To establish alternative targeted therapies against triple negative (TN) breast cancer, the energy metabolism and the sensitivity of cell growth, migration, and invasiveness toward metabolic, canonical, and NSAID inhibitors were analyzed in MDA-MB-231 and MDA-MB-468, two TN metastatic breast cancer cell lines, under both normoxia (21% O) and hypoxia (0.1% O). For comparative purposes, the analysis was also carried out in the less-metastatic breast MCF-7 cancer cells. Under normoxia, oxidative phosphorylation (OxPhos) was significantly higher (2-times) in MDA-MB-468 than in MDA-MB-231 and MCF-7, whereas their glycolytic fluxes and OxPhos and glycolytic protein contents were all similar. TN cancer cell lines mainly depended on OxPhos (62-75%), whereas MCF-7 cells equally depended on both pathways for ATP supply. Hypoxia for 24 h promoted a significant increase (>20 times) in the glycolytic transcriptional master factor HIF1-α in its target proteins GLUT-1, HKI and II, and LDH-A (2-4 times) as well as in the glycolytic flux (1.3-2 times) vs normoxia in MDA-MB-468, MDA-MB-231, and MCF-7. On the contrary, hypoxia decreased (15-60%) the contents of COXIV, 2OGDH, ND1, and ATP synthase as well as the OxPhos flux (50-75%), correlating with a high mitophagy level in the three cell lines. Under hypoxia, the three cancer cell lines mainly depended on glycolysis (70-80%). Anti-mitochondrial drugs (oligomycin, casiopeina II-gly, and methoxy-TEA) and celecoxib, at doses used to block OxPhos, significantly decreased TN cancer cell proliferation (IC = 2-20 μM), migration capacity (10-90%), and invasiveness (25-65%). The present data support the use of mitochondrially targeted inhibitors for the treatment of TN breast carcinoma.

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The accelerated growth of solid tumors leads to episodes of both hypoxia and hypoglycemia (HH) affecting their intermediary metabolism, signal transduction, and transcriptional activity. A previous study showed that normoxia (20% O ) plus 24 h hypoglycemia (2.5 mM glucose) increased glycolytic flux whereas oxidative phosphorylation (OxPhos) was unchanged versus normoglycemia in HeLa cells. However, the simultaneous effect of HH on energy metabolism has not been yet examined. Therefore, the effect of hypoxia (0.1-1% O ) plus hypoglycemia on the energy metabolism of HeLa cells was analyzed by evaluating protein content and activity, along with fluxes of both glycolysis and OxPhos. Under hypoxia, in which cell growth ceased and OxPhos enzyme activities, ΔΨm and flux were depressed, hypoglycemia did not stimulate glycolytic flux despite increasing H-RAS, p-AMPK, GLUT1, GLUT3, and HKI levels, and further decreasing mitochondrial enzyme content. The impaired mitochondrial function in HH cells correlated with mitophagy activation. The depressed OxPhos and unchanged glycolysis pattern was also observed in quiescent cells from mature multicellular tumor spheroids, suggesting that these inner cell layers are similarly subjected to HH. The principal ATP supplier was glycolysis for HH 2D monolayer and 3D quiescent spheroid cells. Accordingly, the glycolytic inhibitors iodoacetate and gossypol were more effective than mitochondrial inhibitors in decreasing HH-cancer cell viability. Under HH, stem cell-, angiogenic-, and EMT-biomarkers, as well as glycoprotein-P content and invasiveness, were also enhanced. These observations indicate that HH cancer cells develop an attenuated Warburg and pronounced EMT- and invasive-phenotype. J. Cell. Physiol. 232: 1346-1359, 2017. © 2016 Wiley Periodicals, Inc.

Cancer development, growth, and metastasis are highly regulated by several transcription regulators (TRs), namely transcription factors, oncogenes, tumor-suppressor genes, and protein kinases. Although TR roles in these events have been well characterized, their functions in regulating other important cancer cell processes, such as metabolism, have not been systematically examined. In this review, we describe, analyze, and strive to reconstruct the regulatory networks of several TRs acting in the energy metabolism pathways, glycolysis (and its main branching reactions), and oxidative phosphorylation of nonmetastatic and metastatic cancer cells. Moreover, we propose which possible gene targets might allow these TRs to facilitate the modulation of each energy metabolism pathway, depending on the tumor microenvironment.

This study shows that the non-steroidal anti-inflammatory drug (NSAID) celecoxib and its non-cyclooxygenase-2 (COX2) analogue dimethylcelecoxib (DMC) exert a potent inhibitory effect on the growth of human cervix HeLa multi-cellular tumor spheroids (MCTS) when added either at the beginning (“preventive protocol”; IC50 = 1 ± 0.3 nM for celecoxib and 10 ± 2 nM for DMC) or after spheroid formation (“curative protocol”; IC50 = 7.5 ± 2 µM for celecoxib and 32 ± 10 µM for DMC). These NSAID IC50 values were significantly lower than those attained in bidimensional HeLa cells (IC50 = 55 ± 9 µM celecoxib and 48 ± 2 µM DMC) and bidimensional non-cancer cell cultures (3T3 fibroblasts and MCF-10A mammary gland cells with IC50 from 69 to >100 µM, after 24 h). The copper-based drug casiopeina II-gly showed similar potency against HeLa MCTS. Synergism analysis showed that celecoxib, DMC, and casiopeinaII-gly at sub-IC50 doses increased the potency of cisplatin, paclitaxel, and doxorubicin to hinder HeLa cell proliferation through a significant abolishment of oxidative phosphorylation in bidimensional cultures, with no apparent effect on non-cancer cells (therapeutic index >3.6). Similar results were attained with bidimensional human cervix cancer SiHa and human glioblastoma U373 cell cultures. In HeLa MCTS, celecoxib, DMC and casiopeina II-gly increased cisplatin toxicity by 41–85%. These observations indicated that celecoxib and DMC used as adjuvant therapy in combination with canonical anti-cancer drugs may provide more effective alternatives for cancer treatment.