Apart from its high affinity the selection of GW as
Apart from its high affinity, the selection of GW2580 as IWR-1-endo receptor lead in our radiotracer development program was motivated by its exceptional selectivity. The observed profound impact of small structural modifications on the selected panel of four kinases tested prompted us to conduct comprehensive kinase selectivity profiling of our fluorinated inhibitors. Compound , as well as the less potent derivative , were tested in enzymatic assays on a panel of 342 kinases (Reaction Biology Corporation, full wild type kinase panel, , ). In the presence of 1.0μM of , only TrkB, TrkC and CSF-1R were strongly inhibited—remaining activity inferior to 25% compared to DMSO control (A). Compound also showed moderate TrkA inhibition (55.7±1.4% remaining activity) and negligible inhibition of only few other kinases (CDK6, HGK/MAP4K4, TXK). A similar kinase profile was obtained for with few more off-target minor inhibitions (Fig. B, 10–25% inhibition: CDK6, CK1g3, DAPK2, ERK5/MAPK7, HGK/MAP4K4, SNARK/NUAK2, TXK). Those results confirmed the retention of the high selectivity of towards TrkB/TrkC/CSF-1R relative to the non-fluorinated lead and encourage its further development into a radiolabeled probe.
In preliminary radiolabeling experiments, [F] was prepared via the alkylation of precursor with [F]fluorobenzyl bromide ([F], , ). Alkylation leading to [F] was performed in radiochemical yield (RCY) of 13% (HPLC incorporation yield, non-decay corrected) in the presence of CsCO and tetrabutylammonium iodide (TBAI) at 100°C for 10min. [F] was obtained following an on-cartridge procedure adapted form Lemaire et al. starting with 4-formyl--trimethylanilinium triflate (). The radiosynthesis of [F] was typically accomplished in 25–30% RCY (non-decay corrected) as a crude mixture (>85% [F] containing residual 4-[F]fluorobenzaldehyde—[F], ) which was used directly in the alkylation step. The multi-step approach used, carried out manually, despite being sufficient for evaluation purposes, will be difficult to implement into an automated synthesis unit for routine production. Therefore, an alternative and more straightforward route to [F] will be developed, such as a diaryliodonium salt strategy.
In conclusion, starting from the known inhibitor a potent and highly selective fluorinated TrkB/TrkC/CSF-1R inhibitor, , was designed and labeled with fluorine-18. The remarkable selectivity profile of was confirmed by exhaustive kinase profiling. Those initial results, together with our preliminary radiosynthesis study, warrant further evaluation of [F] as a uniquely selective tool to assess TrkB, TrkC and CSF-1R level in vivo with PET. Imaging studies in Trk-positive tumor bearing nude mice, especially TrkB-overexpressing neuroblastoma xenograft models, will be reported in due course.
Macrophages are characterized as classical M1- and alternative M2-subtypes depending on how they influence the immune response. M1 macrophages influence the inflammatory response and anti-tumor immunity, whereas M2 macrophages are involved in immunosuppressive and tumor promoting activities. Tumor-associated macrophages (TAMs), which are classified as M2-polarized macrophages, are recruited by tumor cells and infiltrate into the stromal environment of malignant tumors. TAMs in the tumor microenvironment are known to play a crucial role in tumor progression, including tumor initiation and development, matrix remodeling and metastasis, and immune suppression., Colony stimulating factor-1 (known as CSF-1 or macrophage-colony stimulating factor, M-CSF) is a growth factor for monocyte and macrophage-derived cells. CSF-1 and its receptor, colony stimulating factor-1 receptor (CSF-1R or M-CSFR, also known as FMS) have been reported as having important roles in regulating the production, migration, differentiation, survival, and function of macrophages and their precursors. Therefore, over-activation of CSF‐1/CSF-1R signaling in macrophages has been implicated in many disease states including osteoclast proliferation in bone osteolysis, and a number of inflammatory disorders as well as the growth/metastasis of cancer., , , , , , , , , Especially, a number of CSF1/CSF1R-targeting agents such as Pexidartinib and ARRY-382 are undergoing clinical trials in several cancer including breast cancer, prostate cancer, and melanoma.