In Vivo E2F Reporting Reveals Efficacious Schedules of MEK1/2–CDK4/6 Targeting and mTOR–S6 Resistance Mechanisms

Abstract

Targeting cyclin-dependent kinases 4/6 (CDK4/6) represents a therapeutic option in combination with BRAF inhibitor and/or MEK inhibitor (MEKi) in melanoma; however, continuous dosing elicits toxicities in patients. Using quantitative and temporal reporting, we show that continuous MEKi with intermittent CDK4/6 inhibitor (CDK4/6i) led to more complete tumor responses versus other combination schedules. Nevertheless, some tumors acquired resistance that was associated with enhanced phosphorylation of ribosomal S6 protein. These data were supported by phospho-S6 staining of melanoma biopsies from patients treated with CDK4/6i plus targeted inhibitors. Enhanced phospho-S6 in resistant tumors provided a therapeutic window for the mTORC1/2 inhibitor AZD2014. Mechanistically, upregulation or mutation of was associated with resistance in models and patient samples, respectively, and mutant NRAS was sufficient to enhance resistance. This study utilizes an reporter model to optimize schedules and supports targeting mTORC1/2 to overcome MEKi plus CDK4/6i resistance. Mutant BRAF and NRAS melanomas acquire resistance to combined MEK and CDK4/6 inhibition via upregulation of mTOR pathway signaling. This resistance mechanism provides the preclinical basis to utilize mTORC1/2 inhibitors to improve MEKi plus CDK4/6i drug regimens. .

A Functional Landscape of Resistance to MEK1/2 and CDK4/6 Inhibition in NRAS-Mutant Melanoma

“…S1G and S1H) and an increase in CCND1 ( Supplementary Fig. S1H) total protein levels, both of which are consistent with previous studies (7,11). Palbociclib inhibition did not alter the MAPK signaling components RAF and MEK, however we did observe an increase in phospho-ERK levels (Fig.…”

“…4H). These data suggest that reengagement of mTOR signaling is associated with resistance to trametinib and palbociclib treatment in NRAS mutant melanoma, which is consistent with known in vivo and clinical mechanisms of inhibitor resistance (7,11).…”

Overcoming CDK4/6 inhibitor resistance in ER-positive breast cancer

“…While no changes in mTOR signalling components have been documented in long-term resistance models, CDK4/6 inhibitor-resistant cell lines have demonstrated sensitivity to mTORC1/2 inhibition (Michaloglou et al 2018). Other signalling changes include NRAS amplification or mutation in an NRAS model of melanoma co-treated with MEK1 inhibition and CDK4/6 inhibition (Teh et al 2018), and alterations in FGF/FGFR signalling in resistance to CDK4/6 inhibitors both alone and in combination with endocrine therapy (Cruz et al 2018, Mao et al 2018, Shee et al 2018. Finally, an activating mutation in PIK3CA E545K , was shown to display a resistance phenotype to combination MEK and CDK4/6 inhibitors, and the outgrowth of a clone expressing this mutation caused recurrence in a melanoma patient treated with these inhibitors (Romano et al 2018).…”

Differential expression of phospho‐S6 in hair follicle tumors: Evidence of mammalian target of rapamycin pathway activation

“…Phospho‐S6 is a reliable surrogate biomarker/readout of mTOR axis activation and has been investigated in numerous studies focusing on neoplastic and non‐neoplastic conditions . In a recent study, phospho‐S6 was shown to be at least partially responsible for resistance to combinational mitogen‐activated protein kinase MEKi therapy with CDK4/6 inhibitors in melanoma . To date, there are no studies investigating mTOR pathway activation in hair follicle tumors through protein expression on formalin fixed tissue.…”