Adenylate cyclase-stimulating, bone-resorbing and B TGF-like activities in canine apocrine cell adenocarcinoma of the anal sac

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Abstract
Canine apocrine cell adenocarcinoma of the anal sac (APO-AS) is a spontaneously occurring tumor that causes humorally medicated hypercalcemia in 90% of cases. To further define the nature of the responsible mediator in APO-AS, we examined tumor extracts from five APO-AS and four control tumors for adenylate cyclase-stimulating activity (ACSA). All extracts from APO-AS contained potent ACSA, whereas the four control tumors did not. The ACSA extracted from one tumor demonstrated a dose response curve parallel to that of synthetic bovinePTH-(1–34) and was 80% inhibited by Nle8,18,Tyr34 bPTH-(3–34)amide at a concentration of 10−5 M. Extracts from three APO-AS and three control tumors were also examined forin vitro bone-resorbing activity (BRA). All APO-AS contained significant BRA, stimulating resorption 1.47 to 2.13-fold over basal, whereas none of the control tumors stimulated resorption. Purification of one extract using C18 reverse-phase high pressure liquid chromatography (RP-HPLC) resulted in a single sharp peak of ACSA which was 400-fold purified compared with the initial extract. This pool also contained significant bone-resorbing activity, whereas none of the adjacent pools did. Purification of a second extract using sequential CN and C18 RP-HPLC followed by size exclusion HPLC resulted in material that was at least 10,000-fold purified, and showed co-purification of ACSA and B TGF-like activity.