Abstract P1-19-25: Wee1 inhibition enhances the anti-tumor effects of capecitabine in preclinical models of triple negative breast cancer
Authors: Todd M Pitts, Dennis M Simmons, Kyrie Dailey, Stacey M Bagby, Sarah J Hartman, Betelehem W Yacob, Brian Gittleman, John J Tentler, Diana Cittely, D. Ryan Ormond, Wells A Messersmith, S Gail Eckhardt, Jennifer R Diamond
Journal: Cancer Research (2020)

Abstract Background: Triple-negative breast cancer (TNBC) is an aggressive subtype defined by lack of hormone receptor expression and non-amplified HER2. TNBC accounts for approximately 15% of breast cancer cases, however, is associated with an increased risk of cancer recurrence, brain metastasis, and death due to metastatic breast cancer. Mutations in p53 are common in TNBC, occurring in approximately 85% of tumors. While a number of promising targeted therapies are on the horizon in TNBC including immunotherapy, there remains an unmet need for active targeted therapies where chemotherapy remains the standard treatment for metastatic disease and results in a median survival of 12-18 months. 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. AZD1775 potentiates the activity of DNA-damaging and antimetabolite chemotherapeutics in preclinical models without TP53-deficiency, possibly due to baseline replicative stress or compromised DNA repair proficiency. A previous unbiased screen of CTEP compounds in TNBC PDX models demonstrated that the combination of adavosertib and capecitabine/5FU had greater anti-proliferative effects than either of the single agents. The purpose of this study was to further investigate the combination of adavosertib and capecitabine/5FU in preclinical TNBC models. Methods: HCC1937, CAL51, MDA-MB-231 and MDA-MB-468 cells were plated in 96-well plates and exposed to increasing concentrations of adavosertib, 5FU, or the combination. 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 CDC2, phospho-CDC2, H2AX, and Bcl-xL. TNBC PDX models CU_TNBC_012 and CU_TNBC_013 were treated with vehicle, adavosertib, 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&amp;lt;0.05 combo vs adavosertib or capecitabine, TNBC013, p&amp;lt;0.01 combo vs adavosertib or capecitabine ). An enhanced antiproliferative effect was observed in the adavosertib/5FU 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. Citation Format: Todd M Pitts, Dennis M Simmons, Kyrie Dailey, Stacey M Bagby, Sarah J Hartman, Betelehem W Yacob, Brian Gittleman, John J Tentler, Diana Cittely, D. Ryan Ormond, Wells A Messersmith, S Gail Eckhardt, Jennifer R Diamond. Wee1 inhibition enhances the anti-tumor effects of capecitabine in preclinical models of triple negative breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P1-19-25.