The need for reliable serum parathyroid hormone measurements
2006; Elsevier BV; Volume: 70; Issue: 2 Linguagem: Inglês
10.1038/sj.ki.5001658
ISSN1523-1755
Autores Tópico(s)Bone health and osteoporosis research
ResumoSerum parathyroid hormone (PTH) is a recognized marker of bone remodeling in patients with renal osteodystrophy. However, identification of N-terminal truncated PTH fragments and a new form of PTH that interfere with second-generation PTH assays may be responsible for the great variability of PTH values and the difficulties of implementing the recommendations of the National Kidney Foundation/Kidney Disease Outcomes Quality Initiative. Serum parathyroid hormone (PTH) is a recognized marker of bone remodeling in patients with renal osteodystrophy. However, identification of N-terminal truncated PTH fragments and a new form of PTH that interfere with second-generation PTH assays may be responsible for the great variability of PTH values and the difficulties of implementing the recommendations of the National Kidney Foundation/Kidney Disease Outcomes Quality Initiative. Parathyroid hormone (PTH) is a polypeptide molecule of 84 amino acids with a molecular weight of 9500 daltons. It is synthesized by and secreted from the chief epithelial cells of parathyroid glands. Developmentally, there are four orthotopic parathyroid glands derived from the pharyngeal pouch; however, supernumerary, scanty, and/or ectopic parathyroid glands can be seen in a small percentage of subjects.1.Hindié E. Ureña P. Jeanguillaume C. et al.Preoperative imaging of parathyroid glands with 99m-labelled sestamibi and iodine-123 subtraction scanning in secondary hyperparathyroidism.Lancet. 1999; 2: 2200-2204Abstract Full Text Full Text PDF Scopus (62) Google Scholar The formation of these glands is under the control of a key regulatory gene coding for the transcription factor Gcm-2 (glial cell missing), which is almost exclusively expressed in these cells.2.Günther T. Chen Z. Kim J. et al.Genetic ablation of parathyroid glands reveals another source of parathyroid hormone.Nature. 2000; 406: 199-203Crossref PubMed Scopus (287) Google Scholar PTH is synthesized as a 115-amino acid precursor molecule called pre-pro-PTH, which is encoded by a single gene located on the short arm of chromosome 11 (11p15). This gene has three exons separated by two introns; the first exon contains most of the 5′ non-coding region of the gene, and the second exon codes for most of the pre-pro-PTH sequence. The third exon codes for three regions: the cleavage site Lys-Arg, the mature 1–84 PTH sequence, and the untranslated 3′ region.3.Hendy G.N. Kronenberg H.M. Potts Jr, J.T. Rich A. Nucleotide sequence of cloned cDNAs encoding human preproparathyroid hormone.Proc Natl Acad Sci USA. 1981; 78: 7365-7369Crossref PubMed Scopus (132) Google Scholar The mature PTH is stocked in secretory granules that fuse with the cellular membrane and release PTH in response to reduction in the extracellular ionized-calcium concentration. Most of this PTH is secreted in its intact form, 1–84; however, it can also be secreted as N-terminal truncated fragments or C-terminal fragments after intracellular degradation, as in case of hypercalcemia.4.D'Amour P. Räkel A. Brossard J.H. et al.Acute regulation of circulating parathyroid hormone (PTH) molecular forms by calcium: utility of PTH fragments/PTH (1-84) ratios derived from three generations of PTH assays.J Clin Endocrinol Metab. 2006; 91: 283-289Crossref PubMed Scopus (41) Google Scholar No PTH fragment has been found to be the result of another gene or alternative splicing of the PTH gene. Normally, on the basis of the percentage of the total immunoreactivity detected by different PTH assays, sometimes validated by the evaluation of PTH molecular-form concentrations derived from high-performance liquid chromatography profiles,4.D'Amour P. Räkel A. Brossard J.H. et al.Acute regulation of circulating parathyroid hormone (PTH) molecular forms by calcium: utility of PTH fragments/PTH (1-84) ratios derived from three generations of PTH assays.J Clin Endocrinol Metab. 2006; 91: 283-289Crossref PubMed Scopus (41) Google Scholar approximately 10%–20% of the total circulating PTH molecules are in its intact and bioactive form (1–84) (Table 1). Indeed, once the secreted 1–84 PTH reaches the circulation, it is rapidly taken up by the liver and the kidney and metabolized by several endoproteases in two main fragments: N-terminal and C-terminal fragments (the half-life of the entire molecule varies between 2 and 5 minutes). In the liver, all intact and N-terminal fragments are metabolized, and no N-terminal fragment is released. This degradation gives rise to several C-terminal fragments, starting from residues at positions 34, 37, 41, and 43 and ending probably at position 84 or shorter, which are often released back into the circulation.5.Habener J.F. Mayer G.P. Dee P.C. Potts Jr, J.T. Metabolism of amino- and carboxyl-sequence immunoreactive parathyroid hormone in the bovine: evidence for peripheral cleavage of the hormone.Metabolism. 1976; 25: 385-395Abstract Full Text PDF PubMed Scopus (30) Google Scholar, 6.Murray T.M. Rao L.G. Divieti P. Bringhurst F.R. Parathyroid hormone secretion and action: evidence for discrete receptors for the carboxyl-terminal region and related biological actions of carboxyl-terminal ligands.Endocr Rev. 2005; 26: 78-113Crossref PubMed Scopus (214) Google Scholar Quantitatively, less than 20% of the intact PTH is converted into C-terminal fragments by the liver. However, because of their longer half-life (five to ten times longer than that of the intact form) and because they are normally cleared by the kidneys, those C-terminal PTH fragments represent 80% of the total circulating PTH molecules in normal individuals (Table 1). The kidney is the second organ where PTH is catabolized. After crossing the glomerular filtration membrane, all PTH molecules are actively reabsorbed by the proximal tubular cells through two mechanisms: binding to its specific receptor (PTHR1) and internalization through the megalin/cubulin endocytosis system.7.Hilpert J. Nikjaer A. Jacobsen C. et al.Megaline antagonizes activation of the parathyroid hormone receptor.J Biol Chem. 1999; 274: 5620-5625Crossref PubMed Scopus (88) Google Scholar Thus, no detectable PTH fragments can be found in the final urine.Table 1Approximate estimation of the percentage of the total immunoreactivity detected by three different parathyroid hormone (PTH) assays, before and after validation by the evaluation of PTH molecular-form concentrations derived from high-performance liquid chromatography profiles 4.D'Amour P. Räkel A. Brossard J.H. et al.Acute regulation of circulating parathyroid hormone (PTH) molecular forms by calcium: utility of PTH fragments/PTH (1-84) ratios derived from three generations of PTH assays.J Clin Endocrinol Metab. 2006; 91: 283-289Crossref PubMed Scopus (41) Google Scholar,15.D'Amour P. Brossard J.H. Rousseau L. et al.Aminoterminal form of parathyroid hormone (PTH) with immunologic similarities to hPTH (1-84) is overproduced in primary and secondary hyperparathyroidism.Clin Chem. 2003; 49: 2037-2044Crossref PubMed Scopus (88) Google ScholarIntact 1–84 PTH (%)C-terminal PTH fragments (%)Normal subjects208014.0 of intact 1–84 PTH, 4.4 of N-terminal truncated PTH, 1.6 of amino-PTHChronic renal failure5952.5 of intact 1–84 PTH, 1.8 of N-terminal truncated PTH, 0.7 of amino-PTH Open table in a new tab Because the kidneys play an important role in the catabolism of intact PTH and also of its fragments, when they fail, there is a tendency toward the accumulation of circulating PTH fragments. However, this accumulation seems to be disproportionate to the degree of renal insufficiency and could be the result of two phenomena: the renal clearance of C-terminal fragments is sensibly lower than that of intact 1–84 PTH, and secondly, it is now recognized that parathyroid glands can also release C-terminal fragments, namely 7–84 PTH,6.Murray T.M. Rao L.G. Divieti P. Bringhurst F.R. Parathyroid hormone secretion and action: evidence for discrete receptors for the carboxyl-terminal region and related biological actions of carboxyl-terminal ligands.Endocr Rev. 2005; 26: 78-113Crossref PubMed Scopus (214) Google Scholar thereby increasing further the burden of these fragments. The skeletal tissue is another organ that can also clear a small fraction of circulating intact PTH molecules though a receptor-mediated process.8.Rouleau M.F. Warshawsky H. Goldzman D. Parathyroid hormone binding in vivo to renal, hepatic, and skeletal tissues of the rat using a adioautographic approach.Endocrinology. 1986; 118: 919-931Crossref PubMed Scopus (89) Google Scholar Chronic kidney disease is often associated with skeletal and mineral disorders globally described as renal osteodystrophy (ROD), ranging from high-turnover bone diseases such as secondary hyperparathyroidism, with its extreme and severe manifestation osteitis fibrosa, to low-turnover bone diseases such as osteomalacia and adynamic bone disease.9.deVernejoul M.C. Kuntz D. Miravet L. et al.Bone histomorphometry in hemodialysed patients.Metab Bone Dis Relat Res. 1981; 3: 175-179Abstract Full Text PDF PubMed Scopus (38) Google Scholar The gold-standard method to make the diagnosis of the type of ROD is still the histomorphometric analysis of iliac crest bone biopsies obtained after double tetracycline labeling.9.deVernejoul M.C. Kuntz D. Miravet L. et al.Bone histomorphometry in hemodialysed patients.Metab Bone Dis Relat Res. 1981; 3: 175-179Abstract Full Text PDF PubMed Scopus (38) Google Scholar However, this invasive and complicated method cannot be used in daily practice, so alternative methods must be used. Because the circulating concentration of PTH is well correlated with the degree of bone turnover, it has been largely used as a surrogate of bone biopsy, and nephrologists have been searching for decades for a better and more reliable method to measure it. The first generation of PTH assays were developed and used from 1959 to 1987. They used a single antibody directed against PTH (epitopes on amino acids 39–84 or 53–84) in the radioimmunoassay and measured many blood-borne fragments of PTH along with 1–84 PTH.10.Berson S.A. Yalow R.S. Immunochemical heterogeneity of parathyroid hormone in plasma.J Clin Endocrinol Metab. 1968; 28: 1037-1047Crossref PubMed Scopus (265) Google Scholar The second-generation PTH assays, called 'intact' PTH assays, have been developed since 1987.11.Nussbaum S. Zahradnik R. Lavigne J. et al.Highly sensitive two-site immunoradiometric assay of parathyrin, and its clinical utility in evaluating patients with hypercalcemia.Clin Chem. 1987; 33: 1364-1367PubMed Google Scholar They use two different antibodies; the first, directed against the C-terminal region of PTH (epitopes on amino acids 39–84, 44–84, 55–64, or 53–84), is often immobilized onto plastic surfaces and binds intact 1–84 PTH and C-terminal PTH fragments. The second antibody is directed against the N-terminal region (epitopes on amino acids 13–34, the 'proximal epitope,' as in the Allegro assay from Nichols Institute Diagnostics; or 26–32, the 'distal epitope,' as in the Elecsys assay from Roche Diagnostics), and it is often radiolabeled, biotinylated, bound to an enzyme to allow its easy detection and quantification, or served as the coated antibody. These 'sandwich' assays were the first radioimmunometric assays, and they were thought to measure only the full-length 1–84 PTH; however, they also measure large PTH fragments (namely 7–84 PTH).12.Brossard J. Cloutier M. Roy L. et al.Accumulation of a non-(1-84) molecular form of parathyroid hormone (PTH) detected by intact PTH assay in renal failure: importance in the interpretation of PTH values.J Clin Endocrinol Metab. 1996; 81: 3923-3929Crossref PubMed Scopus (264) Google Scholar These N-terminal truncated PTH fragments have now been demonstrated to inhibit the action of PTH by blocking its binding to its normal receptor, PTHR1. They may also bind to a specific C-terminal PTH receptor and may have multiple not-yet-known biological functions in the skin, bone, hematopoietic system, and placenta.16.Gao P. Scheibel S. D'Amour P. et al.Development of a novel immunoradiometric assay exclusively for biologically active whole parathyroid hormone 1-84: implication for improvement of accurate assessment of parathyroid function.J Bone Miner Res. 2001; 16: 605-614Crossref PubMed Scopus (312) Google Scholar, 13.Murray T.M. Rao L.G. Muzaffar S.A. Ly H. Human parathyroid hormone carboxyterminal peptide (53-84) stimulates alkaline phosphatase activity in dexamethasone-treated rat osteosarcoma cells in vitro.Endocrinology. 1989; 124: 1097-1099Crossref PubMed Scopus (75) Google Scholar, 14.Kaji H. Sugimoto T. Kanatani M. et al.Carboxy-terminal parathyroid hormone fragments stimulate osteoclastic-like cell formation and osteoclastic activity.Endocrinology. 1994; 134: 1897-1904Crossref PubMed Scopus (67) Google Scholar The third generation of PTH assays have been developed since 2000; they use the same sandwich and radioimmunometric techniques, with the first antibody directed against amino acids 39–84, but the second antibody has been restricted and directed against the first six N-terminal residues of 1–84 PTH (epitopes on amino acids 1–4 or 1–5). They have been demonstrated to be most sensitive, and more specific, when measuring bioactive intact 1–84 PTH, and probably another new form of PTH named amino-PTH, recently identified in patients with parathyroid carcinoma and with severe cases of primary and secondary hyperparathyroidism.15.D'Amour P. Brossard J.H. Rousseau L. et al.Aminoterminal form of parathyroid hormone (PTH) with immunologic similarities to hPTH (1-84) is overproduced in primary and secondary hyperparathyroidism.Clin Chem. 2003; 49: 2037-2044Crossref PubMed Scopus (88) Google Scholar From the beginning, the utility of the third-generation PTH assays has generated a highly charged debate. And it should be emphasized here that the economic stakes of this debate should lead clinicians to assess all claims carefully. Many of the studies published found an excellent correlation of serum PTH values between the second- and third-generation PTH assays16.Gao P. Scheibel S. D'Amour P. et al.Development of a novel immunoradiometric assay exclusively for biologically active whole parathyroid hormone 1-84: implication for improvement of accurate assessment of parathyroid function.J Bone Miner Res. 2001; 16: 605-614Crossref PubMed Scopus (312) Google Scholar, 17.John M. Goodman W. Gao P. et al.A novel immunoradiometric assay detects full-length human PTH but not amino-terminally truncated fragments: implications for PTH measurements in renal failure.J Clin Endocrinol Metab. 1999; 84: 4287-4290Crossref PubMed Google Scholar, 18.Slatopolsky E. Finch J. Clay P. et al.A novel mechanism for skeletal resistance in uremia.Kidney Int. 2000; 58: 753-761Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar in chronic kidney disease patients undergoing dialysis. However, mean circulating PTH levels are 30%–60% lower with the third-generation PTH assays than with the second-generation assays. Furthermore, most uremic patients exhibit a normal ratio of intact to N-terminal truncated PTH fragments (intact PTH/N-terminal truncated PTH – intact PTH); however, in certain cases, this ratio can be abnormal, which has led several investigators to propose that a ratio lower than 1 could be the best predictor of a low bone-turnover disease.19.Monier-Faugère M.C. Geng Z. Mawad H. et al.Improved assessment of bone turnover by the PTH (1-84)/C-PTH fragments ratio in ESRD patients.Kidney Int. 2001; 60: 460-468Abstract Full Text Full Text PDF Scopus (229) Google Scholar Other investigators have been unable to confirm these findings.20.Reichel H. Esser A. Roth H.J. Schmidt-Gayk H. Influence of PTH assay methodology on differential diagnosis of renal bone disease.Nephrol Dial Transplant. 2003; 18: 759-768Crossref PubMed Scopus (75) Google Scholar Souberbielle et al.21.Souberbielle J.-C. Boutten A. Carlier M.-C. et al.Inter-method variability in PTH measurement: implication for the care of CKD patients.Kidney Int. 2006; 70: 345-350Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar (this issue) have elegantly looked at the inter-method variability in the measurement of circulating PTH concentration of 15 different PTH assays (13 second-generation and two third-generation PTH assays) in 47 pools of serum samples from a French population of dialyzed patients. They also looked at the recovery of graded amounts of synthetic full-length 1–84 PTH and N-terminal truncated 7–84 PTH added separately to a normal serum pool. They demonstrate that, when the second-generation Allegro intact PTH assay (Nichols Institute Diagnostics) was used as the reference assay, the median bias between the different PTH assays varied from -44.9% to +123.0% (Figure 1). This implies that we may be under- or overestimating the true serum PTH concentration depending on the type of PTH assay we have chosen. A patient with a serum PTH value of 150 pg/ml or 300 pg/ml in one place may have values between 83 and 323 pg/ml and between 160 and 638 pg/ml, respectively, in another place. This clearly illustrates that the method used to measure circulating PTH concentration is a very important clinical, economic, and ethical issue. The most striking example is that one would never indicate a surgical parathyroidectomy or start aggressively and expensively to treat secondary hyperparathyroidism in a patient with serum PTH levels lower than 300 pg/ml, unless the laboratory inaccurately reported a value above 600 pg/ml because of the use of an unreliable method. Indeed, this variability in the PTH measurement may explain the drift and rise in the incidence of surgical parathyroidectomy in the United States between 1998 and 2002.22.Foley R.N. Li S. Liu J. et al.The fall and rise of parathyroidectomy in U.S. hemodialysis patients, 1992 to 2002.J Am Soc Nephrol. 2005; 16: 210-218Crossref PubMed Scopus (102) Google Scholar At that time, Nichols Institute intended to replace the second-generation Allegro intact PTH assay with the automated Intact PTH Advantage, already introduced in the market in 1992; however, it was not well calibrated or shifted upward such that it gave greater values for serum PTH concentration. The economic impact is huge, not only because we might be overtreating our patients with expensive drugs, but also because the frequency of PTH determinations in uremic patients has dramatically increased during the last 7 years, mainly because of the need for a closer monitoring of serum PTH levels in patients treated by calcimimetics, and because of the recommendations given by the Kidney Disease Outcomes Quality Initiative of the National Kidney Foundation (NKF/K-DOQI) since 1998. Thus, this market has exploded and can be easily estimated to exceed several hundred million dollars per year worldwide. It appears unethical that all the companies making different PTH assays could not meet and decide how to standardize and calibrate a consistent circulating PTH measurement. Many physicians are making huge efforts to keep their patients within the target values recommended by the NKF/K-DOQI; however, most of them are misinformed by their PTH measurement. It should also be stressed here that the bone response to a given circulating PTH concentration in uremic patients is very variable; it depends on the level of PTH receptor expression and function,23.Ureña P. Kubrusly M. Mannstadt M. et al.The renal PTH/PTHrP receptor is down-regulated in rats with chronic renal failure.Kidney Int. 1994; 45: 605-611Abstract Full Text PDF PubMed Scopus (148) Google Scholar, 24.Ureña P. Ferreira A. Morieux C. et al.PTH/PTHrP receptor mRNA is down-regulated in epiphyseal cartilage growth plate of uraemic rats.Nephrol Dial Transplant. 1996; 11: 2008-2016Crossref PubMed Scopus (83) Google Scholar and/or on the existence of a specific C-terminal PTH receptor.6.Murray T.M. Rao L.G. Divieti P. Bringhurst F.R. Parathyroid hormone secretion and action: evidence for discrete receptors for the carboxyl-terminal region and related biological actions of carboxyl-terminal ligands.Endocr Rev. 2005; 26: 78-113Crossref PubMed Scopus (214) Google Scholar For the moment, this variability limits any attempt to predict the type of ROD with only the measurement of whatever type of PTH assay — intact, N-terminal truncated PTH fragments or C-terminal PTH fragments. However, as these naturally occurring PTH fragments may also exhibit biological actions, it is quite conceivable that in the future their independent measurement may increase the diagnostic sensitivity of the type of ROD. The most appropriate strategy would be to combine these PTH measurements with specific biochemical markers of bone formation rate — such as bone-specific alkaline phosphatase — and of bone resorption rate — such as collagen type I breakdown cross-linked peptides (cross-laps) — in order to gather more information about the bone response to a given serum PTH concentration.25.Ureña P. Ferreira A. Kung V. et al.Serum pyridinoline as a specific marker of collagen breakdown and bone metabolism in hemodialysis patients.J Bone Miner Res. 1995; 10: 932-939Crossref PubMed Scopus (90) Google Scholar, 26.Ureña P. Hruby M. Ferreira A. et al.Plasma total versus bone alkaline phosphatase as marker of bone turnover in hemodialysis patients.J Am Soc Nephrol. 1996; 7: 506-512PubMed Google Scholar The combination of these measures might improve the prediction of the type of ROD. However, this kind of study remains to be performed. Finally, until there is a generalized use of a unique, true, and accurate intact 1–84 PTH assay, it would be worthwhile for current PTH assays to be calibrated against a validated international standard of synthetic or recombinant 1–84 PTH. Furthermore, it is the responsibility of all companies fabricating second-generation PTH assays to provide a way to correct all results obtained. These values could be corrected, as proposed by Souberbielle et al.,21.Souberbielle J.-C. Boutten A. Carlier M.-C. et al.Inter-method variability in PTH measurement: implication for the care of CKD patients.Kidney Int. 2006; 70: 345-350Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar by a specific factor such as the inverse of the slope reported in Table 2 of their article, or the inverse of the bias presented in Table 3 of their article. Ultimately, even if these measures are taken, it will be necessary to re-evaluate the impact of new intact 1–84 PTH values on the NKF/K-DOQI recommendations and on the morbidity and mortality risks associated with high serum intact PTH levels as measured with the second-generation PTH assays.
Referência(s)