FGF23 neutralization improves chronic kidney disease-associated hyperparathyroidism yet increases mortality - PubMed (original) (raw)
. 2012 Jul;122(7):2543-53.
doi: 10.1172/JCI61405. Epub 2012 Jun 25.
Edward M Shatzen, Sabrina C Ward, James Davis, Jennitte Stevens, Vivian Bi, Lisa Renshaw, Nessa Hawkins, Wei Wang, Ching Chen, Mei-Mei Tsai, Russell C Cattley, Thomas J Wronski, Xuechen Xia, Xiaodong Li, Charles Henley, Michael Eschenberg, William G Richards
Affiliations
- PMID: 22728934
- PMCID: PMC3386816
- DOI: 10.1172/JCI61405
FGF23 neutralization improves chronic kidney disease-associated hyperparathyroidism yet increases mortality
Victoria Shalhoub et al. J Clin Invest. 2012 Jul.
Abstract
Chronic kidney disease-mineral and bone disorder (CKD-MBD) is associated with secondary hyperparathyroidism (HPT) and serum elevations in the phosphaturic hormone FGF23, which may be maladaptive and lead to increased morbidity and mortality. To determine the role of FGF23 in the pathogenesis of CKD-MBD and development of secondary HPT, we developed a monoclonal FGF23 antibody to evaluate the impact of chronic FGF23 neutralization on CKD-MBD, secondary HPT, and associated comorbidities in a rat model of CKD-MBD. CKD-MBD rats fed a high-phosphate diet were treated with low or high doses of FGF23-Ab or an isotype control antibody. Neutralization of FGF23 led to sustained reductions in secondary HPT, including decreased parathyroid hormone, increased vitamin D, increased serum calcium, and normalization of bone markers such as cancellous bone volume, trabecular number, osteoblast surface, osteoid surface, and bone-formation rate. In addition, we observed dose-dependent increases in serum phosphate and aortic calcification associated with increased risk of mortality in CKD-MBD rats treated with FGF23-Ab. Thus, mineral disturbances caused by neutralization of FGF23 limited the efficacy of FGF23-Ab and likely contributed to the increased mortality observed in this CKD-MBD rat model.
Figures
Figure 1. Characterization of FGF23-Abs in normal rodents.
(A) Rat FGF23 dose-response in CHO–Kl–ELK-1 luciferase reporter cells. §P < 0. 001 versus CHO–ELK-1 and parental CHO cells and FGF23, ANOVA plus Tukey’s post test. (B) Rat anti-rat FGF23 hybridoma-conditioned medium inhibited FGF23 binding to FGFR-Kl. Each bar represents the mean result from duplicate wells. Blank, no FGF23 control. (C) FGF23-Ab inhibited binding to FGFRs 1c, 3c, and 4. (D) FGF23-Ab inhibited FGF23 ELK-1 signaling in CHO-Kl cells. *P < 0.05, **P < 0.01 versus FGF23-Ab; #P < 0.05, ##P < 0.01 versus FGF23-Ab, t test. (E) A single FGF23-Ab injection increased serum phosphate in normal mice. Balb/c male mice were injected with FGF23-Ab, control Ab or PBS vehicle (n = 3 PBS, n = 7 control Ab, n = 10 FGF23-Ab; mean ± SEM; *P < 0. 05 versus PBS or control Ab, ANOVA plus Tukey’s post-test). (F) A single FGF23-Ab injection increased serum phosphate in normal rats. Serum phosphate was measured daily for 4 days. (n = 4/group; mean ± SEM; **P < 0.01, §P < 0.001, §§P < 0.0001 versus respective day control Ab, unpaired t test. ###P < 0.001 versus baseline, ANOVA plus Dunnett’s multiple comparison test). BL, baseline (time 0) before each treatment was administered.
Figure 2. Survival plot demonstrates higher mortality in the high-dose FGF23-Ab group.
Started with normal + control Ab, ND (n = 10); sham-treated + control Ab, HPD (n = 12); 5/6Nx + control Ab, HPD (n = 16); 5/6Nx + FGF23-Ab (3 mg/kg), HPD (n = 16); 5/6Nx + FGF23-Ab (10 mg/kg), HPD (n = 16). Note that the normal + control Ab, ND and the sham-treated + control Ab group results are superimposable at 100% survival throughout the 6-week treatment period.
Figure 3. Effect of FGF23-Ab on renal function, serum FGF23, and PTH levels.
(A) GFR was estimated using the formula [(Ucr × 24-h vol)/Scr, where Ucr = urinary creatinine, Scr = serum creatinine, and 24-h vol = 24-hour urinary volume]. (B) Serum BUN levels. (C) Serum FGF23 levels. FGF23 levels were undetectable in treated rat serum, likely due to a common epitope for FGF23-Ab and an Ab in the ELISA. (D) Serum PTH levels. *P < 0.05, **P < 0.01, §P < 0.001, §§P < 0.0001 for 5/6Nx + control Ab versus sham-treated + control Ab; #P < 0.05, ##P < 0.01, ‡P ≤ 0.0001 for 5/6Nx + FGF23-Ab (10 mg/kg or 3 mg/kg) versus 5/6Nx + control Ab; ††P < 0.0001 for 5/6Nx groups versus sham-treated group at baseline. Data points represent mean ± SEM derived from the following experimental groups: normal + control Ab, ND (n = 10); sham-treated + control Ab, HPD (n = 12); 5/6Nx + control Ab, HPD (n = 14); 5/6Nx + FGF23-Ab (3 mg/kg), HPD (n = 14); 5/6Nx + FGF23-Ab (10 mg/kg), HPD (n = 9).
Figure 4. Treatment of 5/6 nephrectomized rats with FGF23-Ab increased serum phosphate, calcium, and 1,25(OH)2D3 levels.
(A) Serum phosphate levels. (B) Fractional excretion of phosphate, FEPi. (C) Serum total calcium levels. (D) Urinary calcium. (E) Serum Vitamin D (1,25[OH]2D3) levels. *P < 0.05, **P < 0.01, §§P < 0.0001 for 5/6Nx + control Ab versus sham-treated plus control Ab; #P < 0.05, ##P < 0.01, ‡P < 0.0001 for 5/6Nx plus FGF23-Ab (10 mg/kg or 3 mg/kg) versus 5/6Nx + control Ab; †P < 0.05, ††P < 0.0001 for 5/6Nx groups versus sham-treated group at baseline. For urinary calcium, †P < 0.05 for 5/6Nx low–dose group versus sham-treated group at baseline. Data points represent mean ± SEM.
Figure 5. FGF23-Ab treatment did not significantly affect expression of markers of kidney pathology.
(A) Cyp27b1 (1α[OH]ase). (B) Cyp24a1 (24-[OH]ase). (C) Slc34a1 (NaPi2a). (D) Klotho. Transcript levels were normalized to mRNA levels for Gapdh, Hprt1, Ppib in the same sample. ***P < 0.001 for comparisons as designated on figures. Animal numbers in each group are the same as those described in Figure 3, except for 5/6Nx + FGF23-Ab (10 mg/kg), HPD (n = 8 instead of 9). Data represent mean ± SEM.
Figure 6. FGF23-Ab treatment had no effect on increased expression of cardiac hypertrophy markers.
(A) Acta1. (B) Acta2. (C) Myh7. (D) Nppb. Transcript levels were normalized to mRNA levels for Gapdh, Hprt1, or Ppib in the same sample. *P < 0.05, **P < 0.01 for comparisons as designated on figures. Animal numbers in each group are the same as those described in Figure 5. Data represent mean ± SEM.
Figure 7. FGF23-Ab increases aortic calcification in 5/6Nx rats on a high-phosphate diet.
(A) Aortic calcification score was higher for FGF23-Ab–treated 5/6Nx rats after 6 weeks of treatment. Semiquantitative mean calcification score: none, 0; minimal, 1; mild, 2; moderate, 3; marked, 4; severe, 5. Overall comparison for CKD groups: P < 0. 0001 Kruskal-Wallis test. (A) **P = 0.0056, ***P = 0.0006, ****P = 0.0001, Wilcoxon’s exact test; (B) Serum phosphorus, ****P < 0.0001. (C) Serum calcium. No significance between sham-treated and/or 5/6Nx groups. Animal numbers in each group are the same as those described in Figure 5. Results are presented for individual animals and mean ± SEM.
Figure 8. Histology of rat proximal tibia showing increased bone/osteoid accumulation in 5/6Nx control animals and improvement upon FGF23-Ab treatment.
Bone sections were subjected to von Kossa and tetrachrome staining. (A) Normal plus control Ab; (B) sham-treated plus control Ab; (C) 5/6Nx plus control Ab. (D) 5/6Nx + FGF23-Ab (10 mg/kg). Original magnification, ×40.
Comment in
- Fibroblast growth factor 23: friend or foe in uremia?
Moe OW. Moe OW. J Clin Invest. 2012 Jul;122(7):2354-6. doi: 10.1172/JCI64184. Epub 2012 Jun 25. J Clin Invest. 2012. PMID: 22728930 Free PMC article.