Radiolabeled alpha(v)beta(3) integrin antagonists: A new class of tracers for tumor targeting

The alpha(v)beta(3) integrins play an important role during tumor metastasi s and tumor-induced angiogenesis. Targeting of this receptor may provide in formation about the receptor status of the tumor and enable specific therap eutic planning. Cyclo(-Arg-Gly-Asp-D-Phe-Val-) has been shown to be a selec tive alpha(v)beta(3) integrin antagonist with high affinity. In this study we describe the synthesis and biological evaluation of [I-125]-3-iodo-D-Tyr (4)-cyclo(-Arg-Gly-Asp-D-Tyr-Val-) ([I-125]P2), [I-125]-3-iodo-Tyr(5)-cyclo (-Arg-Gly-Asp-D-Phe-Tyr-) ([I-125]P4) and the negative control peptide [I-1 25]-3-iodo-D-Tyr(4)-cyclo(-Arg-D-Ala-Asp-Tyr-Val-) ([I-125]P6). Methods: Pe ptides were assembled on a solid support using fluorenylmethoxycarbonyl ami no acid coupling protocols. Radioiodination was performed using the iodogen method. The in vitro binding assays were performed using isolated, immobil ized alpha(IIb)beta(3) and alpha(v)beta(3) integrins. Expression of the alp ha(v)beta(3) receptor on the different tumors was validated by immunohistoc hemical methods using alpha(v) and alpha(v)beta(3) specific antibodies. For biodistribution studies, nude mice with melanoma M21 or mammary carcinoma MaCaF and BALB/c mice with osteosarcoma were used. Results: The in vitro bi nding assays demonstrate that the introduction of tyrosine and subsequent i odination have no influence on the high affinity and selectivity for alpha( v)beta(3). Immunohistochemical staining clearly indicates the presence of t he alpha(v)beta(3) integrins on the tumor tissue of the melanoma and the os teosarcoma. Pretreatment and displacement studies show specific binding of [I-125]P2 on melanoma M21-bearing nude mice and osteosarcoma-bearing BALB/c mice but less specific binding on mammary carcinomas. [I-125]P2 exhibits f ast elimination kinetics. The accumulation in the tumor 10 min postinjectio n is 2.07 +/- 0.32 %ID/g for the melanoma M21 and 3.50 +/- 0.49 %ID/g for t he osteosarcoma and decreases to 1.30 +/- 0.13 %ID/g and 2.03 +/- 0.49 %ID/ g 60 min postinjection, respectively. [I-125]P4 shows even faster eliminati on kinetics, resulting in a tumor accumulation of 0.40 +/- 0.10 %ID/g 60 mi n postinjection for the osteosarcoma-bearing BALB/c mice. Both peptides rev eal predominately hepatobiliary excretion. For [I-125]P2, this also is conf irmed by autoradiography. The negative control peptide [I-125]P6 shows no s pecific activity accumulation. Conclusion: [I-125]P2 exhibits high affinity and selectivity for the alpha(v)beta(3) integrin in vitro and in vivo and, thus, represents the first radiolabeled alpha(v)beta(3) antagonist for the investigation of angiogenesis and metastasis in vivo.