Dataset Information

Generation and characterization of a mitotane-resistant adrenocortical cell line.

ABSTRACT: Mitotane is the only drug approved for the therapy of adrenocortical carcinoma (ACC). Its clinical use is limited by the occurrence of relapse during therapy. To investigate the underlying mechanisms in vitro, we here generated mitotane-resistant cell lines. After long-term pulsed treatment of HAC-15 human adrenocortical carcinoma cells with 70 µM mitotane, we isolated monoclonal cell populations of treated cells and controls and assessed their respective mitotane sensitivities by MTT assay. We performed exome sequencing and electron microscopy, conducted gene expression microarray analysis and determined intracellular lipid concentrations in the presence and absence of mitotane. Clonal cell lines established after pulsed treatment were resistant to mitotane (IC50 of 102.2 ± 7.3 µM (n = 12) vs 39.4 ± 6.2 µM (n = 6) in controls (biological replicates, mean ± s.d., P = 0.0001)). Unlike nonresistant clones, resistant clones maintained normal mitochondrial and nucleolar morphology during mitotane treatment. Resistant clones largely shared structural and single nucleotide variants, suggesting a common cell of origin. Resistance depended, in part, on extracellular lipoproteins and was associated with alterations in intracellular lipid homeostasis, including levels of free cholesterol, as well as decreased steroid production. By gene expression analysis, resistant cells showed profound alterations in pathways including steroid metabolism and transport, apoptosis, cell growth and Wnt signaling. These studies establish an in vitro model of mitotane resistance in ACC and point to underlying molecular mechanisms. They may enable future studies to overcome resistance in vitro and improve ACC treatment in vivo.

SUBMITTER: Seidel E

PROVIDER: S-EPMC6993260 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundMitotane is the only approved treatment for patients with adrenocortical carcinoma (ACC). A better explanation for the variability in the pharmacokinetics (PK) of mitotane, and the optimization and individualization of mitotane treatment, is desirable for patients.ObjectivesThis study aims to develop a population PK (PopPK) model to characterize and predict the PK profiles of mitotane in patients with ACC, as well as to explore the effect of genetic variation on mitotane clearance. Ultimately, we aimed to facilitate mitotane dose optimization and individualization for patients with ACC.MethodsMitotane concentration and dosing data were collected retrospectively from the medical records of patients with ACC taking mitotane orally and participating in the Dutch Adrenal Network. PopPK modelling analysis was performed using NONMEM (version 7.4.1). Genotypes of drug enzymes and transporters, patient demographic information, and clinical characteristics were investigated as covariates. Subsequently, simulations were performed for optimizing treatment regimens.ResultsA two-compartment model with first-order absorption and elimination best described the PK data of mitotane collected from 48 patients. Lean body weight (LBW) and genotypes of CYP2C19*2 (rs4244285), SLCO1B3 699A>G (rs7311358) and SLCO1B1 571T>C (rs4149057) were found to significantly affect mitotane clearance (CL/F), which decreased the coefficient of variation (CV%) of the random inter-individual variability of CL/F from 67.0 to 43.0%. Fat amount (i.e. body weight - LBW) was found to significantly affect the central distribution volume. Simulation results indicated that determining the starting dose using the developed model is beneficial in terms of shortening the period to reach the therapeutic target and limit the risk of toxicity. A regimen that can effectively maintain mitotane concentration within 14-20 mg/L was established.ConclusionsA two-compartment PopPK model well-characterized mitotane PK profiles in patients with ACC. The CYP2C19 enzyme and SLCO1B1 and SLCO1B3 transporters may play roles in mitotane disposition. The developed model is beneficial in terms of optimizing mitotane treatment schedules and individualizing the initial dose for patients with ACC. Further validation of these findings is still required.

Dataset Information

Generation and characterization of a mitotane-resistant adrenocortical cell line.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets