Abstract 4719: Targeting interdependent signaling pathways to increase the durability and magnitude of response: promising combination therapy with dual mTORC1/2 inhibitors and CDK4/6 inhibitors
Authors: Crafter Claire , Curwen Jon , Delpuech Oona , Oplustilova Lenka , Green Stephen , Brown Henry , Trigwell Cath , Cosulich Sabina 
Published: August 2015

Abstract
Two emerging mechanisms of endocrine resistance in estrogen receptor positive (ER+) breast cancers include the activation of the phosphatidylinositol 3-kinase (PI3K) / mammalian target of rapamycin (mTOR) pathway and the de-coupling of cell cycle control from ER signaling via de-regulation of the cyclin D/cyclin dependent kinase (CDK4/6) pathway. In this study, we hypothesized that combining inhibitors of both pathways using the dual mTORC1 and mTORC2 inhibitor AZD2014 and the CDK4/6 inhibitor palbociclib would elicit an improved tumor response over agents that inhibit either pathway alone. Moreover, we hypothesized that combined inhibition of CDK4/6 and TORC1/2 together with inhibition of ER signaling, would cause a profound anti-tumor effect in breast cancer models.
In breast cancer cell lines, the combination of AZD2014 and palbociclib caused a synergistic inhibitory effect on cell growth. These effects occurred under conditions where addition of 300nM AZD2014 resulted in significant blockade on both TORC1 and TORC2 downstream effectors (&gt;80% inhibition of p-AKT, p-S6 and p-4EBP1) as well as significant down-regulation of cyclin D1 levels (&gt;70%). Similarly, inhibition of CDK4/6 using palbociclib (300nM) caused &gt;80% inhibition of p-RB and subsequent cell cycle blockade.The effects observed in breast cancer cell lines were recapitulated in vivo using the MCF7 xenograft model, where tumor regressions (&gt;105%) were observed with the combination. Furthermore, combining AZD2014, palbociclib and fulvestrant in this model also caused tumor regressions.To assess bone marrow tolerability of this combination, we investigated the response of human bone marrow multipotent progenitors (CD34+) in vitro. Palbociclib caused transient cell cycle G1 arrest. The combination of palbociclib with AZD2014 delayed entry into cell cycle, but did not have significant impact on the cell viability (&lt;15%).
We hypothesized that inhibition of CDK4/6 in combination with AZD2014 would also significantly affect pathway outputs and the transcriptional events downstream of the transcription factors E2F (e.g. CENPE, CCNA2, CDC6 and E2F1) and ER (e.g. PR). We therefore measured specific gene signatures downstream of these receptors in breast cancer cell lines. The ability to dose AZD2014 intermittently compared with rapalogues, together with its ability to block signaling from both TORC1 and 2, make this compound an ideal candidate for combining with CDK4/6 inhibitors such as palbociclib. Furthermore, addition of anti-hormonal therapies such as fulvestrant to this combination may provide additional benefit to breast cancer patients.
Citation Format: Claire Crafter, Jon Curwen, Oona Delpuech, Lenka Oplustilova, Stephen Green, Henry Brown, Cath Trigwell, Sabina Cosulich. Targeting interdependent signaling pathways to increase the durability and magnitude of response: promising combination therapy with dual mTORC1/2 inhibitors and CDK4/6 inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4719. doi:10.1158/1538-7445.AM2015-4719