Biosimilar erythropoiesis-stimulating agents and the risk of developing anti-drug antibodies-a systematic review.

PURPOSE: We systematically reviewed published observational studies and randomized controlled trials (RCT) reports of clinical trials on erythropoiesis-stimulating agents (ESA's). Only studies evaluating the risk of developing anti-drug antibodies (ADA) of both original and biosimilar drugs were chosen. METHODS: Databases including PubMed, EMBASE and Cochrane Library were searched up to 17 March 2015. Two reviewers independently assessed the relevant studies for risk of bias. RESULTS: Twenty-one publications were included. The overall prevalence of ADA in the studies was about 0.2 to 0.5 %. Most studies were not designed to monitor the development of ADA and often the study duration was too short (less than 6 months) and the patient population too small. Moreover, in many studies, the assays used only determined the presence of ADA and did not measure therapy failure due to ADA. In one RCT, as many as 13 cases (4 %) of ADA were identified. CONCLUSION: ADA development seems to be low in short-term studies with ESA. None of the efficacy and safety issues for ESA biosimilars were judged to be adequately addressed in the evaluated literature, with respect to ADA formation, due to the study design and the assay method used.

Is the decision on the use of biosimilar growth hormone based on high quality scientific evidence? - a systematic review.

BACKGROUND: The authors carried out a systematic and critical review of the scientific literature regarding the possible development of neutralising antibodies developed in patients treated with growth hormone biosimilars (defined as a drug expected to be similar to the originator or original pharmaceutical -European Medicines Agency) as compared to the reference drug. As a consequence, we discovered two major issues, namely, the poor quality of the comparative clinical trials and the poor quality of the antibody assays used during the trials. METHODS: The literature review was performed according to the principle of the Cochrane Collaboration and SBU. The electronic literature search included the databases PubMed, EMBASE and The Cochrane Library up to December 2012. Two independent reviewers assessed abstracts and full-text articles. RESULTS: The search identified 1,553 abstracts related to the subject. Only six articles contained data on biosimilar growth hormone or antibody results obtained with appropriate methods. None of the studies fulfilled the criteria for high quality randomised controlled trials. Qualitative rather than quantitative assays were used for monitoring antibody formation. CONCLUSIONS: It is our firm opinion , that since biosimilars are not identical, emphasis must be placed on the quality of the comparative clinical trials performed and the quality of the analytical studies in order to guarantee patient safety. Clinical trials should follow established quality rules for controlled comparative randomised clinical trials. A whole set of new guidelines is required.

Network of European studies of genes in growth (NESTEGG).

The network of European studies of genes in growth (NESTEGG) is an international growth genomics project, focusing on the birth size phenotypes of small for gestational age (SGA) and idiopathic short stature. Seven hundred controls and 1,275 cases with their parents have been recruited. Detailed clinical histories and auxological measurements are recorded in a clinical database. Candidate gene studies are being undertaken with the study DNA samples. These genetic data will be used to explore associations with the clinical phenotypes of short stature and SGA birth size, and, in a subset, response to growth hormone (GH) therapy. This article describes the study methodology and reviews the association of the exon 3-deleted genotype of the GH receptor with GH responsiveness in GH-treated children born SGA.

Exon 3-deleted/full-length growth hormone receptor polymorphism genotype frequencies in Spanish short small-for-gestational-age (SGA) children and adolescents (n = 247) and in an adult control population (n = 289) show increased fl/fl in short SGA.

CONTEXT: A polymorphism in the human GH receptor gene (d3/fl-GHR) resulting in genomic deletion of exon 3 has been associated with the degree of height increase in response to GH therapy. OBJECTIVE: The objective of the study was to evaluate the frequencies of d3/fl-GHR polymorphism genotypes in control and short small-for-gestational-age (SGA) populations. DESIGN: An adult control population with heights normally distributed (ACPNH) between -2 and +2 sd score (SDS) and a short non-GH-deficient SGA child population were selected. SETTING: Thirty Spanish hospitals participated in the selection of the short non-GH-deficient SGA children in the setting of a controlled, randomized trial, and one of these hospitals selected the ACPNH. CONTROLS AND PATIENTS: Two hundred eighty-nine adult subjects of both sexes constituted the ACPNH and 247 children and adolescents of both sexes the short SGA patients. MAIN OUTCOME MEASURES: Heights and weights were recorded in the ACPNH, and auxologic and biochemical data were recorded at each hospital for the SGA patients; d3/fl-GHR genotypes were determined and data analyzed in a single hospital. RESULTS: In short SGA patients, d3/fl-GHR genotype frequencies were significantly different from those in ACPNH, with a higher frequency of fl/fl genotype (P < 0.0001). In ACPNH, a trend toward diminished d3/d3 genotype frequency was observed in the shortest height group (height or=-2 SDS, n = 60). CONCLUSIONS: Our data showed significant differences in the frequency distribution of the d3/fl-GHR genotypes between a normally distributed adult height population and short SGA children, with the biologically less active fl/fl genotype being almost twice as frequent in SGA patients. These data suggest that the d3/fl-GHR polymorphism might be considered among the factors that contribute to the phenotypic expression of growth.

Improvement in insulin sensitivity without concomitant changes in body composition and cardiovascular risk markers following fixed administration of a very low growth hormone (GH) dose in adults with severe GH deficiency.

OBJECTIVE: Untreated GH-deficient adults are predisposed to insulin resistance and excess cardiovascular mortality. We showed previously that short-term treatment with a very low GH dose (LGH) enhanced insulin sensitivity in young healthy adults. The present study was therefore designed to explore the hypothesis that LGH, in contrast to the standard GH dose titrated to normalize serum IGF-I levels (SGH), may have differing effects on insulin sensitivity, body composition, and cardiovascular risk markers [lipid profile, C-reactive protein (CRP), interleukin-6 (IL-6), tumour necrosis factor-alpha (TNF-alpha) and adiponectin] in adults with severe GH deficiency. PATIENTS AND METHODS: In this 12-month open, prospective study, 25 GH-deficient adults were randomized to receive either a fixed LGH (0.10 mg/day, n = 13) or SGH (mean dose 0.48 mg/day, n = 12), and eight age- and body mass index (BMI)-matched GH-deficient adults acted as untreated controls. Fasting blood samples were collected at baseline and at months 1, 3, 6, 9 and 12. Assessments of insulin sensitivity, using the hyperinsulinaemic euglycaemic clamp technique, and body composition, using dual-energy X-ray absorptiometry, were performed at baseline and at month 12. RESULTS: The LGH decreased fasting glucose levels (P < 0.01) and enhanced insulin sensitivity (P < 0.02), but body composition, nonesterified fatty acid (NEFA) levels and cardiovascular risk markers were unchanged. The SGH did not modify insulin sensitivity, decreased truncal fat mass (P < 0.05), CRP (P < 0.05) and IL-6 (P < 0.05) levels, and increased NEFA levels (P < 0.05). No changes were observed with the untreated controls. CONCLUSION: Our data indicate that, in contrast to the SGH, fixed administration of the LGH enhances insulin sensitivity with no apparent effects on body composition, lipolysis and other surrogate cardiovascular risk markers in adults with severe GH deficiency. Thus, the LGH may potentially be a beneficial replacement dose in reducing type 2 diabetes risk in adults with severe GH deficiency.

Changes in free rather than total insulin-like growth factor-I enhance insulin sensitivity and suppress endogenous peak growth hormone (GH) release following short-term low-dose GH administration in young healthy adults.

High-dose GH administration is commonly associated with impaired insulin sensitivity (S(I)) in humans. Paradoxically we have shown that low-dose GH (1.7 microg/kg.d) administration enhances beta-cell function in young healthy adults. In the present double-blind, placebo-controlled, cross-over study, we explored the physiological effects of this low GH dose on glucose metabolism in 12 young healthy adults (seven males, 19-29 yr). At pretreatment and after each 14-d treatment block, overnight metabolic profiles were assessed followed by a hyperinsulinemic euglycemic clamp, whereas fasting blood samples were collected weekly. In subjects treated with GH first (group A, n = 6), GH treatment increased total IGF-I (P < 0.05) and IGF binding protein-3 (P < 0.01) after 7 d, but these levels subsequently returned to pretreatment levels after 14 d. In contrast, free IGF-I increased (P < 0.05), and overnight GH pulse peak amplitude decreased (P < 0.01) after 14 d. In subjects treated with placebo first (group B, n = 6), all biochemical parameters were unchanged after placebo treatment, whereas the changes in free and total IGF-I were similar to those of group A after GH treatment. Combined clamp data from both groups A and B (n = 12) showed that 14-d GH treatment decreased overnight plasma insulin levels (P < 0.02) and hepatic glucose appearance (P < 0.05) and increased S(I) (P < 0.01). Of note, the GH-induced changes in S(I) positively correlated with the changes in free IGF-I (r = 0.72, P < 0.01). In conclusion, low-dose GH administration enhanced S(I) and suppressed endogenous peak GH release, and we hypothesize that these effects are the direct result of increased serum levels of free IGF-I.

Short-term low-dose growth hormone administration in subjects with impaired glucose tolerance and the metabolic syndrome: effects on beta-cell function and post-load glucose tolerance.

OBJECTIVE: Modest elevations in circulating IGF-I levels have been suggested to protect against the development of glucose intolerance in insulin-resistant subjects. To further understand the interactions of GH and IGF-I on beta-cell function and post-load glucose tolerance in glucose-intolerant subjects predisposed to diabetes, we performed a pilot study in 12 subjects with impaired glucose tolerance and the metabolic syndrome using a low GH dose (1.7 microg/kg per day) known to increase endogenous IGF-I production. DESIGN: Fourteen daily GH or placebo injections in a double-blind cross-over study. METHODS: Baseline and post-treatment oral glucose tolerance tests were performed. The homeostasis model assessment and the insulinogenic index was used to estimate fasting insulin sensitivity (S(I)) and beta-cell function respectively, whereas changes in the incremental area under the curve were used to estimate post-load glucose tolerance (DeltaAUC(glu)) and post-load insulin levels (DeltaAUC(ins)). RESULTS: GH increased total IGF-I (P<0.02), free IGF-I (P<0.04) and fasting insulin (P<0.04) levels, but did not modify plasma IGF-binding proteins (IGFBPs)-1 and -3, fasting glucose, non-esterified fatty acid and C-peptide levels, and fasting S(I). After oral glucose intake, glucose tolerance improved (P<0.03), but post-load insulin levels and beta-cell function remained unchanged. CONCLUSION: Short-term low-dose GH administration induced fasting hyperinsulinaemia possibly by reducing insulin clearance but improved post-load glucose tolerance, suggesting that increased bioavailable IGF-I enhanced post-load S(I) without altering beta-cell function. Longer-term studies are required to ascertain whether these positive effects on post-load glucose tolerance and the preservation of beta-cell function can be sustained by this GH dose in these high-risk subjects.

A novel dysfunctional growth hormone variant (Ile179Met) exhibits a decreased ability to activate the extracellular signal-regulated kinase pathway.

The pituitary-expressed GH1 gene was screened for mutation in a group of 74 children with familial short stature. Two novel mutations were identified: an Ile179Met substitution and a -360A-->G promoter variant. The Ile179Met variant was shown to exhibit a similar degree of resistance to proteolysis as wild-type GH, indicating that the introduction of Met does not cause significant misfolding. Secretion of Ile179Met GH from rat pituitary cells was also similar to that of wild type. Although receptor binding studies failed to show any difference in binding characteristics, molecular modeling studies suggested that the Ile179Met substitution might nevertheless perturb interactions between GH and the GH receptor loop containing the hotspot residue Trp169, thereby affecting signal transduction. The ability of the Ile179Met variant to activate a signal transducer and activator of transcription (STAT) 5-responsive luciferase reporter gene and induce phosphorylation of STAT 5 and ERK was therefore studied. In contrast to its ability to activate STAT 5 normally, activation of ERK by the Ile179Met variant was reduced to half that observed with wild type. Although differential effects on the activation of distinct signaling pathways by a mutant receptor agonist are unprecedented, these findings also suggest that the ERK pathway could play a role in mediating the action of GH.

Human growth hormone 1 (GH1) gene expression: complex haplotype-dependent influence of polymorphic variation in the proximal promoter and locus control region.

The proximal promoter region of the human pituitary expressed growth hormone (GH1) gene is highly polymorphic, containing at least 15 single nucleotide polymorphisms (SNPs). This variation is manifest in 40 different haplotypes, the high diversity being explicable in terms of gene conversion, recurrent mutation, and selection. Functional analysis showed that 12 haplotypes were associated with a significantly reduced level of reporter gene expression whereas 10 haplotypes were associated with a significantly increased level. The former tend to be more prevalent in the general population than the latter (p<0.01), possibly as a consequence of selection. Although individual SNPs contributed to promoter strength in a highly interactive and non-additive fashion, haplotype partitioning was successful in identifying six SNPs as major determinants of GH1 gene expression. The prediction and functional testing of hitherto unobserved super-maximal and sub-minimal promoter haplotypes was then used to test the efficacy of the haplotype partitioning approach. Electrophoretic mobility shift assays demonstrated that five SNP sites exhibit allele-specific protein binding. An association was noted between adult height and the mean in vitro expression value corresponding to an individual's GH1 promoter haplotype combination (p=0.028) although only 3.3% of the variance of adult height was found to be explicable by reference to this parameter. Three additional SNPs, identified within sites I and II of the upstream locus control region (LCR), were ascribed to three distinct LCR haplotypes. A series of LCR-GH1 proximal promoter constructs were used to demonstrate that 1) the LCR enhanced proximal promoter activity by up to 2.8-fold depending upon proximal promoter haplotype, and that 2) the activity of a given proximal promoter haplotype was also differentially enhanced by different LCR haplotypes. The genetic basis of inter-individual differences in GH1 gene expression thus appears to be extremely complex.

Novel mutations of the growth hormone 1 (GH1) gene disclosed by modulation of the clinical selection criteria for individuals with short stature.

Subtle mutations in the growth hormone 1 (GH1) gene have been regarded as a comparatively rare cause of short stature. Such lesions were sought in a group of 41 individuals selected for short stature, reduced height velocity, and bone age delay; a group of 11 individuals with short stature and idiopathic growth hormone deficiency (IGHD); and a group of 154 controls. Heterozygous mutations were identified in all three groups but disproportionately in the individuals with short stature, both with (odds ratio 25.2; 95% CI, 5.1-132.2) and without (odds ratio 3.6; 95% CI, 1.0-12.9) IGHD. Twenty-four novel GH1 gene lesions were found. Thirteen novel missense mutations were characterized by assaying the signal transduction activity of in vitro expressed variants; six (T27I, K41R, N47D, S71F, S108R, and T175A) exhibited a reduced ability to activate the JAK/STAT pathway. Molecular modeling suggested that both K41R and T175A might compromise GH receptor binding. Seven GH variants (R16C, K41R, S71F, E74K, Q91L, S108C, and a functional polymorphism, V110I) manifested reduced secretion in rat pituitary cells after allowance had been made for the level of expression attributable to the associated GH1 proximal promoter haplotype. A further leader peptide variant (L-11P) was not secreted. Eleven novel mutations in the GH1 gene promoter were assessed by reporter gene assay but only two, including a GH2 gene-templated gene conversion, were found to be associated with a significantly reduced level of expression. Finally, a novel intron 2 acceptor splice-site mutation, detected in a family with autosomal dominant type II IGHD, was shown to lead to the skipping of exon 3 from the GH1 transcript. A total of 15 novel GH1 gene mutations were thus considered to be of probable phenotypic significance. Such lesions are more prevalent than previously recognized and although most may be insufficient on their own to account for the observed clinical phenotype, they are nevertheless likely to play a contributory role in the etiology of short stature.

The effects of short-term administration of two low doses versus the standard GH replacement dose on insulin sensitivity and fasting glucose levels in young healthy adults.

GH has both diabetogenic and insulin-like actions. Supraphysiological GH doses are known to reduce insulin sensitivity (S(I)), but lower doses are less well studied. We therefore compared the effects of two physiological GH doses (intermediate, 0.0033 mg/kg x d; low, 0.0017 mg/kg x d) with the standard adult GH deficiency replacement dose (standard, 0.008 mg/kg x d) on S(I), beta-cell function, IGF-I, and IGF binding proteins (IGFBPs)-1 and -3 in healthy adults. Eleven healthy nonobese volunteers (4 males and 7 females, aged 21-38 yr) received 7 daily injections of the standard and intermediate GH doses, and 10 (5 males and 5 females, aged 21-38 yr) received the low dose. Fasting blood samples were collected daily (days 1-8). S(I) and beta-cell function were calculated using the Homeostasis model assessment. All GH doses increased IGF-I and IGFBP-3 levels, with the standard dose inducing the greatest rise (P < 0.001). At day 2 vs. baseline, all three doses increased the IGF-I/IGFBP-3 ratio, but only the standard dose lowered IGFBP-1 levels (P = 0.03). The standard dose reduced S(I) (P = 0.01), whereas the intermediate dose increased S(I) (P < 0.005) and lowered fasting insulin levels (P < 0.01). The low dose did not modify S(I), but reduced fasting glucose levels (P < 0.0001) and increased beta-cell function (P = 0.001). Males demonstrated higher IGF-I and IGFBP-3 responsiveness to the standard dose than females. Males also showed greater increase in S(I) and decrease in fasting glucose levels on both intermediate and low doses. In conclusion, the metabolic effects of GH are dose- and gender-dependent. The standard adult GH deficiency replacement dose induced insulin resistance, whereas lower doses improved S(I), especially in males. The low GH dose lowered fasting glucose levels and could represent the optimal dose to stimulate beta-cell function without compromising S(I) in insulin-resistant GH-deficient adults.