Abstract P2-06-15: Rational combination of Wee1 and BCL-2 inhibition in preclinical models of triple-negative breast cancer
Authors: Simmons DM , Tse TE , Dailey K , Hartman SJ , Bagsby S , Pitts TM , Tentler JJ , Diamond JR 
Published: May 2019

Background:

Triple-negative breast cancer (TNBC) is an aggressive subtype distinguished by its lack of expression of receptors for estrogen, progesterone, and normal human epidermal growth factor 2 expression. TNBC is difficult to treat and is associated with a high risk of recurrence and mortality. In order to effectively treat TNBC, alternative therapeutic targets need to be identified. Wee1 is a tyrosine kinase that phosphorylates CDC2 to pause the cell cycle at the G2 checkpoint as a means to delay mitosis while DNA damage undergoes repair. Inactivation of Wee1 via adavosertib (AZD1775, MK1775), a highly selective inhibitor of Wee1, allows CDC25 to dephosphorylate the CDC2/cyclin B complex resulting in premature initiation of mitosis and, ultimately, mitotic catastrophe and apoptosis. An unbiased screen of adovosertib in combination with other targeted compounds in TNBC patient-derived xenograft (PDX) models demonstrated that the combination of adavosertib and navitoclax, an inhibitor of anti-apoptotic BCL-2 and BCL-XL proteins, had greater efficacy than the single agents alone. The purpose of this study was to investigate the combination of adavosertib and navitoclax in preclinical TNBC models, both in vitro and in vivo.

Methods:

HCC1937, CAL51, MDA-MB-231 and MDA-MB-468 cells were plated in 96-well plates and exposed to increasing concentrations of adavosertib (125nM – 1000nM), navitoclax (2500nM – 10000nM), or the combination. Cellular proliferation was assessed in real-time using IncuCyte Live Cell Analysis, followed by endpoint sulforhodamine B (SRB) assay. Combination effects were analyzed using Calcusyn to determine combination indexes (CI). Apoptosis was assessed via the Caspase 3/7 assay. Western blotting was used to assess changes in expression of CDC2, phospho-CDC2, and BCL2. TNBC PDX models CU_TNBC_013 and CU_TNBC_014 were treated with vehicle, adavosertib (50mg/kg), navitoclax (100mg/kg), or the combination and assessed for tumor growth inhibition.

Results:

The combination of adavosertib and navitoclax resulted in greater antiproliferative effects in vitro compared to either single agent (p&lt; 0.05). This effect was classified as synergistic with CI values &lt;1. We observed a significant increase in apoptosis with the combination treatment as measured by Caspase 3/7 (p &lt;0.005). The combination of adavosertib and navitoclax treatment resulted in a decrease in phospo-CDC2, and BCL2 in cell lines. In vivo, the combination treatment resulted in greater tumor growth inhibition as compared to adavosertib or navitoclax alone in the CU_TNBC_013 and CU_TNBC_014 PDX models.

Conclusions:

The combination of adavosertib and navitoclax is active in preclinical TNBC models and induces apoptosis and tumor growth inhibition. This data supports the continued development of this combination in TNBC with investigation of potential selective markers.

Citation Format: Simmons DM, Tse TE, Dailey K, Hartman SJ, Bagsby S, Pitts TM, Tentler JJ, Diamond JR. Rational combination of Wee1 and BCL-2 inhibition in preclinical models of triple-negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-06-15.