Identification of a natural soluble neuropilin-1 that binds vascular endothelial growth factor: In vivo expression and antitumor activity

Neuropilin-1 (NRP1) is a 130-kDa transmembrane receptor for semaphorins, me diators of neuronal guidance, and for vascular endothelial growth factor 16 5 (VEGF(165)), an angiogenesis factor. A 2.2-kb truncated NRP1 cDNA was clo ned that encodes a 644-aa soluble NRP1 (sNRP1) isoform containing just the a/CUB and b/coagulation factor homology extracellular domains of NRP1. sNRP 1 is secreted by cells as a 90-kDa protein that binds VECF165. but not VEGF (121). It inhibits I-125-VEGF(165) binding to endothelial and tumor cells a nd VEGF(165)-induced tyrosine phosphorylation of KDR in endothelial cells. The 3' end of sNRP1 cDNA contains a unique, 28-bp intron-derived sequence t hat is absent in full-length NRP1 cDNA, Using a probe corresponding to this unique sequence, sNRP1 mRNA could be detected by in situ hybridization dif ferentially from full-length NRP1 mRNA, for example, in cells of liver, kid ney, skin, and breast. Analysis of blood vessels in situ showed that NRP1, but not sNRP1, was expressed. sNRP1 was functional in vivo. Unlike control tumors, tumors of rat prostate carcinoma cells expressing recombinant sNRP1 were characterized by extensive hemorrhage. damaged vessels, and apoptotic tumor cells. These results demonstrate the existence of a naturally occurr ing, soluble NRP1 that is expressed differently from intact NRP1 and that a ppears to be a VEGF165 antagonist.