[11C]nintedanib as TKI-PET tracer for angiogenesis imaging in vivo.

1153 Objectives PET imaging is a unique technique to assess the in vivo distribution and targeting of radiolabeled anti-cancer drugs to form predictive markers for therapy, as was demonstrated with radiolabeled EGFR targeting tyrosine kinase inhibitors (TKIs).[1, 2] To extend our knowledge on the tumor targeting of radiolabeled TKIs and its potential predictive value for therapy, we developed radiolabeled anti-angiogenesis tracer [11C]nintedanib targeting VEGFR, PDGFR and FGFR and analyzed the in vivo behavior in tumor xenografted mice. As is known from literature, nintedanib has a carboxylic acid as primary metabolite with anti-tumor activity and increased tumor retention.[3] Therefore LC-MS/MS was used to detect the formation of intra-tumoral [11C]nintedanib metabolites and to correlate this with the tumor targeting of this PET tracer. Methods Synthesis of [11C]nintedanib was achieved by methylation of the precursor using [11C]MeI under basic conditions. To establish in vivo tumor targeting, [11C]nintedanib was analysed in xenografted mice bearing FaDu or in house cultured SCC-VU-OE tumors, for which target expression was established by immunohistochemical staining. LC-MS/MS in combination with Multi-Reaction Monitoring (MRM) on an AB Sciex QTRAP 5500 mass spectrometer was used for the characterization of [11C]nintedanib and its metabolites in vivo. The specific fragmentation profile of [11C]nintedanib and the therapeutically active carboxylic acid metabolite allowed the detection of both compounds at picomolar concentrations and thus at tracer level. Results [11C]Nintedanib was obtained in 25.6% ± 3.3% yield by methylation with [11C]MeI in DMF and K2CO3 as base at 80oC for 5 min. In biodistribution experiments, [11C]nintedanib accumulated up to 1.66% ID/g in FaDu xenografts at 60 min post injection compared to 0.7% ID/g in the SCC-VU-OE xenografts. Metabolite analysis from these biodistribution experiments displayed the formation of a polar metabolite from [11C]nintedanib, hinting towards the formation of the described active carboxylic acid. Therefore, incubation experiments in mice and rat plasma were conducted which indeed revealed the formation of the described active carboxylic acid from nintedanib up to 60%, however, in human plasma this metabolite could not be identified. Unexpectedly, the in vivo metabolite analysis on the tumor tissue did not reveal carboxylic acid formation from [11C]nintedanib when the developed MRM LC-MS/MS method was used. Therefore, based on these results a potential role of the carboxylic acid metabolite in tumor targeting and therapeutic efficacy remains uncertain. Conclusions [11C]Nintedanib was synthesized in high and reliable yields. Encouraging results were obtained from biodistribution experiments in FaDu xenografted mice. Nevertheless, it should be noted that it is unclear whether the active carboxylic acid metabolite from [11C]nintedanib is involved in antitumor activity since this metabolite formation could not be detected using MRM LC-MS/MS. A possible explanation for the relatively low accumulation of [11C]nintedanib and absence of [11C]nintedanib metabolites might be the absence of human vasculature and thus kinase receptors of human origin in xenografted tumors in mice. Research Support: This research was financially supported by VUMC-CCA References: [1] J. Baselga et al., Science, 2006, 312, 1175; [2] P. Slobbe et al., Drug Discov. Today, 2012, 17, 1175 [3] F. Hilberg et al., Cancer Res., 2008, 68, 4774. $$graphic_D0D616D7-E66B-4D9F-8251-0225F7E10018$$