Synthesis and evaluation of a 99mTc-MAMA-propyl-thymidine complex as a potential probe for in vivo visualization of tumor cell proliferation with SPECT

IntroductionCytosolic thymidine kinase (TK1) catalyzes phosphorylation of thymidine to its monophosphate. TK1 activity is closely related with DNA synthesis, and thymidine analogs derivatized with bulky carboranylalkyl groups at the N-3 position were reported to be good substrates for TK1. Accordingly, we have synthesized 99mTc-MAMA-propyl-thymidine and evaluated it as a potential tumor tracer.MethodsThe bis(S-trityl)-protected MAMA-propyl-thymidine precursor (3-N-[S-trityl-2-mercaptoethyl]-N-[N′-(S-trityl-2-mercaptoethyl)amidoacetyl]-aminopropyl-thymidine) was prepared in three steps, and its structure was confirmed with 1H NMR and mass spectrometry. Deprotection of the thiols and labeling with 99mTc were done in a two-step, one-pot procedure, yielding 99mTc-MAMA-propyl-thymidine, which was analyzed with high-performance liquid chromatography, radio-LC-MS analysis (ESI+) and electrophoresis, and its log P was determined. The biodistribution in normal mice was evaluated, and its biodistribution in a radiation-induced fibrosarcoma (RIF) tumor mouse was compared with that of 3′-deoxy-3′-[18F] fluorothymidine [18F]FLT.Results99mTc-MAMA-propyl-thymidine was obtained with a radiochemical yield of 70%. Electrophoresis indicated that the complex is uncharged, and its log P was 1.0. The molecular ion mass of the Tc complex was 589 Da, which is compatible with the hypothesized N2S2-oxotechnetium structure. Tissue distribution showed fast clearance from plasma primarily by the hepatobiliary pathway. Whole-body planar imaging after injection of 99mTc-MAMA-propyl-thymidine in an RIF tumor-bearing mouse showed high uptake in the liver and the intestines. No uptake was observed in the tumor, in contrast to the clear uptake observed for [18F] FLT visualized with μPET.ConclusionsAlthough it has been reported that TK1 accepts large substituents at the N-3 position of the thymine ring, the results of this study show that 99mTc-MAMA-propyl-thymidine cannot be used as a single photon emission computed tomography tumor tracer, probably because the 99mTc-MAMA ligand is too bulky to be tolerated by TK1.