Potential SPECT imaging radiotracers targeting necroptosis: I-123 labeled necrostatin-1 and necrostatin-3 derivatives

Objectives Comparing with the two well-established biochemical cell death pathways apoptosis and necrosis, necroptosis is a relatively novel concept. Our goals are to develop radiolabeled necrostatin (Nec) derivatives, which act as imaging agents targeting necroptosis, to help diagnose diseases and monitor their treatment.

Methods 7-¹²³I-Nec-1 was synthesized by Cu(I) catalyzed halogen exchange reaction with precursor 7-Br-Nec-1. Three ¹²³I labeled Nec-3 derivatives (R isomers, 8-¹²³I-Nec-3R, 3’-¹²³I-Nec-3R, and 4’-¹²³I- Nec-3R) were prepared by iodo-destannylation reactions through tributyltin precursors. The stability of the tracers was tested in rat serum at 37 °C for 1 day. In vitro cellular uptake of 7-¹²³I-Nec-1was assessed in h9c2 cardiomyocytes, treated with TNFα and zVAD to stimulate necroptosis. The biodistribution properties of 7-¹²³I-Nec-1and 3’-¹²³I-Nec-3R were evaluated in normal rats at 2 hr post injection (p.i.).

Results 7-¹²³I-Nec-1 and three ¹²³I labeled Nec-3 derivatives were synthesized and purified by RP-HPLC to achieve > 95% radiochemical purity. All tracers were stable in rat serum for 1 day. 7-¹²³I-Nec-1 showed higher uptake in stimulated necroptotic h9c2 cardiomyocytes compared with control cells. 3'-¹²³I-Nec-3R was mainly excreted through liver and intestine, while 7-¹²³I-Nec-1was mostly cleared through kidney and urine, which may related to the higher lipophilicity of the Nec-3 compound. The stomach and thyroid uptake of 3'-¹²³I-Nec-3R was significantly lower than 7-¹²³I-Nec-1, indicating its better in vivo stability. 3'-¹²³I-Nec-3R also showed faster blood clearance, which may be beneficial for in vivo SPECT vascular imaging studies.

Conclusions Four Nec compounds were successfully labeled with ¹²³I. 7-¹²³I-Nec-1 was taken by necroptotic h9c2 cardiomyocytes. 3'-¹²³I-Nec-3R showed less deiodination and a more favorable biodistribution profile than 7-¹²³I-Nec-1. These ¹²³I-Nec tracers are potential SPECT imaging agents targeting necroptosis.