A Novel Human Pth Analog [Cys25]Hpth(1-34) Restores Bone Mass In Ovariectomized Mice

Context: Recently, an arginine-to-cysteine homozygous mutation at position 25 in mature PTH was reported in a Korean patient with hypoparathyroidism.Objective: To clarify whether the high bone mass phenotype observed in this patient was related to the hypoparathyroidism itself or to chronic elevation of mutant PTH.Methods: A series of in vitro and in vivo experiments were performed in MC3T3E1, ROS 17/2.8, and SAOS2 cells treated with human (h) PTH(1-34), Cys(25)hPTH(1-34), Ala1Cys(25)hPTH(1-34), and Bpa(1)Cys(25)hPTH(1-34). The peptides were then sc delivered to ovariectomized mice as daily single injections.Results: Compared with hPTH(1-34) and Ala(1)Cys(25)hPTH(1-34), treatment with Cys(25)hPTH(1-34) or Bpa(1)Cys(25)hPTH(1-34) resulted in decreases in the cAMP response and promoter-cAMP-response element luciferase reporter activity. Although the cAMP response was sustained with hPTH(1-34) in MC3T3E1 cells, such response was not observed with the other mutated peptides. Meanwhile, all PTH analogues exhibited ERK phosphorylation and cytoplasmic Ca++ signals comparable with hPTH(1-34). On microcomputed tomography analyses, trabecular and cortical bone parameters improved after 6 weeks of respective treatments as follows: hPTH(1-34) (80 mu g/kg) = Ala(1)Cys(25)hPTH(1-34) (80 mu g/kg) = Cys(25)hPTH(1-34) (80 mu g/kg) > Bpa(1)Cys(25)hPTH(1-34) (80 mu g/kg) = hPTH(1-34) (40 mu g/kg). The increment of RANKL to OPG mRNA ratio in the MC3T3E1 cells after 6 hours of treatment of Cys(25)hPTH(1-34), AL(1)Cys(25)hPTH(1-34), and Bpa(1)Cys(25)hPTH(1-34) was less than that was obtained after hPTH(1-34) treatment. On bone histomorphometric analysis, AL(1)Cys(25)hPTH(1-34) increased the bone formation rate in both trabecular and periosteal bones compared with the control group.Conclusion: The high bone mass phenotype observed in this patient with hypoparathyrodism caused by a Cys mutation at the 25th residue of hPTH(1-84) may have arisen from both direct and indirect effects exerted by the mutant PTH itself on bone.