Protein C inhibitor biochemistry and pathology

Protein C inhibitor (PCI) is a heparin-binding serine protease inhibitor that regulates a variety of proteases, including some in tumor metastasis. We are characterizing both the structural (heparin binding site and pathobiological properties (in normal and malignant prostate) of PCI. Prostate Pathology: PCI is in normal, hyperplastic and malignant human prostatic epithelial cells. PCI antigen (and mRNA) was in LNCap tumor cells and production was altered when grown either without (charcoal-stripped serum) or with testosterone. We detected PCI by immunohistochemistry in a human prostate cancer xenograft (CWR22). Mice with CWR22 tumors were orchiectomized to reduce testosterone and sacrificed on consecutive days. Sections stained for PCI post-castration (up to 6 days) had an increase in the amount of immunoreactivity, this was confirmed by immunoblot and Northern blot studies. PCI antigen and mRNA was present in the androgen independent recurrent tumor that is the product of extended growth in the orchiectomized mice. Heparin Binding: PCI binds heparin in its H helix region. Previous studies identified R269 and K270 ("H1" rPCI) as important in heparin-accelerated protease inhibition. H1 rPCI was used as a template for further site directed mutagenesis of other basic H helix residues: K266A and K273A. The mutant rPCI's were expressed using a Baculovirus system. Compared to wild-type rPCI and HI rPCI, heparin-accelerated protease inhibition was greatly reduced and the proteins interacted poorly with heparin-Sepharose. Conclusions: Our data show that (i) the heparin binding domain of PCI is dependent on these Arg/Lys H helix residues and that (ii) PCI may help regulate the "proteolytic cascade" which aids prostatic tumor metastasis.

Duke Scholars

Author Maureane Hoffman Pathology