Abstract P5-03-02: Pre-clinical investigation of PP2A inhibitor LB-100 in overcoming and preventing lapatinib resistance in HER2-positive breast cancer
Authors: Conlon N , McDermott M , Browne B , Roche S , O'Neill F , Meiller J , Browne A , Eustace A , Collins DM , O'Donovan N , Crown J 
Published: May 2019

Abstract
Background: HER2-positive breast cancer (BC) accounts for approximately 15% of all BC. HER2-targeted therapies, such as trastuzumab and lapatinib, have significantly improved the outcome for these patients. However, HER2-targeted therapy resistance is a common clinical issue. We have previously shown that protein phosphatase 2A (PP2A) plays a role in mediating acquired lapatinib resistance in HER2-positive BC and that response to lapatinib is enhancedin vitroby the lab-grade PP2A inhibitor, okadaic acid. The aim of this study was to examine the in vitro and in vivo efficacy of LB-100, a PP2A inhibitor that has completed phase I clinical testing (NCT01837667), in models of HER2-positive BC with acquired resistance to lapatinib.
Methods: HER2-positiveSKBR3 and HCC1954 BC cell lines were treated with 250 nM or 1 μM lapatinib, respectively, for 6 months to generate lapatinib-resistant SKBR3-L and HCC1954-L cell lines. In vitro sensitivity to lapatinib and LB-100 was assessed by 2D acid phosphatase assay. Combination index (CI) values were generated to identify synergistic combinations. Propidium iodide staining was used to determine cell cycle arrest and apoptosis. In order to examine the in vivo efficacy of LB-100, HCC1954-L cells were implanted into the mammary fat pad of BALB/c nude mice and treated with vehicle, lapatinib, LB-100, or lapatinib plus LB-100. To examine the prevention of the development of lapatinib resistance, SKBR3 and HCC1954 cells were treated twice weekly with lapatinib, LB-100 or the combination and stained with crystal violet when confluent.
Results: SKBR3-L and HCC1954-L cells were resistant to lapatinib at clinically relevant concentrations (IC50values = 2.37 ± 0.58 μM and 1.67 ± 0.34 μM). This represents a 46- and 5.2-fold decrease in lapatinib sensitivity. LB-100 had a greater anti-proliferative effect in the lapatinib-resistant SKBR3-L and HCC1954-L cell lines compared to their respective parental cell lines (IC50values = 2.12 ± 0.2 μM v 5.38 ± 0.6 μM, and 2.31 ± 0.19 μM v 5.32 ± 0.82 μM, respectively). LB-100 overcame lapatinib resistance in both models, as lapatinib plus LB-100 was synergistic in both cell lines (CI values = 0.56 ± 0.13 and 0.68 ±0.26). LB-100 caused cell death through the induction of apoptosis in SKBR3- L (p = 0.019) and HCC1954-L (p = 0.046) and the addition of lapatinib to LB-100 increased apoptotic induction in HCC1954-L cells (p=0.046).Lapatinib plus LB-100 was well tolerated in vivo. The HCC1954-L cell line maintained resistance to lapatinib in vivo and the combination of lapatinib and LB-100 significantly reduced HCC1954-L tumour volume compared to all other treatment arms (p = 0.0006). Interestingly, in vitro short-term resistance assays showed that the addition of LB-100 to lapatinib could also block the emergence of lapatinib resistance in both parental SKBR3 and HCC1954 cell lines.
Conclusions: This study indicates that LB-100 has in vitro and in vivo efficacy against lapatinib-resistant HER2-positive BC cell line models and justifies further investigation into its potential to circumvent or prevent lapatinib resistance in HER2-positive BC.
Citation Format: Conlon N, McDermott M, Browne B, Roche S, O'Neill F, Meiller J, Browne A, Eustace A, Collins DM, O'Donovan N, Crown J. Pre-clinical investigation of PP2A inhibitor LB-100 in overcoming and preventing lapatinib resistance in HER2-positive 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 P5-03-02.