Mitochondrial ATP generation is more proteome efficient than glycolysis.

Metabolic efficiency profoundly influences organismal fitness. Nonphotosynthetic organisms, from yeast to mammals, derive usable energy primarily through glycolysis and respiration. Although respiration is more energy efficient, some cells favor glycolysis even when oxygen is available (aerobic glycolysis, Warburg effect). A leading explanation is that glycolysis is more efficient in terms of ATP production per unit mass of protein (that is, faster). Through quantitative flux analysis and proteomics, we find, however, that mitochondrial respiration is actually more proteome efficient than aerobic glycolysis. This is shown across yeast strains, T cells, cancer cells, and tissues and tumors in vivo. Instead of aerobic glycolysis being valuable for fast ATP production, it correlates with high glycolytic protein expression, which promotes hypoxic growth. Aerobic glycolytic yeasts do not excel at aerobic growth but outgrow respiratory cells during oxygen limitation. We accordingly propose that aerobic glycolysis emerges from cells maintaining a proteome conducive to both aerobic and hypoxic growth.

Methionine synthase supports tumour tetrahydrofolate pools.

Mammalian cells require activated folates to generate nucleotides for growth and division. The most abundant circulating folate species is 5-methyl tetrahydrofolate (5-methyl-THF), which is used to synthesize methionine from homocysteine via the cobalamin-dependent enzyme methionine synthase (MTR). Cobalamin deficiency traps folates as 5-methyl-THF. Here, we show using isotope tracing that MTR is only a minor source of methionine in cell culture, tissues or xenografted tumours. Instead, MTR is required for cells to avoid folate trapping and assimilate 5-methyl-THF into other folate species. Under conditions of physiological extracellular folates, genetic MTR knockout in tumour cells leads to folate trapping, purine synthesis stalling, nucleotide depletion and impaired growth in cell culture and as xenografts. These defects are rescued by free folate but not one-carbon unit supplementation. Thus, MTR plays a crucial role in liberating THF for use in one-carbon metabolism.

IRS2 copy number gain, KRAS and BRAF mutation status as predictive biomarkers for response to the IGF-1R/IR inhibitor BMS-754807 in colorectal cancer cell lines.

Insulin-like growth factor receptor 1 (IGF-1R)-targeting therapies are currently at an important crossroad given the low clinical response rates seen in unselected patients. Predictive biomarkers for patient selection are critical for improving clinical benefit. Coupling in vitro sensitivity testing of BMS-754807, a dual IGF-1R/IR inhibitor, with genomic interrogations in 60 human colorectal cancer cell lines, we identified biomarkers correlated with response to BMS-754807. The results showed that cell lines with BRAF(V600E) or KRAS(G13D) mutation were resistant, whereas cell lines with wild-type of both KRAS and BRAF were particularly sensitive to BMS-754807 if they have either higher RNA expression levels of IR-A or lower levels of IGFBP6. In addition, the cell lines with KRAS mutations, those with either insulin receptor substrate 2 (IRS2) copy number gain (CNG) or higher IGF-1R expression levels, were more sensitive to the drug. Furthermore, cell lines with IRS2 CNG had higher levels of ligand-stimulated activation of IGF-1R and AKT, suggesting that these cell lines with IGF-IR signaling pathways more actively coupled to AKT signaling are more responsive to IGF-1R/IR inhibition. IRS2 siRNA knockdown reduced IRS2 protein expression levels and decreased sensitivity to BMS-754807, providing evidence for the functional involvement of IRS2 in mediating the drug response. The prevalence of IRS2 CNG in colorectal cancer tumors as measured by qPCR-CNV is approximately 35%. In summary, we identified IRS2 CNG, IGF-1R, IR-A, and IGFBP6 RNA expression levels, and KRAS and BRAF mutational status as candidate predictive biomarkers for response to BMS-754807. This work proposed clinical development opportunities for BMS-754807 in colorectal cancer with patient selection to improve clinical benefit.

Cancer cell dependence on unsaturated fatty acids implicates stearoyl-CoA desaturase as a target for cancer therapy.

Emerging literature suggests that metabolic pathways play an important role in the maintenance and progression of human cancers. In particular, recent studies have implicated lipid biosynthesis and desaturation as a requirement for tumor cell survival. In the studies reported here, we aimed to understand whether tumor cells require the activity of either human isoform of stearoyl-CoA-desaturase (SCD1 or SCD5) for survival. Inhibition of SCD1 by siRNA or a small molecule antagonist results in strong induction of apoptosis and growth inhibition, when tumor cells are cultured in reduced (2%) serum conditions, but has little impact on cells cultured in 10% serum. Depletion of SCD5 had minimal effects on cell growth or apoptosis. Consistent with the observed dependence on SCD1, but not SCD5, levels of SCD1 protein increased in response to decreasing serum levels. Both induction of SCD1 protein and sensitivity to growth inhibition by SCD1 inhibition could be reversed by supplementing growth media with unsaturated fatty acids, the product of the enzymatic reaction catalyzed by SCD1. Transcription profiling of cells treated with an SCD inhibitor revealed strong induction of markers of endoplasmic reticulum stress. Underscoring its importance in cancer, SCD1 protein was found to be highly expressed in a large percentage of human cancer specimens. SCD inhibition resulted in tumor growth delay in a human gastric cancer xenograft model. Altogether, these results suggest that desaturated fatty acids are required for tumor cell survival and that SCD may represent a viable target for the development of novel agents for cancer therapy.

Exome sequencing reveals comprehensive genomic alterations across eight cancer cell lines.

It is well established that genomic alterations play an essential role in oncogenesis, disease progression, and response of tumors to therapeutic intervention. The advances of next-generation sequencing technologies (NGS) provide unprecedented capabilities to scan genomes for changes such as mutations, deletions, and alterations of chromosomal copy number. However, the cost of full-genome sequencing still prevents the routine application of NGS in many areas. Capturing and sequencing the coding exons of genes (the "exome") can be a cost-effective approach for identifying changes that result in alteration of protein sequences. We applied an exome-sequencing technology (Roche Nimblegen capture paired with 454 sequencing) to identify sequence variation and mutations in eight commonly used cancer cell lines from a variety of tissue origins (A2780, A549, Colo205, GTL16, NCI-H661, MDA-MB468, PC3, and RD). We showed that this technology can accurately identify sequence variation, providing ∼95% concordance with Affymetrix SNP Array 6.0 performed on the same cell lines. Furthermore, we detected 19 of the 21 mutations reported in Sanger COSMIC database for these cell lines. We identified an average of 2,779 potential novel sequence variations/mutations per cell line, of which 1,904 were non-synonymous. Many non-synonymous changes were identified in kinases and known cancer-related genes. In addition we confirmed that the read-depth of exome sequence data can be used to estimate high-level gene amplifications and identify homologous deletions. In summary, we demonstrate that exome sequencing can be a reliable and cost-effective way for identifying alterations in cancer genomes, and we have generated a comprehensive catalogue of genomic alterations in coding regions of eight cancer cell lines. These findings could provide important insights into cancer pathways and mechanisms of resistance to anti-cancer therapies.

Synergistic antitumor activity of ixabepilone (BMS-247550) plus bevacizumab in multiple in vivo tumor models.

PURPOSE: Angiogenesis is a critical step in the establishment, growth, and metastasis of solid tumors, and combination of antiangiogenic agents with chemotherapy is an attractive therapeutic option. We investigated the potential of ixabepilone, the first in a new class of antineoplastic agents known as epothilones, to synergize with antiangiogenic agents to inhibit tumor growth. EXPERIMENTAL DESIGN: In vitro and in vivo cytotoxicity of ixabepilone as single agent and in combination with two targeted antiangiogenic agents, bevacizumab or sunitinib, were examined in preclinical tumor models. Direct effects of the agents against endothelial cells was also examined and compared with the effects of paclitaxel as single agent and in combination with bevacizumab. RESULTS: Ixabepilone showed robust synergistic antitumor activity in combination with bevacizumab and sunitinib in preclinical in vivo models derived from breast, colon, lung, and kidney cancers. The synergistic antitumor effect was greater with ixabepilone compared with paclitaxel. Furthermore, ixabepilone was more effective than paclitaxel at killing endothelial cells expressing P-glycoprotein in vitro and inhibiting endothelial cell proliferation and tumor angiogenesis in vivo. CONCLUSIONS: Ixabepilone may enhance the antitumor effects of antiangiogenic therapy by direct cytotoxicity and also indirectly via the killing of tumor-associated endothelial cells. Given that ixabepilone has reduced susceptibility to drug efflux pumps compared with taxanes, these data may explain the increased antiangiogenic and antitumor activity of ixabepilone in combination with antiangiogenic agents. Phase II studies to assess the efficacy and safety of ixabepilone plus bevacizumab in locally recurrent or metastatic breast cancer are planned.