Labordiagnostik bei Wachstumshormon-assoziierten Erkrankungen/Biochemical diagnosis of growth hormone related diseases

The symptoms of growth hormone deficiency and growth hormone excess manifest themselves clinically in different ways, before and after the completion of longitudinal growth. Always, however, biochemical diagnostics is based on the measurement of circulating concentrations of growth hormone and insulin-like growth factor I (IGF-I). Immunoassays are practical, sensitive, and mostly specific methods for measuring either hormone. Still, there are serious discrepancies between the measured results of different assays. These discrepancies are mainly due to differences in the isoform specificity of assays, the use of different standard preparations, as well as the interference of binding proteins. The method-related differences in measured results make the general application of published diagnostic decision limits more difficult. At an interdisciplinary consensus conference, with the participation of the Growth Hormone Research Society, the IGF Society, and the International Federal of Clinical Chemistry and Laboratory Medicine (IFCC), the existing problems were analyzed and possible strategies were highlighted to improve the comparability of the measured results of different GH and IFG-I assays. Currently, however, the use of method-specific reference ranges obtained from well-characterized cohorts continues to be essential in clinical practice.Die Krankheitsbilder des Wachstumshormonmangels und des Wachstumshormonexzesses imponieren klinisch unterschiedlich vor und nach dem Abschluss des Längenwachstums. Stets jedoch bilden die Messung der zirkulierenden Konzentrationen von Wachstumshormon und Insulinartigem Wachstumsfaktor I (Insulin-like growth-factor I, IGF-I) die Basis der laborchemischen Diagnostik. Mit den Immunoassays stehen praktikable, sensitive und meist auch spezifische Methoden zur Messung beider Hormone zur Verfügung. Trotzdem bestehen nach wie vor gravierende Diskrepanzen zwischen den Messergebnissen verschiedener Assays. Diese Diskrepanzen sind vor allem bedingt durch Unterschiede in der Isoformspezifität der Assays, die Verwendung unterschiedlicher Standardpräparationen sowie die Interferenz von Bindungsproteinen. Die methodenbedingten Unterschiede in den Messergebnissen erschweren die allgemeine Anwendung publizierter diagnostischer Entscheidungsgrenzen. Auf einer interdisziplinären Konsensuskonferenz unter Beteiligung der Growth Hormone Research Society, der IGF Society, und der International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) wurden die bestehenden Probleme analysiert und mögliche Strategien zu einer Verbesserung der Vergleichbarkeit der Messergebnisse verschiedener GH- und IGF-I-Assays aufgezeigt. Derzeit bleibt jedoch in der klinischen Praxis die Anwendung methodenspezifischer, an gut charakterisierten Kollektiven gewonnener Referenzbereiche unabdingba

Quantification of growth hormone in serum by isotope dilution mass spectrometry

Inter-assay variation of antibody based routine tests is hampering comparability of measurement results for growth hormone (GH) between different laboratories and decision making in clinical practice. Here it is demonstrated, that quantification of GH by isotope dilution mass spectrometry (IDMS) constitutes a way to precise and reliable results which can be referred to in evaluation of performance of commercial test kits. With the IDMS method developed, tryptic cleavage products YSFLQNPQTSLCFSESIPTPSNR (T6) and LEDGSPR (T12) of GH are quantified by LC/MS-MS using the isotopically labeled forms of the peptides as internal standards. The GH cleavage fragments are obtained by whole-serum tryptic proteolysis and then extracted from the resulting mixture by semi-preparative reversed phase liquid chromatography followed by strong cation-exchange chromatography. Method validation basing on recovery of recombinant 22 kDa GH spiked to blank serum in defined amounts covering the intended concentration range (3-30 µg/L) would yield mean recoveries of 101.6% (100.7%), standard deviations of 2.5% (2.4%) and combined uncertainties (_u~c~_) of 3.0% (2.5%) if quantifying T6 (T12) as GH derived fragments, while the LOQ were 1.7 µg/L (2.7 µg/L). Potential to acquisition of reference values is exemplified by application to serum materials used in a recent quality assessment exercise for routine laboratories

Interpreting growth hormone and IGF-I results using modern assays and reference ranges for the monitoring of treatment effectiveness in acromegaly

Standard treatment for acromegaly focuses on the achievement of target absolute levels of growth hormone (GH) and insulin-like growth factor (IGF-I). The appropriateness of these targets when measured using modern assay methods is not well defined. This paper reviews biochemical status assessed using methods available at the time and associated clinical outcomes. GH measurements were shown to provide an indication of changes in tumor size, and failure of GH suppression after glucose stimulation is associated with tumor recurrence. IGF-I levels were more closely associated with changes in symptoms and signs. Reduced GH and IGF-I concentrations were shown to be associated with increased longevity, although the degree of increase has only been analyzed for GH. Lowering of GH and IGF-I has consistently been associated with improved outcomes; however, absolute levels reported in previous studies were based on results from methods and reference ranges that are now obsolete. Applying previously described absolute thresholds as targets (e.g. “normal” IGF-I level) when using current methods are best applied to those with active acromegaly symptoms who could benefit from further lowering of biochemical markers. In asymptomatic individuals with mild IGF-I or GH elevations, targeting biochemical “normalization” would result in the need for combination pharmacotherapy in many patients without proven benefit. Measurement of both GH and IGF-I remains an essential component of diagnosis and monitoring the effectiveness of treatment in acromegaly; however, treatment goals based only on previously identified absolute thresholds are not appropriate without taking into account the assay and reference ranges being employed. Treatment goals should be individualized considering biochemical improvement from an untreated baseline, symptoms of disease, risks, burdens and costs of complex treatment regimens, comorbidities, and quality of life

Interpreting growth hormone and IGF-I results using modern assays and reference ranges for the monitoring of treatment effectiveness in acromegaly

Standard treatment for acromegaly focuses on the achievement of target absolute levels of growth hormone (GH) and insulin-like growth factor (IGF-I). The appropriateness of these targets when measured using modern assay methods is not well defined. This paper reviews biochemical status assessed using methods available at the time and associated clinical outcomes. GH measurements were shown to provide an indication of changes in tumor size, and failure of GH suppression after glucose stimulation is associated with tumor recurrence. IGF-I levels were more closely associated with changes in symptoms and signs. Reduced GH and IGF-I concentrations were shown to be associated with increased longevity, although the degree of increase has only been analyzed for GH. Lowering of GH and IGF-I has consistently been associated with improved outcomes; however, absolute levels reported in previous studies were based on results from methods and reference ranges that are now obsolete. Applying previously described absolute thresholds as targets (e.g. “normal” IGF-I level) when using current methods is best applied to those with active acromegaly symptoms who could benefit from further lowering of biochemical markers. In asymptomatic individuals with mild IGF-I or GH elevations, targeting biochemical “normalization” would result in the need for combination pharmacotherapy in many patients without proven benefit. Measurement of both GH and IGF-I remains an essential component of diagnosis and monitoring the effectiveness of treatment in acromegaly; however, treatment goals based only on previously identified absolute thresholds are not appropriate without taking into account the assay and reference ranges being employed. Treatment goals should be individualized considering biochemical improvement from an untreated baseline, symptoms of disease, risks, burdens and costs of complex treatment regimens, comorbidities, and quality of life

Harmonization of growth hormone measurements with different immunoassays by data adjustment

Background: The aim of our study was to evaluate the between-assay variability of commercially available immunoassays for the measurement of human growth hormone (hGH). In addition, we asked whether the comparability of the diagnosis of childhood onset growth hormone deficiency could be improved by adjusting hGH results by statistical methods, such as linear regression, conversion factors, and quantile transformation. Methods: In archived sera from 312 children and adolescents (age: 17 days-17 years) hGH values between 0.01 and 16.5 ng/mL were determined by using the following immunoassays: AutoDELFIA (PerkinElmer), BC-IRMA (Beckman-Coulter), ELISA (Mediagnost), IMMULITE 2000 (Siemens), iSYS (IDS), Liaison (DiaSorin), UniCel DxI 800 Access (BeckmanCoulter) and "In house"-RIA (Tubingen). Results: The assays differed in median hGH concentrations by as much as 5.44 ng/mL (Immulite), and as little as 2.67 ng/mL (BC-IRMA). The mean difference between assays ranged from 0.35 to 2.71 ng/mL, whereas several samples displayed differences up to 11.4 ng/mL. The best correlation (r=0.992) was found between AutoDELFIA and Liasion, the lowest (r=0.864) was between an in-house RIA and iSYS. The between-assay CV (mean +/- SD) of values within the cut-off range was 24.3%+/- 7.4%, resulting in an assay-dependent diagnosis of growth hormone deficiency (GHD) in more than 27% of patients. Yet, adjustment of this data by linear regression or a conversion factor reduced the CV below 14%, and the ratio of assay-dependent diagnoses below 8%. Using quantile transformation, the CV and ratio were reduced to 11.4% and < 1%, respectively. Conclusions: hGH measurements using different assays vary significantly. Linear regression, conversion factors, or particularly quantile transformation are useful tools to improve comparability in the diagnostic procedure for the confirmation of GHD in childhood and adolescence

Control of (pre)-analytical aspects in immunoassay measurements of metabolic hormones in rodents

The measurement of circulating hormones by immunoassay remains a cornerstone in preclinical endocrine research. For scientists conducting and interpreting immunoassay measurements of rodent samples, the paramount aim usually is to obtain reliable and meaningful measurement data in order to draw conclusions on biological processes. However, the biological variability between samples is not the only variable affecting the readout of an immunoassay measurement and a considerable amount of unwanted or unintended variability can be quickly introduced during the pre-analytical and analytical phase. This review aims to increase the awareness for the factors 'pre-analytical' and 'analytical' variability particularly in the context of immunoassay measurement of circulating metabolic hormones in rodent samples. In addition, guidance is provided how to gain control over these variables and how to avoid common pitfalls associated with sample collection, processing, storage and measurement. Furthermore, recommendations are given on how to perform a basic validation of novel single and multiplex immunoassays for the measurement of metabolic hormones in rodents. Finally, practical examples from immunoassay measurements of plasma insulin in mice address the factors 'sampling site and inhalation anesthesia' as frequent sources of introducing an unwanted variability during the pre-analytical phase. The knowledge about the influence of both types of variability on the immunoassay measurement of circulating hormones as well as strategies to control these variables are crucial, on the one hand, for planning and realization of metabolic rodent studies and, on the other hand, for the generation and interpretation of meaningful immunoassay data from rodent samples

Toward a Diagnostic Score in Cushing's Syndrome

Cushing's syndrome (CS) is a classical rare disease: it is often suspected in patients who do not have the disease;at the same time, it takes a mean of 3 years to diagnose CS in affected individuals. The main reason is the extreme rarity (1-3/million/year) in combination with the lack of a single lead symptom. CS has to be suspected when a combination of signs and symptoms is present, which together make up the characteristic phenotype of cortisol excess. Unusual fat distribution affecting the face, neck, and trunk;skin changes including plethora, acne, hirsutism, livid striae, and easy bruising;and signs of protein catabolism such as thinned and vulnerable skin, osteoporotic fractures, and proximal myopathy indicate the need for biochemical screening for CS. In contrast, common symptoms like hypertension, weight gain, or diabetes also occur quite frequently in the general population and per se do not justify biochemical testing. First-line screening tests include urinary free cortisol excretion, dexamethasone suppression testing, and late-night salivary cortisol measurements. All three tests have overall reasonable sensitivity and specificity, and first-line testing should be selected on the basis of the physiologic conditions of the patient, drug intake, and available laboratory quality control measures. Two normal test results usually exclude the presence of CS. Other tests and laboratory parameters like the high-dose dexamethasone suppression test, plasma ACTH, the CRH test, and the bilateral inferior petrosal sinus sampling are not part of the initial biochemical screening. As a general rule, biochemical screening should only be performed if the pre-test probability for CS is reasonably high. This article provides an overview about the current standard in the diagnosis of CS starting with clinical scores and screenings, the clinical signs, relevant differential diagnoses, the first-line biochemical screening, and ending with a few exceptional cases

Guidance for the treatment of adult growth hormone deficiency with somapacitan, a long-acting growth hormone preparation

Adult growth hormone deficiency (AGHD) is a rare endocrine disorder characterized by an abnormal body composition, metabolic abnormalities associated with increased cardiovascular diseases, bone loss, and impaired quality of life. Daily subcutaneous injections with recombinant growth hormone (GH) can alleviate the abnormalities associated with AGHD. Several long-acting GH (LAGH) preparations are currently in development that aim to reduce treatment burden for patients receiving daily GH injections. Somapacitan (Sogroya®; Novo Nordisk, Denmark) is the first LAGH preparation that has been approved for treatment of AGHD in the United States, Europe, and Japan. The recent approval of somapacitan and anticipated approval of other LAGH molecules presents new questions for physicians planning to treat AGHD with LAGH in the future. Differences in the technologies used to prolong the half-life of recombinant GH are expected to result in variations in pharmacokinetic and pharmacodynamic profiles between preparations. Therefore, it is essential that physicians understand and consider such variations when treating patients with these novel GH replacement therapies. Here, we present a set of treatment recommendations that have been created to guide physicians initiating therapy with somapacitan in patients with AGHD who are eligible for GH replacement. Furthermore, we will review the published data that underlie these recommendations to explain the rationale for the treatment and monitoring advice provided