The bromodomain inhibitor OTX015 (MK-8628) exerts anti-tumor activity in triple-negative breast cancer models as single agent and in combination with everolimus

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive subtype defined by lack of hormone receptor expression and non-amplified HER2. Adavosertib (AZD1775) is a potent, small molecule, ATP-competitive inhibitor of the Wee1 kinase that potentiates the activity of many DNA-damaging chemotherapeutics and is currently in clinical development for multiple indications. The purpose of this study was to investigate the combination of AZD1775 and capecitabine/5-FU in preclinical TNBC models. Methods: TNBC cell lines were treated with AZD1775 and 5-FU and cellular proliferation was assessed in real-time using IncuCyte® Live Cell Analysis. Apoptosis was assessed via the Caspase-Glo 3/7 assay system. Western blotting was used to assess changes in expression of downstream effectors. TNBC PDX models were treated with AZD1775, capecitabine, or the combination and assessed for tumor growth inhibition. Results: From the initial PDX screen, two of the four TNBC PDX models demonstrated a better response in the combination treatment than either of the single agents. As confirmation, two PDX models were expanded for statistical comparison . Both PDX models demonstrated a significant growth inhibition in the combination versus either of the single agents. (TNBC012, p<0.05 combo vs adavosertib or capecitabine, TNBC013, p<0.01 combo vs adavosertib or capecitabine ). An enhanced antiproliferative effect was observed in the adavosertib/5-FU combination treatment as measured by live cell analysis. An increase in apoptosis was observed in two of the four cell lines in the combination when compared to single agent treatment. Treatment with single agent adavosertib resulted in an increase in p-CDC2 in a dose dependent manner that was also observed in the combination treatment. Similar results were observed with γH2AX in two of the four cell lines tested. No significant changes were observed in Bcl-xL following treatment in any of the cell lines. Conclusions: The combination of adavosertib and capecitabine/5-FU demonstrated enhanced combination effects both in vitro and in vivo in preclinical models of TNBC. These results support the clinical investigation of this combination in patients with TNBC, including those with brain metastasis given the CNS penetration of both agents.

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Dasatinib Treatment Increases Sensitivity to c-Met Inhibition in Triple-Negative Breast Cancer Cells

Cancers ◽

10.3390/cancers11040548 ◽

2019 ◽

Vol 11 (4) ◽

pp. 548 ◽

Cited By ~ 4

Author(s):

Patricia Gaule ◽

Nupur Mukherjee ◽

Brendan Corkery ◽

Alex Eustace ◽

Kathy Gately ◽

...

Keyword(s):

Breast Cancer ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Kinase Inhibitor ◽

Combined Treatment ◽

Single Agent ◽

Src Kinase ◽

Ic50 Value ◽

Met Inhibitor ◽

Dasatinib Treatment

In pre-clinical studies, triple-negative breast cancer (TNBC) cells have demonstrated sensitivity to the multi-targeted kinase inhibitor dasatinib; however, clinical trials with single-agent dasatinib showed limited efficacy in unselected populations of breast cancer, including TNBC. To study potential mechanisms of resistance to dasatinib in TNBC, we established a cell line model of acquired dasatinib resistance (231-DasB). Following an approximately three-month exposure to incrementally increasing concentrations of dasatinib (200 nM to 500 nM) dasatinib, 231-DasB cells were resistant to the agent with a dasatinib IC50 value greater than 5 μM compared to 0.04 ± 0.001 µM in the parental MDA-MB-231 cells. 231-DasB cells also showed resistance (2.2-fold) to the Src kinase inhibitor PD180970. Treatment of 231-DasB cells with dasatinib did not inhibit phosphorylation of Src kinase. The 231-DasB cells also had significantly increased levels of p-Met compared to the parental MDA-MB-231 cells, as measured by luminex, and resistant cells demonstrated a significant increase in sensitivity to the c-Met inhibitor, CpdA, with an IC50 value of 1.4 ± 0.5 µM compared to an IC50 of 6.8 ± 0.2 µM in the parental MDA-MB-231 cells. Treatment with CpdA decreased p-Met and p-Src in both 231-DasB and MDA-MB-231 cells. Combined treatment with dasatinib and CpdA significantly inhibited the growth of MDA-MB-231 parental cells and prevented the emergence of dasatinib resistance. If these in vitro findings can be extrapolated to human cancer treatment, combined treatment with dasatinib and a c-Met inhibitor may block the development of acquired resistance and improve response rates to dasatinib treatment in TNBC.

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