Abstract 2579: Combination therapy with MEK inhibition is efficacious in intracranial triple negative breast cancer models
Authors: Van Swearingen Amanda E.D. , Siegel Marni B. , Sambade Maria J. , Sud Shivani , Miller Samantha M. , Silva Grace , Bash Ryan E. , Santos Charlene M. , Darr David B. , Golitz Brian , Parker Joel S. , Miller C. Ryan , Johnson Gary L. , Anders Carey K. 
Published: August 2015

Abstract
Introduction: Nearly half of metastatic triple negative breast cancer (TNBC) patients develop brain metastases (BM) and face a poor prognosis. The blood-brain barrier (BBB) prevents many treatments from reaching intracranial tumors, and there are no FDA-approved systemic therapies to treat TNBC BM. In this study, we evaluated the efficacy of BBB-permeable, clinically-available inhibitors of MEK and identified rational co-target pathways in preclinical models of intracranial (IC) TNBC.
Methods: In vitro IC50s, synergy, and siRNA screens (700 kinase genes) were conducted in 4 human-derived TNBC lines (SUM149, MDA-MB-468, MDA-MB-436, MDA-MB-231Br). We evaluated the efficacy of the MEK1/2 inhibitor AZD6244 (AZD), the pan-PI3K inhibitor BKM120 (BKM), and the PDGFR inhibitor Pazopanib (Pazo) in IC TNBC mouse models. Tumor burden was monitored via bioluminescence, and IC tumors were frozen for gene expression analyses using custom human 4×44K Agilent microarrays or of kinome activity profiles using multiplex kinase inhibitor beads and mass spectrometry. To determine drivers of AZD sensitivity, DNA copy number data (Broad CCLE) was analyzed using SWITCHplus to identify copy number alterations that differ between sensitive (n = 8) vs. resistant (n = 12) TNBC lines based on their IC50s (Sanger Cancerxgene).
Results: In vitro, SUM149 and 231Br TNBC cells exhibited lower (&lt;20 uM) AZD IC50s while 468 cells were resistant (IC50 &gt;40 uM). Several genes synthetically enhanced lethality in SUM149 and 231Br cells: PI3K genes and PDGFRα/β with AZD, and MAPK/MAP2K/MAP3K genes with BKM, suggesting MEK+PI3K and MEK+PDGFR inhibition as rational combinations. AZD plus BKM or Pazo were synergistic in vitro in sensitive cell lines.
In vivo, AZD reduced tumor burden and improved survival in the SUM149 (72 vs. 45 days in controls, p &lt; 0.005) and 231Br (37 vs. 30 days, p &lt; 0.02) models, with no benefit in the other two models. Single agent BKM or Pazo resulted in little to no improvement in any model. However, in AZD-sensitive models, combined AZD+BKM inhibition increased survival (SUM149: 87 vs 45 days, p &lt; 0.0001; MDA-MB-231Br: 52 vs 30 days, p &lt; 0.001) as did AZD+Pazo (SUM149: 88 vs. 35 days, p &lt; 0.0001).
Several DNA segments were significantly altered in sensitive vs. resistant TNBC cell lines. Notably, MEK-pathway genes were lost in the resistant lines. Ongoing work will complete characterization of therapies in all models in vitro and in vivo and will compare genetic, transcriptional, and kinome activity alterations.
Conclusions: TNBC models exhibit different innate sensitivities to the BBB-permeable MEK inhibitor AZD6244. In sensitive models, AZD improves survival and reduces intracranial tumor burden, and rational combined inhibition of PI3K or PDGFR further increases survival. Identification of predictive biomarkers will enable translation of our results to biomarker-driven clinical trials for patients with TNBC BM.
Citation Format: Amanda E.D. Van Swearingen, Marni B. Siegel, Maria J. Sambade, Shivani Sud, Samantha M. Miller, Grace Silva, Ryan E. Bash, Charlene M. Santos, David B. Darr, Brian Golitz, Joel S. Parker, C. Ryan Miller, Gary L. Johnson, Carey K. Anders. Combination therapy with MEK inhibition is efficacious in intracranial triple negative breast cancer models. [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 2579. doi:10.1158/1538-7445.AM2015-2579