Drastically elevated glycolytic activity is a prominent metabolic feature of cancer cells. Until recently it was thought that tumor cells shift their entire energy production from oxidative phosphorylation (OXPHOS) to glycolysis. However, new evidence indicates that many cancer cells still have functional OXPHOS, despite their increased reliance on glycolysis. Growing pre-clinical and clinical evidence suggests that targeting mitochondrial metabolism has anti-cancer effects. Here, we analyzed mitochondrial respiration and the amount and activity of OXPHOS complexes in four melanoma cell lines and normal human dermal fibroblasts (HDFs) by Seahorse real-time cell metabolic analysis, immunoblotting, and spectrophotometry. We also tested three clinically approved antibiotics, one anti-parasitic drug (pyrvinium pamoate), and a novel anti-cancer agent (ONC212) for effects on mitochondrial respiration and proliferation of melanoma cells and HDFs. We found that three of the four melanoma cell lines have elevated glycolysis as well as OXPHOS, but contain dysfunctional mitochondria. The antibiotics produced different effects on the melanoma cells and HDFs. The anti-parasitic drug strongly inhibited respiration and proliferation of both the melanoma cells and HDFs. ONC212 reduced respiration in melanoma cells and HDFs, and inhibited the proliferation of melanoma cells. Our findings highlight ONC212 as a promising drug for targeting mitochondrial respiration in cancer.
Melanomas are genetically and metabolically heterogeneous, which influences therapeutic efficacy and contributes to the development of treatment resistance in patients with metastatic disease. Metabolite phenotyping helps to better understand complex metabolic diseases, such as melanoma, and facilitates the development of novel therapies. Our aim was to characterize the tumor and plasma metabolomes of mice bearing genetically different melanoma xenografts. We engrafted the human melanoma cell lines A375 (BRAF mutant), WM47 (BRAF mutant), WM3000 (NRAS mutant), and WM3311 (BRAF, NRAS, NF1 triple-wildtype) and performed a broad-spectrum targeted metabolomics analysis of tumor and plasma samples obtained from melanoma-bearing mice as well as plasma samples from healthy control mice. Differences in ceramide and phosphatidylcholine species were observed between melanoma subtypes irrespective of the genetic driver mutation. Furthermore, beta-alanine metabolism differed between melanoma subtypes and was significantly enriched in plasma from melanoma-bearing mice compared to healthy mice. Moreover, we identified beta-alanine, p-cresol sulfate, sarcosine, tiglylcarnitine, two dihexosylceramides, and one phosphatidylcholine as potential melanoma biomarkers in plasma. The present data reflect the metabolic heterogeneity of melanomas but also suggest a diagnostic biomarker signature for melanoma screening.
Targeted analysis
of Coffea arabica and Coffea canephora green coffees (total sample size n = 57) confirmed
2-O-β-d-glucopyranosyl-carboxyatractyligenin
(6) as the quantitatively
dominating carboxyatractyligenin derivative. Its abundance in Arabicas
(2425 ± 549 nmol/g, n = 48) exceeded that in
Robustas (34 ± 12 nmol/g, n = 9) roughly by
a factor of 70. Coffee processing involving heat (e.g., steam treatment
and decaffeination) reduced concentrations of 6 and increased
those of the decarboxylated derivative. The bioavailability of compound 6 in Caenorhabditis elegans was demonstrated
by ultraperformance liquid chromatography-tandem mass spectrometry
analysis of extracts prepared from nematode cultures incubated in
a liquid medium containing 6. A toxicity assay performed
to assess the impact of 6 in vivo showed a 20-fold higher
median lethal dose (LD50 = 11.7 ± 1.2 mM) concentration
compared to that of the known phytotoxic adenine-nucleotide transporters
inhibitor carboxyatractyloside (2, LD50 =
0.61 ± 0.05 mM), whereas 1 mM 6 and 0.1 mM 2 were sufficient to decrease the survival of wild type C. elegans, already 10–20-fold lower doses reduced
reproduction. Because the insulin/insulin-like growth factors signaling
cascade (IIS) is a key regulator of life span and stress resistance,
the impact of compound 6 on the survival of long-living daf-2 C. elegans was tested. As the susceptibility of these
nematodes to 6 was as high as that in wild type, an impact
on central metabolic processes independent of IIS was suggested. Analysis
of the in vivo adenosine triphosphate (ATP) content of adult C. elegans revealed no changes after 1 and 24 h, but a 50%
reduction after treatment with 1 mM 6 during the entire
postembryonic development. These data speak for a developmental-stage-dependent
modulation of the ATP pool by 6.
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