title: Anticancer activity of targeted proapoptotic peptides.
abstract: Tumor-induced angiogenesis can be targeted by RGD (Arg-Gly-Asp) peptides, which bind to alpha(v)beta(3)-receptors upregulated on angiogenic endothelial cells. RGD-containing peptides are capable of inducing apoptosis through direct activation of procaspase-3 to caspase-3 in cells. Additionally, tumor cells overexpressing somatostatin receptors can be targeted by somatostatin analogs. Radiolabeled somatostatin analogs are successfully used to image and treat such tumors via receptor-targeted scintigraphy and therapy. We combined these 2 peptides, RGD and somatostatin, to synthesize a new hybrid peptide, RGD-diethylenetriaminepentaacetic acid (DTPA)-octreotate (c(Arg-Gly-Asp-D-Tyr-Asp)-Lys(DTPA)-D-Phe-c(Cys-Tyr-D-Trp-Lys-Thr-Cys)-Thr). An earlier study showed that tumor-bearing rats had high receptor-specific uptake of RGD-(111)In-DTPA-octreotate in somatostatin receptor subtype 2-positive tissues and tumors. Furthermore, RGD-(111)In-DTPA-octreotate showed a pronounced tumoricidal effect, which is probably the result of increased apoptosis, as is shown by an increased caspase-3 activity after incubation with (111)In-labeled RGD-DTPA-octreotate in comparison with the 2 monopeptides (111)In-DTPA-RGD and (111)In-DTPA-Tyr(3)-octreotate. In this study, we evaluated the biodistributions of RGD-(111)In-DTPA-octreotate and (125)I-RGD-octreotate and investigated the caspase-3 activation of the unlabeled compound RGD-DTPA-octreotate in vitro.
  
  
  METHODS
  Biodistribution studies on tumor-bearing rats were performed with RGD-(111)In-DTPA-octreotate and (125)I-RGD-octreotate. The apoptotic activity, by activation of caspase-3 with RGD-DTPA-octreotate and RGD-octreotate, was examined using colorimetric and immunocytochemical assays.
  
  
  RESULTS
  The radiolabeled compound, RGD-(111)In-DTPA-octreotate, showed high uptake and retention in the rats in which rat pancreatic CA20948 tumor had been implanted. A major drawback was high renal uptake. In vitro, the unlabeled peptide RGD-DTPA-octreotate induced a significant increase in caspase-3 levels in various cell lines in comparison with RGD and Tyr(3)-octreotate (P < 0.01). Caspase-3 activation was time dependent. To alter the elimination route, we examined the biodistribution of radioiodinated RGD-octreotate without DTPA [c(Arg-Gly-Asp-D-Tyr-Asp)-D-Phe-c(Cys-Tyr-D-Trp-Lys-Thr-Cys)-Thr], as a model of unlabeled RGD-octreotate, in tumor-bearing rats. (125)I-RGD-octreotate showed a much lower renal uptake than did RGD-(111)In-DTPA-octreotate. Furthermore, the affinity of RGD-octreotate increased in comparison with RGD-DTPA-octreotate (values of 1.4 x 10(-8) mol/L vs. 9.4 x 10(-8) mol/L, respectively, for inhibitory concentration of 50%). Finally, RGD-octreotate was still able to activate caspase-3, as was indicated with immunocytochemistry.
  
  
  CONCLUSIONS
  Because of the high renal uptake, RGD-(111)In-DTPA-octreotate is unsuitable for radionuclide therapy. However, the unlabeled peptides, RGD-DTPA-octreotate and RGD-octreotate, also induced an increase in caspase-3 levels, indicating the therapeutic potential of this compound. Thus, the development of hybrid molecules can become a new approach in the treatment of cancer.
authors.affiliation: Department of Nuclear Medicine, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Astrid
authors.initials: A
authors.lastname: Capello
authors.forename: Eric P
authors.initials: EP
authors.lastname: Krenning
authors.forename: Bert F
authors.initials: BF
authors.lastname: Bernard
authors.forename: Wout A P
authors.initials: WA
authors.lastname: Breeman
authors.forename: Jack L
authors.initials: JL
authors.lastname: Erion
authors.forename: Marion
authors.initials: M
authors.lastname: de Jong
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000493:pharmacokinetics / Q000627:therapeutic use; D051017:Apoptosis Regulatory Proteins / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D016503:Drug Delivery Systems / Q000379:methods; D007205:Indium Radioisotopes / Q000493:pharmacokinetics / Q000627:therapeutic use; D008297:Male; D008657:Metabolic Clearance Rate; D009842:Oligopeptides / Q000493:pharmacokinetics* / Q000627:therapeutic use*; D009928:Organ Specificity; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology / Q000532:radiotherapy*; D016499:Radioimmunotherapy / Q000379:methods; D019275:Radiopharmaceuticals / Q000493:pharmacokinetics; D051381:Rats; D011917:Rats, Inbred Lew; D014018:Tissue Distribution; D016896:Treatment Outcome
chemical_list: D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D007205:Indium Radioisotopes; D009842:Oligopeptides; D019275:Radiopharmaceuticals; C047981:arginyl-glycyl-aspartic acid