99mTc-DTPA-bis-c(RGDfK) a potential alpha(v)beta3 integrin based homobivalent radioligand for imaging neoangiogenesis in malignant glioma and melanoma

Abstract

α_vβ₃ integrin is a marker of tumor neoangiogenesis that specifically binds to RGD containing peptides. Hence, the present study is focused on the development of ^99mTc-labeled bivalent DTPA-bis-c(RGDfK) conjugate and its preclinical evaluation on human tumor cell lines expressing α_vβ₃ targets. Homobivalent DTPA-bis-c(RGDfK) was prepared and assessed for its affinity and specificity in α_vβ₃ positive (and negative) receptor cell lines. DTPA-bis-c(RGDfK) conjugate was labeled with ^99mTc and subjected to cells/tissue sections. Localization of α_vβ₃ expression was corroborated using immunostaining and ex vivo imaging of the distribution pattern of ^99mTc-DTPA-bis-c(RGDfK). The radiolabeling of the DTPA-bis-c(RGDfK) with ^99mTc is obtained after 45 min at 95 °C with a radiochemical yield of about 45%. Radiochemical purity was >95% after C18 Oasis HLB cartridge purification with specific activity of 1475 GBq mmoL⁻¹. In vitro experiments showed high affinity and specificity for α_vβ₃ with IC₅₀ of 32.86 ± 7.83 nmol L⁻¹. Ex vivo imaging on tissue sections confirmed preliminary specificity results. In vivo analysis in a mouse model showed that this tracer was able to detect and readily identify U87MG and B16F10 α_vβ₃ positive tumors 60 minutes post-injection. c(RGDfK) blocking experiments confirmed its excellent affinity and specificity to α_vβ₃ receptors in U87MG tumors. The radiotracer was mainly excreted through the renal route with minimal radioactivity being excreted through the hepatobiliary route. ^99mTc-DTPA-bis-c(RGDfK) can be an excellent scintigraphic agent for imaging of α_vβ₃ receptors being expressed in abundance in malignant glioma and melanoma cancer.