Column chromatographic characterization of complex formation of pro-IGF-II isoforms with acid labile subunit and IGF-binding proteins associated with non-islet cell tumour induced hypoglycaemia

Objective and design: Non-islet cell tumour induced hypoglycaemia (NICTH) is a paraneoplastic phenomenon that is associated with the formation of several isoforms of pro-insulin like growth factor 2 (pro-IGF-II), or so called big IGF-II. Disturbance of ternary complex formation by big IGF-II is assumed to be a crucial early event in the pathogenic cascade of hypoglycaemia. By size-exclusion chromatography, we investigated complex formation by adding different naturally occurring isoforms of pro-IGF-II to pooled normal adult serum. Results were compared with the analysis of the serum from a patient with NICTH. Results: Gel filtration experiments with the serum of a patient with NICTH demonstrated that ternary complex formation was severely compromised. The various forms of pro-IGF-II did not induce a shift of IGF-binding protein 3 (IGFBP-3) from 150 kD towards smaller binary complexes in the normal adult serum, suggesting that they did not interfere with the interaction between the add labile subunit and IGFBP-3. Instead, unglycosylated recombinant pro-IGF-II[1-104] was capable of forming a 150 kD complex. In contrast predominantly glycosylated and unglycosylated pro-IGF-II[1-87] eluted in the free unbound form. We showed that mature IGF-II and isoforms of pro-IGF-II were able to phosphorylate the IGF-I receptors of MC7 cells, albeit to a markedly lesser extent than IGF-I. When the patient's serum was tested in this system, the IGF-I receptor phosphorylation activity was considerably less than that in sera from age matched healthy individuals. Conclusion: We postulate that, alongside the presence of big IGF-II in the circulation, additional steps are required to stimulate the release of and pro-IGF-II isoforms from IGFBPs in vivo. These factors may be proteases, that are present in the local environment of the tumour and in insulin-sensitive tissues. (C) 2014 Elsevier Ltd. All rights reserved.