Investigating mechanisms of acquired resistance to AT13148, an AGC kinase inhibitor

The PI3K/AKT/mTOR (PAM) pathway is a major driver of cell growth, proliferation and survival, and is frequently dysregulated in cancer. AKT, a member of the AGC family of serine/threonine kinases, is a key signalling node within the PAM pathway, and as such, an attractive therapeutic target for the treatment of cancer. AT13148 is an ATP-competitive inhibitor of AKT, currently in phase 1 clinical trial, which also potently inhibits several other clinically relevant AGC kinases, such as ROCK1/2 and p70S6K. Acquired resistance to kinase inhibitors has been a barrier to their success, but resistance to AT13148 is yet to be defined. Therefore, the aim of this thesis was to investigate mechanisms of acquired AT13148 resistance, using preclinical cell line models.

Isogenic AT13148 resistant sub-clones were generated from the A2780 human ovarian carcinoma cell line, which harbours mutations in the PAM pathway. In these sub-clones, phosphorylation of S6RP (p70S6K substrate) and PRAS40 (AKT substrate) was refractory to AT13148 treatment, concurrent with increased ERK 1/2 phosphorylation, when compared to the parental A2780 cell line. Two of the resistant sub-clones were sensitised to AT13148 on exposure to the ERK inhibitor GDC-0994, with the combination of AT13148 and GDC-0994 shown to restore the inhibition of S6RP phosphorylation. This implicates ERK 1/2 as a driver of AT13148 resistance, potentially via the reactivation of the PAM pathway and suggests ERK inhibition as a strategy to overcome this resistance. A loss of DUSP6 expression, an ERK 1/2 phosphatase, was subsequently detected in AT13148 resistant sub-clones, but DUSP6 loss alone was not shown to cause a sustained increase in ERK 1/2 phosphorylation or confer AT13148 resistance, suggesting other factors are required for AT13148 resistance.

In conclusion, the results presented in this thesis identify ERK 1/2 as a driver of AT13148 resistance. This discovery has the potential to aid the ongoing clinical development of AT13148 and provide a therapeutic strategy to overcome AT13148 resistance.