Fatty acid synthase (FAS) is overexpressed in cancer cells, and expression of this enzyme increases fatty acid biosynthesis necessary for cell survival and proliferation. Therefore, it has been a popular therapeutic target for cancer drugs. However, there are several unexpected side effects when using FAS inhibitors alone, such as metabolic changes for cell survival and drug resistance. So to circumvent this phenomenon, a combination therapy that modulates multiple metabolism may be a better method to reduce the side effects.

We were able to confirm in the previous results that fatty acids and LysoPCs were increased when FAS inhibitor was treated. The kennedy pathway is a major pathway for phospholipid synthesis leading to LysoPC synthesis through the lands cycle. Inhibition of the Kennedy pathway affects cell survival by regulating the balance between phospholipids and neutral lipids. A study was conducted on maximizing the anticancer effect by inhibiting the kennedy pathway under the assumption that FAS inhibition would activate it.

In this study, for finding out where choline was obtained from, kennedy pathway-related metabolites were measured by UPLC-MS/MS using FAS inhibitor TVB-3166, orlistat and choline kinase α inhibitor EB-3D in choline-d₉ treated media. Furthermore, Cell viability experiments were performed to confirm the synergistic effect of the two inhibitors together compared to the single treatment.

In UPLC-MS/MS results, choline-d₉, CDP-choline-d₉, and LysoPC-d₉ were increased treat FAS inhibition indicating that the choline was uptaken from the extracellular matrix. When FAS and kennedy pathway were inhibited together using FAS inhibitor and EB-3D, there was a significant synergistic effect on cell viability. Taken together, our results show that inhibiting FAS simultaneously with the intervention of the Kennedy pathway maximizes the anticancer effect.