Abstract
Ribociclib (
RBC
, Kisqali®) is a highly selective CDK4/6 inhibitor that has been approved for breast cancer therapy. Initially, prediction of susceptible sites of metabolism and reactivity pathways were performed by the StarDrop WhichP450™ module and the Xenosite web predictor tool, respectively. Later,
in vitro
metabolites and adducts of
RBC
were characterized from rat liver microsomes using LC-MS/MS. Subsequently,
in silico
data was used as a guide for the
in vitro
work. Finally,
in silico
toxicity assessment of
RBC
metabolites was carried out using DEREK software and structural modification was proposed to reduce their side effects and to validate the bioactivation pathway theory using the StarDrop DEREK module.
In vitro
phase I metabolic profiling of
RBC
was performed utilizing rat liver microsomes (RLMs). Generation of reactive metabolites was investigated using potassium cyanide (KCN) as a trapping nucleophile for the transient and reactive iminium intermediates to form a stable cyano adduct that can be identified and characterized using mass spectrometry. Nine phase I metabolites and one cyano adduct of
RBC
were characterized. The proposed metabolic pathways involved in generation of these metabolites are hydroxylation, oxidation and reduction. The reactive intermediate generation mechanism of
RBC
may provide an explanation of its adverse reactions. Aryl piperazine is considered a structural alert for toxicity as proposed by the DEREK report. We propose that the generation of only one reactive metabolite of
RBC
in a very small concentration is due to the decreased reactivity of the piperazine ring compared to previous reports of similar drugs. Docking analysis was performed for
RBC
and its proposed derivatives at the active site of the human CDK6 enzyme.
Methyl-RBC
exhibited the best ADMET and docking analysis and fewer side effects compared to
RBC
and
fluoro-RBC
. Further drug discovery studies can be conducted taking into account this concept allowing the development of new drugs with enhanced safety profiles that were confirmed by using StarDrop software. To the best of our knowledge, this is the first literature report of
RBC
in vitro
metabolic profiling and structural characterization and toxicological properties of the generated metabolites.