Development of new anti-cancer peptides from conformational energy analysis of the oncogenic ras-p21 protein and its complexes with target proteins.
We have employed a computational approach to design peptides, from known oncogenic proteins, that inhibit tumor growth. This approach has been applied to the ras-p21 protein that becomes oncogenic when single amino acid substitutions occur at critical positions in its polypeptide chain, such as at Gly 12 and Gln 61. In this approach, using two sampling methods, molecular dynamics and the electrostatically driven Monte Carlo (EDMC) method, we have computed the average structures of wild-type and oncogenic forms of ras-p21 alone and bound to a number of its target proteins, such as the ras-binding domain (RBD) of raf, guanine nucleotide exchange protein (GAP) and SOS guanine nucleotide exchange protein (GAP). By superimposing the average structures of the oncogenic forms on those of their wild-type counterparts, we have identified a number of domains that change conformation. These domains are potential effector domains that are involved uniquely in oncogenic ras-p21 signaling. We have therefore synthesized peptides corresponding to these domains and tested them in Xenopus laevis oocytes for their abilities to inhibit oncogenic ras-p21 selectively. Using this approach, we have identified three peptides from ras-p21 and one peptide each from the RBD of raf, GAP and SOS that selectively inhibit oncogenic but not insulin-activated wild-type ras-p21-induced oocyte maturation. We have synthesized the ras-p21 peptides attached to a penetratin leader sequence to enable cell membrane penetration and introduced these peptides into a ras-transformed pancreatic cancer cell line; these peptides, but not an unrelated negative control peptide, cause the cells to undergo complete phenotypic reversion. On the other hand, none of these peptides has any effect on the growth of untransformed pancreatic acinar cells in culture, further suggesting that they may not interfere with normal cell growth. Thus these peptides can be useful agents in the treatment of cancers. We have further used these peptides to demonstrate that oncogenic and wild-type ras-p21 proteins utilize different signal transduction pathways and to identify where these differences occur in cells.
Pincus
Matthew R
MR
Department of Pathology and Laboratory Medicine, New York Harbor VA Medical Center, 800 Poly Place, Brooklyn, NY 1209, USA. Matthew.Pincus2@med.va.gov
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001665:Binding Sites; D002471:Cell Transformation, Neoplastic; D019295:Computational Biology; D018076:DNA, Complementary / Q000378:metabolism; D015195:Drug Design; D015972:Gene Expression Regulation, Neoplastic; D011905:Genes, ras; D006801:Humans; D009154:Mutation; D009865:Oocytes / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D016283:Proto-Oncogene Proteins p21(ras) / Q000737:chemistry*; D015398:Signal Transduction; D012984:Software
D000970:Antineoplastic Agents; D018076:DNA, Complementary; D010455:Peptides; D016283:Proto-Oncogene Proteins p21(ras)