PIK3CA hotspot mutations p. H1047R and p. H1047L sensitize breast cancer cells to thymoquinone treatment by regulating the PI3K/Akt1 pathway.

Abstract

Nigella sativa (N. sativa) exhibits anti-inflammatory, antioxidant, antidiabetic, antimetastatic and antinociceptive effects and has been used to treat dozens of diseases. Thymoquinone (TQ) is an important and active component isolated from N. sativa seeds. Inhibition of cancer-associated activating PIK3CA mutations is a new prospective targeted therapy in personalized metastatic breast cancer (MBC). TQ is reported to be an effective inhibitor of the PI3K/Akt1 pathway in MBC. This study aimed to evaluate the in vitro antitumor effect of TQ in the context of two PIK3CA hotspot mutations, p. H1047R and p. H1047L. Molecular dynamics, free energy landscapes and principal component analyses were also used to survey the mechanistic effects of the p. H1047R and p. H1047L mutations on the PI3K/Akt1 pathway. Our findings clearly confirmed that the p. H1047R and p. H1047L mutants could reduce the inhibitory effect of ΔNp63α on the kinase domain of PIK3CA, resulting in increased activity of PI3K downstream signals. Structurally, the partial disruption of the interaction between the ΔNp63α DNA binding domain and the PIK3CA kinase domain at residues 114-359 and 797-1068 destabilizes the conformation of the activation loop and modifies the PIK3CA/ΔNp63α complex. Alongside these structural changes, we found that TQ treatment resulted in high PI3K/Akt1 pathway inhibition in p. H1047R and p. H1047L-expressing cells versus wild-type cells. These two PIK3CA hotspot mutations therefore not only contribute to tumor progression in patients with MBC but may also serve as targets for the development of novel small molecule therapeutic strategies.