Improvement of a Myostatin Imperacer assay towards a high-sensitive test system for the detection of anabolic manipulations, including gene doping strategies

Code: 08B02PD

Improvement of a Myostatin Imperacer assay towards a high-sensitive test system for the detection of anabolic manipulations, including gene doping strategies

Main findings

Our recent research has demonstrated that monitoring the fingerprint expression of members of the myostatin signalling pathway is a promising tool to detect manipulations of myostatin. An important result of this project was the development of a sensitive ACT IIB Imperacer. This is a very essential step into the fight against the abuse of myostatin inhibitors because a soluble form of the activin type IIB receptor (ACT IIB), as a strategy for myostatin inhibition, is already examined in clinical trials (Acceleron). In our opinion our assay is applicable to detect the abuse of such myostatin inhibitors. First results demonstrate that it also works in capillary blood. As an alternative to antibody based Imperacers we also have started to develop Imperacer assays based on receptor ligand interactions. For example we could demonstrate that myostatin is detectable in the serum via binding to a recombinant ACT IIB receptor linked to DNA. As a basis for an indirect detection of myostatin inhibition ratios of FOLLI, MYPORO and ACT IIB were determined in long term studies with male volunteers and in females in different phases of the menstrual cycle. The results show notably inter-individual variations, however the distinct individual ratios were stable and not affected by training, menstrual cycle and circadian rhythms.

To determine whether the analysis of FOLLI, MYPORO and ACT IIB ratios are suitable to detect anabolic effects of steroids, their expression was analysed in untrained males, bodybuilders abusing anabolic steroids and “clean” bodybuilders. Our data demonstrate a tendency for a lowered FOLLI/MYOPRO ratio in the serum of natural bodybuilders. However, in our opinion, these variations cannot be used to decide whether somebody has abused anabolic steroids or not.

These data are in agreement to data from matching animal experiments. Fortunately, in these experiments we found that the endocrine profile and IGF-1 expression is strongly affected by anabolic steroid abuse. A WADA funded pilot project based on this observation is ongoing (LIVE) in the moment. Finally we have conducted an animal experiment with myostatin siRNA. The results of this experiment, in agreement to data obtained from Myostatin Knockout (KO) mice, indicate that manipulation of myostatin signalling, even by siRNA, is indeed detectable by comparing ratios of FOLLI, MYOPORO and ACT IIB. Interestingly in these experiments we recognised that fat mass was affected by myostatin siRNA much stronger than muscle mass. This knowledge is very helpful for identification of new biological markers for indirect detection of myostatin inhibition.

In summary we succeeded in the development of a high sensitive ACT IIB Imperacer, which can be used for the direct detection of specific myostatin inhibitors. This assay seems to be functional in capillary blood samples. Long term studies in males and females indicate that the ratios of FOLLI, MYPORO and ACT IIB are individually very stable and not effected by training, menstrual cycle and anabolic steroids. The use of specific myostatin inhibitors, in our studies specific siRNA to inhibit myostatin in mice, resulted in a significant shift in these patterns. This knowledge is very helpful for indirect detection of myostatin inhibition.