A Novel Human PTH Analog [Cys25]hPTH(1-34) Restores Bone Mass in Ovariectomized Mice.

Abstract

Recently, an arginine-to-cysteine homozygous mutation at position 25 in mature PTH was reported in a Korean patient with hypoparathyroidism. 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. 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), Cys²⁵hPTH(1-34), Ala¹Cys²⁵hPTH(1-34), and Bpa¹Cys²⁵hPTH(1-34). The peptides were then sc delivered to ovariectomized mice as daily single injections. Compared with hPTH(1-34) and Ala¹Cys²⁵hPTH(1-34), treatment with Cys²⁵hPTH(1-34) or Bpa¹Cys²⁵hPTH(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 μg/kg) = Ala¹Cys²⁵hPTH(1-34) (80 μg/kg) = Cys²⁵hPTH(1-34) (80 μg/kg) > Bpa¹Cys²⁵hPTH(1-34) (80 μg/kg) > hPTH(1-34) (40 μg/kg). The increment of RANKL to OPG mRNA ratio in the MC3T3E1 cells after 6 hours of treatment of Cys²⁵hPTH(1-34), AL¹Cys²⁵hPTH(1-34), and Bpa¹Cys²⁵hPTH(1-34) was less than that was obtained after hPTH(1-34) treatment. On bone histomorphometric analysis, AL¹Cys²⁵hPTH(1-34) increased the bone formation rate in both trabecular and periosteal bones compared with the control group. 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.