Complex formation between PSA isoenzymes and protease inhibitors

Purpose: We have studied complex formation between the isoenzymes of prostate specific antigen (PSA) and protease inhibitors in vitro. Materials and Methods: Ion exchange chromatography and hydrophobic interaction chromatography: (HIC) were used for rapid separation of PSA isoenzymes from inhibitors and for characterization of the complex formation. Immunofluorometric assays (IFMA) specific for free PSA, the 1PSA-alpha(1)-antichymotrypsin (PSA-ACT) complex and for both of these (total PSA) were used to measure various forms of PSA. Loss of free PSA immunoreactivity was used to estimate complex formation with alpha(2)-macroglobulin (A2M) and ACT, which also was measured by PSA-ACT IFMA. Results: Complex formation between PSA and A2M was more rapid than with ACT. After extended:incubation, about 75% of PSA reacted with ACT and 85% with A2M. When added to a mixture of ACT and A2M at concentrations corresponding to those in plasma, only 17% of PSA formed a complex with ACT while 17% remained free and 66% was undetectable, indicating complex formation with A2M. After extended incubation of PSA-ACT at 37C, a significant proportion of PSA was released as free active PSA. When A2M was included in the reaction mixture, the loss of PSA-ACT was not accompanied by appearance of free PSA, an indication that it complexed with A2M. Five to 18% of nicked PSA complexed with ACT whereas 54 to 67% reacted with A2M. Conclusions: alpha(2)-macroglobulin is the major inhibitor of PSA when it reaches the circulation. Contrary to earlier assumptions, nicked PSA can bind to A2M rendering it inaccessible to antibodies.