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Radiolabeled alpha(v)beta3 integrin antagonists: a new class of tracers for tumor targeting
R Haubner1, H J Wester, U Reuning
1Department of Nuclear Medicine, Women's Hospital, Institute of Organic Chemistry and Biochemistry, Technische Universität München, Munich, Germany.
Unlabelled:
The alpha(v)beta3 integrins play an important role during tumor metastasis and tumor-induced angiogenesis. Targeting of this receptor may provide information about the receptor status of the tumor and enable specific therapeutic planning. Cyclo(-Arg-Gly-Asp-D-Phe-Val-) has been shown to be a selective alpha(v)beta3 integrin antagonist with high affinity. In this study we describe the synthesis and biological evaluation of [125I]-3-iodo-D-Tyr4-cyclo(-Arg-Gly-Asp-D-Tyr-Val-) ([125I]P2), [125I]-3-iodo-Tyr5-cyclo(-Arg-Gly-Asp-D-Phe-Tyr-) ([125I]P4) and the negative control peptide [1251]-3-iodo-D-Tyr4-cyclo(-Arg-D-Ala-Asp-Tyr-Val-) ([125I]P6).
Methods:
Peptides were assembled on a solid support using fluorenylmethoxycarbonyl amino acid coupling protocols. Radioiodination was performed using the iodogen method. The in vitro binding assays were performed using isolated, immobilized alphaIIbeta3 and alpha(v)beta3 integrins. Expression of the alphaVbeta3 receptor on the different tumors was validated by immunohistochemical methods using alpha(v) and alpha(v)beta3 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 binding assays demonstrate that the introduction of tyrosine and subsequent iodination have no influence on the high affinity and selectivity for alpha(v)beta3. Immunohistochemical staining clearly indicates the presence of the alpha(v)beta3 integrins on the tumor tissue of the melanoma and the osteosarcoma. Pretreatment and displacement studies show specific binding of [125I]P2 on melanoma M21-bearing nude mice and osteosarcoma-bearing BALB/c mice but less specific binding on mammary carcinomas. [125I]P2 exhibits fast elimination kinetics. The accumulation in the tumor 10 min postinjection is 2.07 +/- 0.32 %ID/g for the melanoma M21 and 3.50 +/- 0.49 %ID/g for the osteosarcoma and decreases to 1.30 +/- 0.13 %ID/g and 2.03 +/- 0.49 %ID/g 60 min postinjection, respectively. [125I]P4 shows even faster elimination kinetics, resulting in a tumor accumulation of 0.40 +/- 0.10 %ID/g 60 min postinjection for the osteosarcoma-bearing BALB/c mice. Both peptides reveal predominately hepatobiliary excretion. For [1251]P2, this also is confirmed by autoradiography. The negative control peptide [125I]P6 shows no specific activity accumulation.
Conclusion:
[125I]P2 exhibits high affinity and selectivity for the alpha(v)beta3 integrin in vitro and in vivo and, thus, represents the first radiolabeled alpha(v)beta3 antagonist for the investigation of angiogenesis and metastasis in vivo.
Insights
This study developed a radiolabeled peptide ([125I]P2) that selectively targets alpha(v)beta3 integrins. This offers a new tool for investigating tumor metastasis and angiogenesis in vivo.
Area of Science:
- Oncology
- Radiochemistry
- Molecular Imaging
Background:
- Alpha(v)beta3 integrins are crucial in tumor metastasis and angiogenesis.
- Targeting alpha(v)beta3 integrins can aid in tumor receptor status assessment and therapeutic planning.
- Cyclo(-Arg-Gly-Asp-D-Phe-Val-) is a known high-affinity alpha(v)beta3 integrin antagonist.
Purpose of the Study:
- To synthesize and biologically evaluate novel radiolabeled alpha(v)beta3 integrin antagonists.
- To assess the in vitro and in vivo affinity and selectivity of [125I]P2 and [125I]P4.
- To establish a radiolabeled antagonist for investigating angiogenesis and metastasis.
Main Methods:
- Peptide synthesis using solid-phase Fmoc chemistry.
- Radioiodination via the iodogen method.
- In vitro binding assays with immobilized integrins.
- Immunohistochemical validation of alpha(v)beta3 expression in tumors.
- Biodistribution studies in tumor-bearing mice models (melanoma, osteosarcoma, mammary carcinoma).
Main Results:
- Iodinated peptides ([125I]P2, [125I]P4) retained high affinity and selectivity for alpha(v)beta3 integrin.
- Immunohistochemistry confirmed alpha(v)beta3 expression in melanoma and osteosarcoma tissues.
- [125I]P2 demonstrated specific binding in vivo, particularly in melanoma and osteosarcoma, with fast clearance and hepatobiliary excretion.
- Negative control peptide [125I]P6 showed no specific accumulation.
Conclusions:
- [125I]P2 is a highly effective alpha(v)beta3 integrin antagonist, showing excellent affinity and selectivity both in vitro and in vivo.
- This represents the first radiolabeled alpha(v)beta3 antagonist suitable for in vivo studies of angiogenesis and metastasis.
- [125I]P2 holds potential for diagnostic imaging and therapeutic planning in relevant cancers.