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Biologic and immunologic characterization and physical separation of ACTH and ACTH fragments in the ectopic ACTH
Abstract:
Extracts of tumors from 32 patients with the ectopic ACTH syndrome were subjected to simultaneous bioassay and radioimmunoassays for ACTH. Radioimmunoassays were performed using three antisera, one of which reacts with the extreme N-terminal 1-13 amino acid sequence of ACTH, the second with the N-terminal 1-23 sequence of the ACTH molecule, and the third with the C-terminal 25-39 amino acid sequence of ACTH. There was, in general, good correlation between bioactivity and N-terminal ACTH immunoreactivity. However, there were large excesses of both extreme N-terminal and C-terminal immunoreactive materials in most tumor extracts, which were not found in extracts of three human pituitaries. Three tumor extracts were subjected to molecular sieve chromatography on Sephadex G-50 fine resin. The bioactive ACTH eluted in the same fractions as pituitary ACTH (mol wt approximately 4,500 daltons) and reacted equally in all three ACTH radioimmunoassay systems. The bioactive tumor ACTH was neutralized by incubation with the C-terminal antiserum, indicating it has an intact C-terminal sequence of amino acids. The next several fractions from the Sephadex column contained a material, mol wt approximately 3,100, which was biologically inactive and had C-terminal immunoreactivity but no N-terminal or extreme N-terminal immunoreactivity. Incubation with the N-terminal 1-23 ACTH antiserum did not adsorb these C-terminal fragments, indicating they lacked an intact sequence of amino acids in this region. A smaller ACTH fragment (mol wt approximately 1,800 daltons) eluted in still later fractions and reacted with the extreme N-terminal antiserum but not with the N-terminal or C-terminal antisera. It had no steroidogenic activity, but appeared to have significant melanocyte-stimulating activity. It is concluded that, in addition to an ACTH similar, if not identical, to pituitary ACTH, tumors of patients with the ectopic ACTH syndrome contain both N-terminal and C-terminal ACTH fragments.
Insights
Tumors in ectopic ACTH syndrome patients produce bioactive adrenocorticotropic hormone (ACTH) and excess N-terminal and C-terminal ACTH fragments. These fragments, unlike pituitary ACTH, show varied biological activity and immunoreactivity.
Area of Science:
- Endocrinology
- Molecular Biology
- Oncology
Background:
- Ectopic ACTH syndrome (EAS) is characterized by non-pituitary tumors producing adrenocorticotropic hormone (ACTH).
- Understanding the nature and processing of ACTH in these tumors is crucial for diagnosis and treatment.
Purpose of the Study:
- To characterize the forms of ACTH present in tumors from patients with EAS.
- To compare tumor-derived ACTH with pituitary ACTH using bioassays and immunoassays.
Main Methods:
- Tumor extracts from 32 EAS patients were analyzed using bioassay and radioimmunoassays with three different ACTH antisera.
- Molecular sieve chromatography (Sephadex G-50) was employed to separate and analyze ACTH components.
Main Results:
- Tumor extracts contained bioactive ACTH similar to pituitary ACTH, correlating with N-terminal immunoreactivity.
- Significant excesses of N-terminal and C-terminal ACTH fragments, not found in pituitary extracts, were detected.
- Chromatography revealed inactive C-terminal fragments and smaller N-terminal fragments with melanocyte-stimulating activity.
Conclusions:
- Ectopic ACTH-producing tumors contain not only bioactive ACTH but also various biologically inactive N-terminal and C-terminal fragments.
- These fragments contribute to the complex immunoreactivity observed in EAS and may have distinct biological functions.
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