Plastocyanins are small, type I blue copper proteins, or cupredoxins, ranging in size from 97 to 105 amino acids that play an essential role in oxygenic photosynthesis in green plants, algae and cyanobacteria, where they act as mobile electron carriers. We have reported that the cupredoxins azurin and rusticyanin, derived from bacterial sources, and peptides derived from them, can penetrate human cancer cells and exert anti-cancer effects in vitro and in vivo. In general, the most selective and active peptides form amphipathic, α- helical and β- sheet type structures with theoretical pI values under 7. We now extend these findings to plastocyanin derivatives, which unlike azurin and rusticyanin, do not have a significant (predicted) α-helical motif. However, these 15- 20 aa (1671 - 2154 kDa) peptides exhibit a β-sheet structure, have pI values that range from 3.8 -8.15, contain motifs structurally homologous to the ephrin G-H loop and bind competitively to ephrin receptors in a 1:1 ratio. Confocal microscopy revealed that amino acid fragments 70-84, Plc-15, aa 66-85, Plc-20 of plastocyanin derived from Ulva pertusa and aa 76-90, Plc-15 b, from Phormidium laminosum penetrate a variety of human cancer cell lines without producing any significant alteration in membrane structure (LDH leakage assays) or exhibiting any hemolytic activity on human or murine RBCs (5-100 uM). Each of these peptides exerted a selective and significant, dose related (5-100 uM) antiproliferative activity on a variety of human solid tumor and hematologic malignancies in vitro. Plc-15 (U. pertusis) inhibited (p < 0.05) the growth of HT-1080 sarcoma, MCF-7 breast, DU-145 prostate, HCT-116 colon and CRL1611 kidney cancer with no effect on 5 leukemia and lymphoma cell lines. Plc-15b (P. laminosum) also inhibited (p < 0.05) the growth of HT-1080 sarcoma, MCF-7 breast, HCT-116 colon and UISO Mel-2 melanoma and CCF-STTG1 astrocytoma cells, but did not alter the growth of DU-145 or CRL-1611 cells or effect the 5 leukemia and lymphoma cell lines tested. In contrast, Plc-20 (U. pertusis) inhibited (p < 0.05) the growth of MCF-7, NCI-H23 NSCLC and HCT-116 cells, and the growth of TIB152Jurkat, HL60, HSB2and U937 cells as determined by MTT assay. Results from MTT assays suggested that these peptides inhibited growth via apoptosis rather than cell cycle inhibition. In conclusion, our initial studies suggest that fragments of plastocyanin that exhibit a β-sheet structure and a motif similar to the ephrin G-H loop inhibit cancer cell proliferation by inducing apoptosis without exerting a direct effect on the cancer cell membrane or hemolytic activity.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA