Inhibition of processing of parathyroid hormone-related peptide by anti-sense furin: effect in vitro and in vivo on rat Leydig (H-500) tumor cells.

Abstract

To attain full biological activity, the precursor pro-parathyroid hormone-related peptide (ProPTHRP) must be converted to the mature peptide PTHRP. We have examined the effect of inhibiting expression of the pro-hormone convertase furin in H-500 rat Leydig tumor cells on PTHRP production and action in vitro and in vivo. H-500 Leydig tumor cells were stably transfected with a mammalian expression plasmid containing furin cDNA in an anti-sense orientation. This resulted in inhibition of endogenous furin mRNA expression and of protein production as assessed by immunocytochemistry. These experimental cells secreted extended NH2-terminal PTHRP forms with reduced adenylate cyclase-stimulating activity. This was associated with a marked decrease in the proliferation of these tumor cells in vitro. Transfected and control cells were then implanted into male Fischer rats. Animals implanted with control cells became hypercalcemic. In contrast, animals implanted with experimental cells maintained near normal levels of plasma calcium. Experimental cells inoculated in vivo developed into tumors of significantly decreased volume compared to control cells and animal survival time was prolonged. Our results indicate that alteration of the processing of PTHRP can diminish the hypercalcemic endocrine actions of PTHRP and can reduce autocrine/paracrine effects of PTHRP on tumor cell growth both in vitro and in vivo. Furin may also exert a broader role in processing other factors required for tumor proliferation. Consequently, anti-sense modulation of furin activity may be a potential modality for understanding the mechanism of neoplastic growth and progression.