Identification of epidermal growth factor (EGF), in an unknown pharmaceutical preparation suspected to contain insulin like growth factor 1 (IGF-1).

The last 30years are marked by a tremendous evolution in the comprehension of human physiology. Although many physiological processes remain enigmatic, our increased understanding has led to a considerable development of medicines. A small, but yet a more progressively important sub fraction of this drug market consists of recombinant proteins and peptides. Although in most western countries a prescription is required for the injection of these compounds, some of them can easily be acquired from illicit internet pharmacies whether or not disguised as ‘research companies’. Some of these drugs can not only be used to cure ourselves but also to improve our sports performance, deepen our tan, and strive to the ideals imposed by society. Reports originating from controlling agencies residing in Germany, the UK, Denmark, Norway, Italy, the USA, Australia, and Belgium show that monoclonal anti‐cancer antibodies, insulins, growth hormones, human chorionic gonadotropin, Clostridium botulinum toxin type A, potential doping peptides, putative anti‐obesity drugs, skin tanning peptides, neuropeptides and a putative anti‐cancer polypeptide were identified in seized samples. The danger of these illegal polypeptides resides in the fact that these polypeptide products are not produced under controlled environment and therefore may contain the wrong ingredient, the wrong dosage, bacteria, viruses, heavy metals, etc., which could result in severe health issues. In some cases these illegal acts had a deadly outcome. Furthermore, for many of these polypeptide drugs, there is a limited knowledge of their effects and/or side‐effects (long and short term) hence clinical studies have not been terminated or even worse were given a negative advice due to overall potential adverse health effects. Global actions are on‐going to protect the sometimes unaware public from these malignant activities. In Belgium, the Federal Agency for Medicines and Health Products (FAMHP) is responsible for this task and upon their request unknown pharmaceutical preparations suspected to contain illegal drugs are regularly subject to analysis in our official medicines control laboratory (OMCL). At the end of 2014, an unknown pharmaceutical preparation, claimed to contain insulin like growth factor 1 (IGF‐1) was submitted for analysis by immunoblotting against IGF‐1 and analogues, prior to liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). Based on the interpretation of the western blot and the LC‐MS data we were not able to identify this substance as being any form of insulin like growth factor (IGF), including IGF‐1, IGF‐2 or synthetic analogues potentially used as doping agent. Further analysis by LC‐MS/MS and subsequent peptide sequencing showed that the substance present in the sample corresponds to epidermal growth factor (EGF). EGF plays a major role in the regulation of cell growth, proliferation and differentiation and is therefore frequently used in cell culture. This polypeptide, consisting of 53 amino acids, with sequence NSDSECPLSHDGYCLHDGVCMYIEALDKYACNCVVGYIGERCQYRDLKWWELR and 3 intramolecular disulfide bonds has an average Molecular weight of 6215.9Da. This polypeptide is known to interact with the EGF‐receptor (EGFR) and can promote tumour cell motility and invasion. Therefore the blocking of EGF binding to the EGFR and further downstream signalling pathway is currently marked as a target for anti‐cancer therapy for certain cancers (reviewed in Yewale et al.). Indeed, a vaccine targeting EGF, named CimaVax‐EGF, results in the production of antibodies directed against EGF and this vaccine is being used as cancer therapy against non‐small‐cell lung carcinoma in Cuba. The CimaVax vaccine consists of recombinant EGF chemically conjugated to the protein P64K from Neisseria meningitides. It induces the production of antibodies directed against EGF and results in the prevention of EGF to attach to EGFR and thus inhibiting the signal that tells cancer cells to grow and divide. Nowadays, the product is undergoing further trials for possible licensing in Europe and the United States. Although no injectable legal medicine is available on the market, the product is used in the cosmetic industry. It is thought that the topical application of EGF would enhance wound healing and stimulate skin rejuvenation. Taken together, we believe that the purchaser was unaware of the presence of EGF in the sample. This case of potential dangerous wrong labelling clearly demonstrates the peril of purchasing potential doping agents via the black market.