Using stable isotope and click-chemistry probes to study fatty acid flux in cancer cells

The free fatty acids in the cell are dynamically regulated and utilized for mitochondrial oxidation, membrane biogenesis and storage in lipid droplets. The fatty acid flux in cancer is dysregulated due to mutations in oncogenes, tumor suppressors and stress factors in the tumor microenvironment. The dysregulated fatty acid flux provides energy and lipids for tumor growth and progression. On the other hand, re-directing fatty acid flux to toxic lipid species could be employed to enhance the efficacy of cancer therapeutics. In this protocol article, we described two different approaches using stable isotope probes and click-chemistry probes to study fatty acid flux in cancer cells. Compared to the traditional tracing approaches using radioactive probes, these two methods are much more accessible. Although we optimized our protocols using pancreatic cancer cell cultures, we can envision the adaptation of these protocols for other in vitro cultures including non-cancer cells and organoid cultures.