Deciphering the impact of lipid metabolism in breast cancer cells

Vazzana, Roberta (2021). Deciphering the impact of lipid metabolism in breast cancer cells. PhD thesis The Open University.



Breast cancer represents the most common cancer among women and, despite increased early detection and improved therapeutic approaches, it also remains among the deadliest cancer worldwide. Accumulating evidence has shed light on the correlation between oncogenic signalling and metabolic reprogramming in cancer cells, and ten years ago metabolic reprogramming has been included among the hallmarks of cancer. In breast cancer a de-regulated metabolism has been described both in patients previously untreated and in treatment-resistant populations. Relapses are believed to arise from residual cancer stem cells that survive the initial therapeutic treatment. Tumour relapse still represents the main cause of breast cancer related death. Therefore, a comprehensive understanding of cancer stem cell biology and metabolism is needed to eventually impair the formation of tumour relapses and obtain more effective therapies.

In my Ph.D. research activity, I first investigated the correlation between lipid metabolism and breast cancer stem cells. Focusing specifically on lipid droplets (LD), the lipid storage organelle of the cell, I investigated the role of LD in breast cancer stem cells and tumorigenesis. Mammosphere assays and gene expression profiling, revealed a strong correlation between lipid droplet number and stemness in a panel of 4 different breast cancer cell lines. Furthermore, in vivo studies revealed the role of lipid droplets as a marker of cell harbouring increased tumorigenic capacity.

The accumulation of lipid droplets in cancer cells is part of a more general metabolic reprogramming characterizing breast tumours. Indeed, from a metabolic perspective, breast cancer cells show high glycolytic rates, intense anabolic growth and an up-regulation of de-novo fatty acid synthesis. Historically, a competitive relationship between glucose and fatty acid usage at the level of tissue metabolism was reported, with the cells ultimately relying upon only one of the two energy sources. In order to better dissect the role of lipid metabolism in breast cancer onset at a molecular level, I sought to investigate the previously described, but never elucidated, biochemical interplay between lipid and glucose metabolism. With a particular focus on the glycolytic enzyme Pyruvate kinase M2 (PKM2), which has been correlated also with the promotion and maintenance of the stemness traits, I focused my attention on the role of lipids as direct possible regulators of glycolysis and glycolytic enzymes. Medium and long-chain fatty acids are involved both in energy metabolism and in signalling pathways. Here, I show a new possible level of regulation, where fatty acids could act as allosteric regulators of PKM2. Using fatty-acid pull-down and in vitro binding assays, I demonstrate that PKM2 is able to directly interact with fatty acids. Particularly, my results suggest that it is the glycolytically inactive, dimeric form of PKM2 that preferentially interacts with FAs. To better dissect the functional implications of this regulation, finally I decided to characterize structurally the PKM2_K422E dimer mutant. Crystallization studies revealed a different conformation of the protein, compared to the wild type enzyme, that could correlate with the fatty acid binding capacity observed in my in vitro assays. Moreover, the structural characterization of the complex formed by PKM2_K422E and oleic acid is still under investigation.

Although further investigations are required to deeply characterize the molecular mechanism underpinning the correlation between altered lipid metabolism, LDs accumulation and stemness traits, the possibility that targeting lipid metabolism and LDs formation could impact on tumorigenesis, opens important scenarios for the development of new therapeutic strategies. My findings pave the way for new possible mechanisms by which deregulated lipid metabolism can impact on cellular proliferation and cancer progression, via the regulation of one of the key metabolic players and glycolytic enzyme PKM2.

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