both factors. The prostaglandin E(2) treatment of E4BP4-up-regulated osteoblasts promptly restored Runx2 and Osterix activities, suggesting that integrity of COX-2 pathway is essential for their transcription. Down-regulation of Osterix by E4BP4 was confirmed in vivo by its inverse levels in osteoblasts from myeloma patients with increased serum PTHrP, whose bone biopsies expressed the E4BP4 protein.

CONCLUSIONS

Our data support the role of E4BP4 as osteoblast transcriptional repressor in inhibiting both Runx2 and Osterix in myeloma bone disease and correlate its effect with the increased PTHrP activity.