Dosing time dependent in vitro pharmacodynamics of Everolimus despite a defective circadian clock

Abstract

Everolimus (EV), a rapamycin analogue mTOR inhibitor, is used in the clinic to treat Estrogen positive (ER) breast cancer in order to avoid the resistance to hormonotherapy. Here, we investigated whether EV efficacy varied according to administration timing by using the ER breast cancer cell line MCF-7 as model system. Our results showed that instead of apoptosis, EV induced a G0/G1 phase blockage of MCF-7 cells. Following serum shock, MCF-7 cells displayed a statistically significant 24h rhythm of mammalian target of Rapamycin (mTOR) activity, but perturbed circadian clock genes oscillations. Interestingly, the different delivery schedule of EV presented different efficacy in G0/G1 phase blockage in serum shocked MCF-7 cells. Moreover, serum shock induced also a circadian-like oscillation in expression or activity of several important G1 phase progression proteins, such as Cyclin D1 and phosphorylated Retinoblastoma protein (RB). Inhibition mTOR activity by EV reduced Cyclin D1 and Cyclin D3 protein level as well as RB phosphorylation level. Taken together, the results indicated that serum shock synchronization induced a circadian oscillation in mTOR activity in MCF-7 cells, which rhythmically regulated the synthesis or phosphorylation of key G1 progression proteins, such as Cyclin D1 and phosphorylated RB, ultimately resulting in different G0/G1 blockage efficiency according to different EV administration timing.

The Cancer Clock Is (Not) Ticking: Links between Circadian Rhythms and Cancer

“…Daytime rather than nighttime administration of seliciclib, a cyclin-dependent kinase inhibitor, reduced Glasgow osteosarcoma tumor growth and improved circadian gene expression within the tumor [194]. mTOR activity has also been found to have a 24-h rhythmic pattern in cultured breast cancer cells and the efficacy of in vitro everolimus, an mTOR inhibitor, varied depending on the timing of the dose [195]. More research is needed into understanding how current drug protocols may be able to be optimized to target physiological and cellular circadian rhythms and improve cancer outcomes.…”

mTOR Signaling, Translational Control, and the Circadian Clock

“…In peripheral tissues, mTOR activities are rhythmic in the liver, cardiac and skeletal muscles, adipocytes, and retinal photoreceptors ( Huang et al, 2013 ; Jouffe et al, 2013 ; Shavlakadze et al, 2013 ; Khapre et al, 2014 ; Drägert et al, 2015a , b ; Lipton et al, 2015 , 2017 ; Chang et al, 2016 ). Interestingly, mTOR also shows circadian rhythms in human osteosarcomas, mouse renal carcinomas as well as human breast cancer cells ( Zhang et al, 2009 , 2018 ; Okazaki et al, 2014 ). These circadian mTOR studies are summarized in Table 1 .…”

Does rapamycin slow down time?

“…Conversely, circadian clock mediators affect the mTOR pathway and aging [ 31 ]. Because mTOR activity is itself part of the circadian clock, its sensitivity to rapamycin can vary widely depending on the time of the day and the phase of the clock [ 32 ]. This should be taken into account when comparing the numerous studies in mouse models.…”