Metabotypes of breast cancer cell lines revealed by non-targeted metabolomics.

We present an analysis of intracellular metabolism by non-targeted, high-throughput metabolomics profiling of 18 breast cell lines. We profiled >900 putatively annotated metabolite ions for >100 samples collected under both normoxic and hypoxic conditions and revealed extensive heterogeneity across all metabolic pathways and cell lines. Cell line-specific metabolome profiles dominated over patterns associated with malignancy or with the clinical nomenclature of breast cancer cells. Such characteristic metabolome profiles were reproducible across different laboratories and experiments and exhibited mild to robust changes with change in experimental conditions. To extract a functional overview of cell line heterogeneity, we devised an unsupervised metabotyping procedure that for each pathway automatically recognized metabolic types from metabolome data and assigned cell lines. Our procedure provided a condensed yet global representation of cell line metabolism, revealing the fine structure of metabolic heterogeneity across all tested pathways and cell lines. In follow-up experiments on selected pathways, we confirmed that different metabolic types correlated to differences in the underlying fluxes and difference sensitivity to gene knockdown or pharmacological inhibition. Thus, the identified metabotypes recapitulated functional differences at the pathway level. Metabotyping provides a powerful compression of multi-dimensional data that preserves functional information and serves as a resource for reconciling or understanding heterogeneous metabolic phenotypes or response to inhibition of metabolic pathways.

Phenobarbital induces cell cycle transcriptional responses in mouse liver humanized for constitutive androstane and pregnane x receptors.

The constitutive androstane receptor (CAR) and the pregnane X receptor (PXR) are closely related nuclear receptors involved in drug metabolism and play important roles in the mechanism of phenobarbital (PB)-induced rodent nongenotoxic hepatocarcinogenesis. Here, we have used a humanized CAR/PXR mouse model to examine potential species differences in receptor-dependent mechanisms underlying liver tissue molecular responses to PB. Early and late transcriptomic responses to sustained PB exposure were investigated in liver tissue from double knock-out CAR and PXR (CAR(KO)-PXR(KO)), double humanized CAR and PXR (CAR(h)-PXR(h)), and wild-type C57BL/6 mice. Wild-type and CAR(h)-PXR(h) mouse livers exhibited temporally and quantitatively similar transcriptional responses during 91 days of PB exposure including the sustained induction of the xenobiotic response gene Cyp2b10, the Wnt signaling inhibitor Wisp1, and noncoding RNA biomarkers from the Dlk1-Dio3 locus. Transient induction of DNA replication (Hells, Mcm6, and Esco2) and mitotic genes (Ccnb2, Cdc20, and Cdk1) and the proliferation-related nuclear antigen Mki67 were observed with peak expression occurring between 1 and 7 days PB exposure. All these transcriptional responses were absent in CAR(KO)-PXR(KO) mouse livers and largely reversible in wild-type and CAR(h)-PXR(h) mouse livers following 91 days of PB exposure and a subsequent 4-week recovery period. Furthermore, PB-mediated upregulation of the noncoding RNA Meg3, which has recently been associated with cellular pluripotency, exhibited a similar dose response and perivenous hepatocyte-specific localization in both wild-type and CAR(h)-PXR(h) mice. Thus, mouse livers coexpressing human CAR and PXR support both the xenobiotic metabolizing and the proliferative transcriptional responses following exposure to PB.