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In this article, we report our efforts towards improving in vitro human clearance in a series of 5-azaquinazolines through a series of C4 truncations and C2 expansions. Extensive DMPK studies enabled us to tackle high Aldehyde Oxidase (AO) metabolism and unexpected discrepancies in human hepatocyte and liver microsomal intrinsic clearance. Our efforts culminated with the discovery of 5-azaquinazoline 35, which also displayed exquisite selectivity for IRAK4, and showed synergistic in vitro activity against MyD88/CD79 double mutant ABC-DLBCL in combination with the covalent BTK inhibitor acalabrutinib.

Rationale To capture all metabolites in metabolite identification studies, MS/MS information is required in both positive and negative ionization mode, usually involving several sample injections to gain all information about samples. A high-resolution and high mass accuracy quadrupole/linear trap/Orbitrap tribrid instrument was used to gain this information in a novel single injection 'capture-all' approach to metabolite identification. Methods Diclofenac, a model compound, was incubated in human and rat hepatocytes. These incubated samples were run using an ultrahigh-performance liquid chromatography/ultraviolet (UHPLC-UV) system coupled to a Thermo Fusion tribrid mass spectrometer. Five parallel scans were used: positive and negative ion full scan, data-dependent MS/MS, both high energy dissociation and collision-induced dissociation, and data-independent all ion fragmentation (AIF) spectra were collected in positive and negative ion mode. Results Nine metabolites were identified; a metabolite observed in the UV trace, but not positive ion full scan MS, was detected in the same sample injection by negative ion full scan MS. This was identified as a sulphate metabolite, and the corresponding negative ion AIF allowed for some structural elucidation. The use of a photo-diode array (PDA) detector allowed for spectral assessment in case of changes in absorbance spectra, and the subsequent semi-quantification of metabolites. Conclusions This method provided good-quality MS/MS data across the m/z range in both positive and negative ion mode. The addition of both negative ion full scan MS and negative ion MS/MS allowed for the detection and structural elucidation of metabolites not observed in positive ion mode. The use of the PDA detector allowed for the semi-quantification of metabolites.

Computer simulation of chemical speciation has been used to assess the ability of the readily-biodegradable ligand SS-EDDS4− to replace the non readily-biodegradable ligand EDTA4−. Potential replac...