ABSTRACT
BACKGROUND: Breast cancer is the most commonly diagnosed cancer among women and its metastases results in poor survival rates in patients. The ability to alter metabolism is a key attribute cancer cells use to survive within different metastatic microenvironments and cause organ failure. We hypothesized that evaluation of metabolic alterations within tumor cells could provide a better understanding of cancer metastasis. Therefore, to investigate underlying metabolic alterations during metastases, we utilized human MDA-MB-231 and mouse 4T1 models that closely mimic human breast cancer metastasis. METHODS: The glycolysis and glutamine pathway-related changes were examined in bone metastatic cells by XF-24 extracellular flux analyzer and western blotting. The expression levels of genes related to metabolism were examined by PCR arrays. RESULTS: The MDA-MB-231 cells isolated after bone metastases showed reduced glucose uptake and glycolysis compared to parental cells, suggesting that these cells could alter metabolic requirements for survival. To understand these metabolic changes, we investigated glutamine, a common and naturally occurring non-essential amino acid. Interestingly, in reduced glucose conditions both cell lines showed dependence on glutamine for cell survival, and with glutamine withdrawal significantly increasing apoptotic cell death. Glutamine was also critical for normal cell proliferation even in the presence of high glucose concentrations. To further understand this metabolic switch in metastatic cells, we examined the genes related to metabolism and identified a more than seven-fold downregulation of protein kinase C zeta (PKC-ζ) expression levels in bone-derived MDA-MB-231 cells compared to the parental population. The PKC-ζ levels were also significantly reduced in metastatic 4T1 cells compared to non-metastatic MT1A2 cells. Since PKC-ζ deficiency promotes glutamine utilization via the serine biosynthesis pathway, we examined glutamine metabolism. The ectopic expression of PKC-ζ inhibited glutamine conversion to glutamate, while mutant PKC-ζ reversed this effect. Furthermore, the gene expression levels of enzymes involved in serine biosynthesis, phosphoserine phosphatase (PSPH), phosphoserine aminotransferase (PSAT1), and phosphoglycerate dehydrogenase (PHGDH) showed upregulation following glucose deprivation with PKC-ζ deficiency. The PHGDH upregulation was inhibited by ectopically expressing wild type but not mutated PKC-ζ in glucose-deprived conditions. CONCLUSIONS: Our results support the upregulation of serine biosynthesis pathway genes and downregulation of PKC-ζ as potential metabolic alterations for bone metastatic breast cancer cells.
Subject(s)
Bone Neoplasms/metabolism , Bone Neoplasms/secondary , Breast Neoplasms/metabolism , Glutamine/metabolism , Protein Kinase C/genetics , Protein Kinase C/metabolism , Animals , Biosynthetic Pathways , Bone Neoplasms/genetics , Breast Neoplasms/genetics , Cell Line, Tumor , Cell Proliferation , Cell Survival , Down-Regulation , Female , Gene Expression Regulation, Neoplastic , Glycolysis , Humans , Mice , Models, Biological , Tumor MicroenvironmentABSTRACT
Breast cancer metastases cause significant patient mortality. During metastases, cancer cells use autophagy, a catabolic process to recycle nutrients via lysosomal degradation, to overcome nutritional stress for their survival. The Runt-related transcription factor, Runx2, promotes cell survival under metabolic stress, and regulates breast cancer progression and bone metastases. Here, we identify that Runx2 enhances autophagy in metastatic breast cancer cells. We defined Runx2 function in cellular autophagy by monitoring microtubule-associated protein light chain (LC3B-II) levels, an autophagy-specific marker. The electron and confocal microscopic analyses were utilized to identify alterations in autophagic vesicles. The Runx2 knockdown cells accumulate LC3B-II protein and autophagic vesicles due to reduced turnover. Interestingly, Runx2 promotes autophagy by enhancing trafficking of LC3B vesicles. Our mechanistic studies revealed that Runx2 promotes autophagy by increasing acetylation of α-tubulin sub-units of microtubules. Inhibiting autophagy decreased cell adhesion and survival of Runx2 knockdown cells. Furthermore, analysis of LC3B protein in clinical breast cancer specimens and tumor xenografts revealed significant association between high Runx2 and low LC3B protein levels. Our studies reveal a novel regulatory mechanism of autophagy via Runx2 and provide molecular insights into the role of autophagy in metastatic cancer cells.
Subject(s)
Autophagy , Bone Neoplasms/metabolism , Breast Neoplasms/metabolism , Cell Movement , Core Binding Factor Alpha 1 Subunit/metabolism , Acetylation , Animals , Antineoplastic Agents/pharmacology , Autophagosomes/metabolism , Autophagy/drug effects , Bone Neoplasms/drug therapy , Bone Neoplasms/genetics , Bone Neoplasms/secondary , Breast Neoplasms/drug therapy , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cell Adhesion , Cell Movement/drug effects , Chloroquine/pharmacology , Core Binding Factor Alpha 1 Subunit/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , MCF-7 Cells , Mice, Inbred NOD , Mice, SCID , Microtubule-Associated Proteins/genetics , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Protein Transport , RNA Interference , Signal Transduction , Transfection , Tubulin/metabolismABSTRACT
This paper reports a temperature-tunable conjugated polymer poly[3-(2-ethyl-isocyanato-octadecanyl)-thiophene] (TCP) laser working in superradiant (SR)-or amplified spontaneous emission (ASE)-mode. The absorption spectra indicated the aggregate (mostly dimer) formation upon increasing concentration and/or decreasing temperature. Amplified spontaneous emission (ASE) was observed at suitable concentration, temperature, and pump energy values. The efficiency of the ASE from the TCP polymer was improved by energy transfer from an oligomer 9,9,9',9',9â³,9â³-hexakis(octyl)-2,7',2',7â³-trifluorene (HOTF). Moreover, the ASE wavelength can be tuned between 550 and 610 nm by changing the temperature of the solution from 60 to 10 °C. To the best of our knowledge, this is the first report of a high-power, temperature-tunable, and conjugated polymer laser.
