MATHEMATICAL MODELING REVEALS THAT G2/M CHECKPOINT OVERRIDE CREATES A THERAPEUTIC VULNERABILITY IN TSC2-NULL TUMORS

Cell cycle checkpoints determine whether cells meet requirements to progress through the next stage. In response to DNA damage, how cells activate checkpoints have been well studied, but little is known about checkpoint deactivation (recovery), which directly impacts on cell fate. In tumor cells, the signaling network has been rewired due to epigenetic and genetic alterations, which result in resistance to the cell cycle control, and thus resistance to chemotherapy or radiation therapy. Therefore, it is critical to identify molecules required for checkpoint recovery or adaptation after DNA damage. To achieve this goal, we performed a multidisciplinary study combining reverse phase protein array (RPPA) data, molecular biology and mathematical modeling to systematically identify molecules required for DNA damage checkpoint recovery. The mTOR complex 1 (mTORC1) plays an essential role to regulate mitotic entry after irradiation. Inhibition of the mTOR pathway delayed G2/M checkpoint recovery, while TSC2-null cells with hyperactivity of mTORC1 exhibited the opposite results. Furthermore, our mechanistic study revealed that mTOR signaling pathway controls a transcriptional program of mitotic entry through regulating histone lysine demethylase KDM4B, which is required for the epigenetic regulation of key mitosis-related genes including CCNB1 and PLK1. Given accelerated G2/M checkpoint recovery in TSC2-null cells with mTORC1 hyperactivity, we postulated that further abrogation of the G2/M checkpoint may facilitate mitotic catastrophe and selectively kill cells. As we expected, TSC2-null cells were more sensitive to the WEE1 inhibitor, a negative regulator of mitotic entry, compared to wild-type cells. In summary, we reported a novel mechanism of the mTORC1 signaling in regulating a transcriptional program required for G2/M checkpoint recovery after DNA damage. This mechanism provided a therapeutic strategy for TSC patients with mTORC1 hyperactivity using the WEE1 inhibitor, which has a potential to be translated into clinical trials