Prostate-specific Antigen Isoforms Research Articles

Biochemical characteristics and biological profile of prostate specific antigen (PSA)

Since its identification in seminai fluid in 1971, a tot more knowledge has been obtained about the biology and expression of prostate specific antigen (PSA). PSA is a glycoprotein consisting of 93% aminoacids and 7% carbohydrates with a molecular weight of about 30000 dalton. Functionally and structurally PSA is a kallikrein-like serine protease and its physiological role is the degradation of the major proteins of the seminal coagulum (semenogelin I and II, fibronectin) leading to semen liquefaction. The PSA gene is located on the 13q region of chromosome 19 and has a high degree of homology (more than 80%) with the gene of human glandular kallikrein (hGK1). PSA production and expression are preferentially but not exclusively associated with the normal, benign hyperplastic and cancerous tissues of the prostate. In fact, it has been demonstrated that PSA is also present in the accessory male sex glands of Cowper, Littre and Morgagni and other extra-prostatic neoplasms, such as salivary gland tumours and breast cancer. Many factors may influence PSA synthesis and production and among them the most important are androgen and growth factor stimulation. Advances have recently been made on the molecular isoforms of PSA. In the seminai fluid PSA seems partially bound to a serpine (protein C inhibitor), whilst in the serum PSA is predominantly associated to α-antichymotrypsin (ACT) and in low quantity to α2-macroglobulin (α2M). All these findings will have implications on clinical applications of PSA as a tumour marker for prostate cancer.

Evaluation of free psa isoforms, psa complex formation, and specificity of anti‐psa antibodies by hplc and page‐immunoblotting techniques

Both high performance liquid chromatographic (HPLC) and polyacrylamide gel electrophoresis-immunoblotting (PAGE-immunoblotting) procedures have been established for the study of isoforms of free prostate-specific antigen (PSA) and the complex formation between free PSA and protease inhibitors, and for the evaluation of the specificities of various anti-PSA antibodies. We found multiple isoforms of free PSA on PAGE, which were all capable of forming complexes with protease inhibitors. The same isoform pattern can be produced from the original seminal fluid. The PSA isoforms differ from each other most likely in charge because they could be converted to one band on SDS-PAGE and to a single peak by gel filtration chromatography. We found it difficult to form large quantities of PSA complex when mixing free PSA from seminal fluid with protease inhibitors, regardless of whether the free PSA or the protease inhibitors were in excess. Except for the PSA-ACT complex, which was consistently detectable by both HPLC and PAGE-immunoblotting techniques after incubation, these two procedures disagreed in their detection of PSA-A2M and PSA-AT complexes. The PSA-A2M complex was usually observable by immunoblotting techniques but barely detectable on HPLC, whereas PSA-AT was totally invisible by immunoblotting but appeared as a peak in the HPLC elution profile. Mixing free PSA with serum clearly resulted in both PSA-ACT and PSA-A2M complexes. However, more PSA-ACT than PSA-A2M was formed; the result was also confirmed by using 125I-PSA for mixing. PSA could be separated into active and inactive PSA by DEAE Sepharose chromatography with a 14-fold difference in protease activity. The difference in enzymatic activity apparently had no effect on complex formation. All the anti-PSA antibodies examined in this study reacted with PSA isoforms, PSA-ACT and PSA-A2M complexes. We conclude that it would be almost impossible to establish an assay to measure all forms of PSA in the serum and to expect to produce precise and accurate PSA values.