99mTc-labeled annexin V fragments:: a potential SPECT radiopharmaceutical for imaging cell death

Introduction, Annexin V is a protein that binds to phosphatidylserine exposed on dying cells. The phosphatidylserine-specific sequence is attributed to a chain on the N-terminal of annexin consisting of 13 amino acid sequence. Radiolabeled annexin V is used for imaging apoptosis. Methods: With an aim to synthesize a probe that can detect cell death akin to annexin V but smaller in size, annexin-13 fragments were derivatized to contain cysteine, cysteine-cysteine and histidine in their sequence at N terminal and were labeled with Tc-99m via nitrido and carbonyl precursors. The Tc-99m-labeled annexin-13 derivatives were characterized by HPLC and studied for their stability. In vitro and in vivo studies were carried out in apoptotic HL-60 cells and fibrosarcoma tumor-bearing Swiss mice, respectively. Results: The Tc-99m complexes were formed in high yields and were found to be stable. HPLC pattern of Tc-99m nitrido complex of cysteine cysteine-annexine 13 (CC-Anxi 3) and Tc-99m carbonyl complex of histdine-annexin 13 (H-Anx13) revealed the formation of single species. In vitro cell uptake studies with Tc-99m nitrido complex of cysteine-cysteine-annexin 13 fragment showed 6.5% uptake in apoptotic HL-60 cells. The uptake was found to be specific on testing with apoptotic HL-60 cells. Biodistribution studies of Tc-99m nitrido complex with CC-Anx 13 in fibrosarcoma tumor-bearing Swiss mice revealed optimum tumor uptake of 0.52 (0.17) %ID/g at 1 h pi. Conclusion: Tc-99m(N)-CC-anx 13 showed specific uptake in apoptotic tumor cells and warrants further evaluation. (c) 2006 Elsevier Inc. All rights reserved.