Development of 68Ga-labeled Novel Dasatinib Analogues for PET Imaging of Discoidin Domain Receptors

Introduction: The discoidin domain receptors (DDRs), including DDR1 and DDR2, belong to an unique subfamily of receptor tyrosine kinases, which are characterized by the binding of collagen to their extracellular domain for signaling pathways activation. These DDRs have been crucial for the regulation of cellular functions such as adhesion, migration, proliferation, differentiation, survival, matrix remodeling and oncogenic transformation. Their dysregulation has led to the progression of various human diseases including inflammation, fibrosis and cancer. DDRs have therefore been considered as attractive targets for drug discovery. Despite the availability of DDRs inhibitors, their efficacy in treatment of these disorders remains to be ambiguous. Imaging of DRRs using radiolabeled inhibitors has been proposed for target engagement, dose optimization, patient selection and therapeutic effect prediction. Here, novel dasatinib analogues were designed, synthesized and radiolabelled with Ga-68 for PET imaging of DDRs in vivo for the first time.

Methods: Two amino carboxylate analogues of dasatinib were designed, synthesized and radiolabelled with ⁶⁸GaCl₃ to obtain radiotracers [⁶⁸Ga]Ga-PEMPC and [⁶⁸Ga]Ga-AEMPC.The radiochemical yields, radiochemical purity, molar activities and in vitro stability were assessed by raido-HPLC. The partition coefficient (log P) values were determined through measuring the distribution of radioactivity in 1-octanol and PBS (pH 7.4). Small-animal PET imaging studies in normal mice and acute lung injury models were performed to verify the pharmacokinetics and DDRs-targeted imaging capability in vivo.

Results: Two amino carboxylate analogues of dasatinib were designed as specific DDRs inhibitors, synthesized and characterized by ¹H-NMR and HRMS. [⁶⁸Ga]Ga-PEMPC and [⁶⁸Ga]Ga-AEMPC were successfully prepared with a high radiochemical yield of 81.6 ± 9.5 % and 90.3 ± 1.1 % (n = 4), radiochemical purity of > 99 % (n = 3) and molar activity of 42.5 ± 6.1 GBq/μmol and 37.3 ± 5.5 GBq/μmol (n = 4), respectively. The Log P values were calculated to be -1.14 ± 0.15 and -2.25 ± 0.14 (n = 6), indicating their hydrophilicity. Both [⁶⁸Ga]Ga-PEMPC and [⁶⁸Ga]Ga-AEMPC were confirmed by non-radioactive standard compounds and their excellent in vitro stability in saline and serum within 4 h was proved by radio-HPLC analysis. PET imaging in normal mice revealed the primary distribution of [⁶⁸Ga]Ga-PEMP in liver and intestine as well as [⁶⁸Ga]Ga-AEMPC in liver and kindey, demonstrated that the differences in hydrophilicity lead to changes in metabolism pathway. In experimental acute lung injury models, the uptake of [⁶⁸Ga]Ga-PEMPC and [⁶⁸Ga]Ga-AEMPC was significantly increased in lung inflammation regions, as compared to control and blocking groups, indicating the specificity and targeted imaging capability of two tracers.

Conclusions: Two radiotracers [⁶⁸Ga]Ga-PEMPC and [⁶⁸Ga]Ga-AEMPC have been firstly developed and preliminarily demonstrated their specificity and imaging capability for DDRs in vivo, which support their further validation for specific PET imaging of DDRs expression in preclinical and clinical settings.