Optimization of Tetrahydroindazoles as Inhibitors of Human Dihydroorotate Dehydrogenase and Evaluation of Their Activity and In Vitro Metabolic Stability

Human
dihydroorotate dehydrogenase (DHODH), an enzyme in the de
novo pyrimidine synthesis pathway, is a target for the treatment of
rheumatoid arthritis and multiple sclerosis and is re-emerging as
an attractive target for cancer therapy. Here we describe the optimization
of recently identified tetrahydroindazoles (HZ) as DHODH inhibitors.
Several of the HZ analogues synthesized in this study are highly potent
inhibitors of DHODH in an enzymatic assay, while also inhibiting cancer
cell growth and viability and activating p53-dependent transcription
factor activity in a reporter cell assay. Furthermore, we demonstrate
the specificity of the compounds toward the de novo pyrimidine synthesis
pathway through supplementation with an excess of uridine. We also
show that induction of the DNA damage marker γ-H2AX after DHODH
inhibition is preventable by cotreatment with the pan-caspase inhibitor
Z-VAD-FMK. Additional solubility and in vitro metabolic stability
profiling revealed compound 51 as a favorable candidate
for preclinical efficacy studies.