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 (99m)Tc via nitrido and carbonyl precursors. The (99m)Tc-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 (99m)Tc complexes were formed in high yields and were found to be stable. HPLC pattern of (99m)Tc nitrido complex of cysteine-cysteine-annexine 13 (CC-Anx13) and (99m)Tc carbonyl complex of histdine-annexin 13 (H-Anx13) revealed the formation of single species. In vitro cell uptake studies with (99m)Tc 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 (99m)Tc nitrido complex with CC-Anx13 in fibrosarcoma tumor-bearing Swiss mice revealed optimum tumor uptake of 0.52 (0.17) %ID/g at 1 h pi.
Conclusion: (99m)Tc(N)-CC-anx13 showed specific uptake in apoptotic tumor cells and warrants further evaluation.