Doctor of Philosophy
School of Chemistry
Sumner, Pamela Allison, Diagnostic and therapeutic potential of growth factor RTK inhibitors as radiopharmaceuticals in cancer, Doctor of Philosophy thesis, School of Chemistry, University of Wollongong, 2011. http://ro.uow.edu.au/theses/3342
The anilinoquinazoline compounds have proven to be effective in current cancer treatment as EGF and VEGF RTK inhibitors. They were also excellent candidates for radiopharmaceutical development (with 18F, 123I and 11C) due to the precedence of halogen substituents present in active compounds and the ease of synthesis.
A series of chemical precursors and standards were synthesized in preparation for radiochemical synthesis and biological evaluation. The Series I synthesis yielded the fluorine standards ,  and their tosylate precursors ,  in eleven steps, while the iodine standard  and stannane precursor  were obtained through the five step Series II synthesis. The carbon precursors and standards ,  -  were also produced during Series I and Series II syntheses respectively.
Radiolabelling of the fluorine-18 derivatives ,  using Kryptofix 222 was achieved in 30-38% RCY. These yields had improved significantly with changes to solvent (DMA), base (K2C2O4) and higher reaction temperature (150 °C). The iodine-123 derivative  was also successfully synthesised using electrophilic radioiodination, giving both cis and trans isomers in 71% RCY (E:Z = 8:1).
In vitro evaluation of the fluorine derivatives ,  in a kinase assay (including EGFR, VEGFR (KDR and Flt1), FGFR, PDGFR, IGFR and Akt) showed significant activity and selectivity for EGFR and VEGFR. Since EGFR had a strong inhibition profile, an IC50 was obtained for this RTK, revealing IC50 values of 1.2 nM  and 3.4 nM . In vivo imaging and biodistribution studies of the radioiodinated compound  were performed in rats expressing a mammary adenocarcinoma EGFR tumour model. There was some uptake of  in the tumour, but also signs of in vivo metabolism and deiodination.
In addition, structure-based drug design was used to investigate the EGF and VEGF receptor targets, gaining further insight into developing RTK inhibitors as radiopharmaceuticals. The crystal structures of each receptor were analysed, to find both conserved regions within the binding pockets and the differences which could provide selectivity in drug design. AutoDock studies also revealed key binding patterns within each receptor. For example, the anilinoquinazoline compounds docked into EGFR with the aniline group reaching into the hydrophobic pocket, while the 6,7-alkoxy chains stretched along the edge of the binding pocket, interacting with residues such as Lys721/Asp831/Phe832 (hydrophobic pocket) and Gly772/Cys773 (edge). Preliminary VEGFR docking studies showed a more diverse binding pattern throughout the open channel-like binding area with key binding residues, Glu915 and Cys917.
Docking analysis was also performed for the target molecules, including the fluorine and iodine derivatives. Very good results were obtained for both fluorine compounds ,  in the EGFR, which was consistent with initial in vitro evaluation. The iodine derivative was not as well suited to the EGFR due to the bulkier iodoethenyl substituent on the aniline, with slightly better results obtained in VEGFR. Therefore, the fluorine derivatives may have greater potential as radiopharmaceuticals.