Combined administration of microdoses of growth hormone and erythropoietin: Effects on performance and evaluation of GH detection capability using anti‐doping methods

Abstract

The combination of growth hormone (GH) and recombinant erythropoietin (rEPO) is thought to be used particularly in endurance sports. Our objective was to reproduce a 2‐week administration of rEPO microdoses alone or in combination with GH microdoses (three times a week) on healthy and athletic male subjects and to evaluate if GH had any additional effects compared to EPO treatment alone. The effects of the treatments on hematological parameters and VO2max were studied as well as the detection of GH in serum. While the rEPO microdose regimen was associated with a significant increase in reticulocytes, no clear elevation in hemoglobin concentration (HGB) was observed. Using a correction by plasma volume did not reveal more effects of EPO on HGB. Our results did not show any additional effect when the GH microdoses were co‐administered. In addition, no clear increase in VO2max was observed after treatment, with an elevation in only half the subjects in both groups (EPO and EPO+GH). A clear effect of GH on insulin‐like growth factor I (IGF‐I) was seen but it was lower on procollagen III amino‐terminal propeptide (P‐III‐NP). GH detection using the direct isoform test identified only one subject 24 hours after receiving GH. The GH biomarker test combining IGF‐I and P‐III‐NP was not able to detect the GH administration. However, a longitudinal follow‐up of the intraindividual variations showed a significant increase in IGF‐I 24 and 48 hours after GH administration in most subjects, while the effect of GH microdoses on P‐III‐NP was less straightforward.

Longitudinal studies of putative growth hormone (GH) biomarkers and hematological and steroidal parameters in relation to 2 weeks administration of human recombinant GH

“…To our knowledge this is the first observation that recGH influences erythropoiesis in healthy adults. In a recent study, it was noted that a recGH micro‐dose regimen did not show any additional effect on reticulocytes when co‐administered with rEPO 8 . However, in that study recGH was administered only every second day which might explain the lack of response.…”

“…A future test approach may be a new endocrine module of the athlete biological passport (ABP) including the longitudinal monitoring of the GH‐2000 score and its components insulin‐like growth factor (IGF‐I) and procollagen type III N‐terminal peptide (P‐III‐NP) 6 . It has been shown that such a longitudinal approach may increase the likelihood of detecting low doses of recGH 7,8 …”

The intra‐individual stability of GH biomarkers IGF‐I and P‐III‐NP in relation to GHRH administration, menstrual cycle, and hematological parameters

“…recGH. These direct methods have short detection times, 5,6,19 especially when microdosing is used, and any information on how the athlete is actually doping with these substances are needed to collect the samples at an accurate time point to get an adverse analytical finding (AAF). The detection window for GHRH in this controlled study seems to be superior to the direct method where a 500-μg dose 19 could be detected for 2 h, whereas using the longitudinal approach described here, with a dose less than 100 μg, suspicious results were detected for more than 12 h following the last administration.…”

“…3,4 For recGH, we and others have shown that longitudinally monitoring IGF-I may be a good approach to detect microdose doping with recGH. 5,6 The intra-individual variation of the GH biomarkers IGF-I and P-III-NP has not previously been studied with respect to the administration of growth hormone-releasing hormone (GHRH). GHRH is an endogenous peptide hormone that contains 44 amino acids, is produced in the hypothalamus, and is responsible for the stimulation of GH production.…”

The 37th Manfred Donike workshop on doping analysis

“…Studies concerning the (improved) detection of peptidic drugs and drug candidates focused, amongst others, on erythropoietin, growth hormone, and the myostatin‐neutralizing antibody domagrozumab. For erythropoietin, a superior immunoaffinity‐based sample preparation protocol was established, which allowed for a higher sample throughput and reduced manual workload while conserving the assay's analytical sensitivity, and controlled microdose growth hormone administration studies corroborated the potential of intra‐individual biomarker monitoring (particularly IGF‐I and P‐III‐NP) for an enhanced traceability of growth hormone misuse in sports . Moreover, two complementary test methods concerning domagrozumab, a representative of the novel class of therapeutic antibodies, were developed, enabling the detection of the drug candidate at serum or plasma levels of 50 ng/mL using either Western blotting or bottom‐up LC‐HRMS analytical approaches …”