Abstract
Anchorage-independent growth of cancer cells in vitro is correlated to metastasis formation in vivo. Metformin use is associated with decreased breast cancer incidence and currently evaluated in cancer clinical trials. The combined treatment with metformin and 2-deoxy-D-glucose (2DG) in vitro induces detachment of viable MDA-MB-231 breast cancer cells that retain their proliferation capacity. This might be important for cell detachment from primary tumors, but the metabolic changes involved are unknown. We performed LC/MS metabolic profiling on separated attached and detached MDA-MB-231 cells treated with metformin and/or 2DG. High 2DG and metformin plus 2DG altered the metabolic profile similarly to metformin, inferring that metabolic changes are necessary but not sufficient while the specific effects of 2DG are crucial for detachment. Detached cells had higher NADPH levels and lower fatty acids and glutamine levels compared to attached cells, supporting the role of AMPK activation and reductive carboxylation in supporting anchorage-independent survival. Surprisingly, the metabolic profile of detached cells was closer to untreated control cells than attached treated cells, suggesting detachment might help cells adapt to energy stress. Metformin treated cells had higher fatty and amino acid levels with lower purine nucleotide levels, which is relevant for understanding the anticancer mechanisms of metformin.
Highlights
Anchorage-independent growth of cancer cells in vitro is correlated to metastasis formation in vivo
The effects of metformin and 2DG treatment on cell proliferation and viability were determined in a separate experiment using direct cell counting for total cell number (Fig. 1b) and annexin/Propidium iodide (PI) staining for the percentage of dead and apoptotic cells in the samples (Fig. 1c)
We present metabolic profiling results of attached and detached MDA-MB-231 breast cancer cells treated either with 2DG, metformin or with both compounds
Summary
Anchorage-independent growth of cancer cells in vitro is correlated to metastasis formation in vivo. The combined treatment with metformin and 2-deoxy-D-glucose (2DG) in vitro induces detachment of viable MDA-MB-231 breast cancer cells that retain their proliferation capacity. This might be important for cell detachment from primary tumors, but the metabolic changes involved are unknown. We observed that treating breast cancer cells with metformin and physiologically relevant concentration of 2DG induced detachment of viable cells that retained their proliferation capacity following r eseeding[14] This effect might be relevant for metastasis formation, where cell detachment from the invasion front is the crucial step in the early s tages[15,16]. Identifying the metabolic differences of attached and detached populations could help us better understand the metabolic changes involved in avoiding anoikis and surviving in anchorage-independent state
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