Fig. 1: Design and characterization of Palm-PTH(1-34).
From: Prolonging parathyroid hormone analog action in vitro and in vivo through peptide lipidation
A Schematic of Palm-PTH(1-34). PTH(1-34) is C-terminally fused to a hepta-residue tag EYEK(palm)EYE optimized for albumin binding and containing a palmitoyl group (palm) linked to the central lysine side chain amine; a (PEG)2 (20 atom) spacer joins Phe34 of PTH to Glu1 of the tag. B cAMP responses in SGS-72 human osteoblast-derived cells expressing the glosensor cAMP reporter; data are calculated as the area-under-the-curve (AUC) of plots of luminescence vs. time (60’) normalized to the PTH(1-34) maximum and are means ± SEM of 10 independent experiments. C–F Pharmacodynamic and pharmacokinetic measurements of PTH(1-34) and Palm-PTH(1-34) in 10-week-old female CD1 mice; peptides injected at a dose of 10 nmol/kg (C–E) or 50 nmol/kg (F). Blood concentrations of ionized calcium (Ca++, C) and inorganic phosphorus (Pi, D) as a function of time after subcutaneously (SC) injection of peptide or vehicle; p < 0.05 (2-way ANOVA, followed by LSD test): *, peptide vs. vehicle; +, PTH(1-34) vs. Palm-PTH(1-34). E Peptide concentrations in blood plasma as a function of time after SC injection. The inset shows the data plotted with the y-axis in log10 scale. Data for x = 1–24 h were fit to a monophasic decay curve (dashed lines); the corresponding values of T1/2, AUC, Cmax (peak y), and Tmax (peak x) are shown. F Concentrations of tetramethylrhodamine (tmr)-labeled ligands in urine as a function time after SC injection of PTH(1-34)tmr or Palm-PTH(1-34)tmr; tmr fluorescence (lex = 531 ± 25 nm, lem = 595 ± 60 nm) at each time point was corrected for background fluorescence (t = 0) and net fluorescence was converted to peptide concentration using a standard curve formed with the corresponding tmr-peptide; p < 0.05 (two-sided t-test): *, PTH(1-34) vs. Palm-PTH(1-34). The inset shows the data plotted with the x-axis extended to 8 h; corresponding values of AUC (0-24 h) and Tmax (peak x) are also shown. In C-F, t = 0 represents pre-dose. Data are means ± SEM of values from six (C, D), three (E) or five (F) mice per group. Source data with exact p values are provided in the Source Data File. Corresponding changes in blood Ca++ and plasma peptide concentrations after IV injection are shown in Supplementary Fig. S-2.
