In Vitro and in Vivo Antitumor Effects of Cytotoxic Camptothecin-Bombesin Conjugates Are Mediated by Specific Interaction with Cellular Bombesin Receptors

摘要

Most human tumors overexpress or ectopically express peptide hormoneeurotransmitter receptors,which are being increasingly studied as a means to selectively deliver cytotoxic agents.Although a number of peptide ligand-constructs demonstrate tumor cytotoxicity,the role of specific tumoral receptor interaction in its mediation is unclear.To address this question,we synthesized camptothecin(CPT)bombesin(Bn)analogs,in which CPT was coupled via a novel carbamate linker,L2 [N-(N-methyl-amino-ethyl)-glycine carbamate],that were chemically similar but differed markedly in their potency/affinity for human Bn receptors.We then examined their ability to interact with Bn receptors and cause in vitro and in vivo tumor cytotoxicity.CPT-L2-[D-Tyr~6,beta-Ala~(11),D-Phe~(13),Nle~(14)] Bn(6-14)(BA3)bound with high affinity and had high potency for all three human Bn receptor subtypes,whereas CPT-L2-[D-Tyr~6,beta-Ala~(11),D-Phe~(13),Nle~(14)] Bn(6-14)[ D-Phe-CPT-L2-BA3] had >1400-fold lower affinity/potency.~(125)I-CPT-L2-BA3 but not ~(125)I-D-Phe-CPT-L2-BA3 was internalized by Bn receptor subtype-containing cells.CPT-L2-BA3 displayed significantly more cytotoxicity than D-Phe-CPT-L2-BA3 toward NCI-H1299 lung cancer cells in both 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazo-lium bromide and clonogenic assays and more potently inhibited H1299 xenograft growth in nude mice.CPT-L2-BA3 was also metabolically more stable than its parent peptide and inhibited growth of a number of other tumor cell lines in vitro and in vivo.These results demonstrate that specific tumoral receptor interaction is important in mediating the ability of peptide ligand-cytotoxic constructs to cause cytotoxicity.Because many tumors overexpress Bn receptors,these results also demonstrate that CPT-L2-BA3 will be a useful agent for delivering receptor-mediated cytotoxicity to many different human tumors.