Abstract 3333: Pre-clinical investigation of neratinib plus dasatinib in breast cancer
Authors: Conlon Neil T. , Roche Sandra , O'Neill Fiona , Meiller Justine , Crown John , Collins Denis M. 
Published: June 2022

Background: HER2-targeted therapies have revolutionized the prognosis of HER2-positive (HER2+) breast cancer (BC). However, the emergence of acquired resistance or de novo resistance are persistent clinical problems. The objective of this study was to examine neratinib (NER), an irreversible pan-HER inhibitor approved for early-stage and metastatic HER2+ BC, and dasatinib (DAS), a multi-kinase inhibitor used to treat certain leukemias, as a new treatment for HER2-targeted therapy resistant HER2+ BC and triple negative BC (TNBC).

Methods: The anti-proliferative effect of NER plus DAS was screened in a panel of 20 breast cancer cell lines using a 5-day acid phosphatase assay. This panel included 12 HER2+ (3 trastuzumab (TRAS) sensitive (SKBR3, BT474, EFM192A), 2 acquired TRAS resistant (BT474-T, SKBR3-T), 3 innately TRAS resistant (HCC1954, HCC1569, JIMT1), and 4 acquired neratinib resistant cell lines (HCC1954-N, HCC1569-N, SKBR3-N, BT474-N), 6 triple negative (BT20, HCC1143, HCC1937, MDA-MB- 157/231/468), and 2 estrogen receptor positive (ER+) cell lines (MCF7, ZR-75-1). Drug synergy was examined using Combenefit and Calcusyn software. Combination Index (CI) values &lt; 1 indicated synergy, &gt; 1 antagonism. Neratinib IC50 values &lt; 150nM were considered physiologically relevant. Prolonged exposure in vitro efficacy was tested in these cell lines by treating twice weekly with NER +/- DAS until resistance emerged. Cell confluence was then quantified using a crystal violet-based method. The safety and efficacy of NER (10 mg/kg) plus DAS (15 mg/kg) was investigated in vivo in HER2+ HCC1954 xenografts in BALB/c nude mice.

Results: NER was highly effective against HER2+ BC cell lines regardless of TRAS sensitivity (IC50 values range 1-172 nM), showed moderate activity in TNBC (IC50 value range 31 nM - &gt;1 μM), and did not achieve &lt;1 μM activity in ER+ BC cell lines. NER resistant cell lines had IC50 values greater than 150 nM DAS alone displayed IC50 values from 8-135 nM in 4/6 TNBC cell lines, with all 6 TNBC cell lines sensitive to either the single agents or the combination at concentrations less than 100 nM. DAS enhanced the anti-proliferative activity of NER in the HCC1954, HCC1569, JIMT1 cell lines (CI values = 0.27-0.62). The NER/DAS combination overcame NER resistance in all acquired NER-resistant cell line models and no antagonism was observed in any cell line tested. Matrix assays confirmed synergy at low nanomolar concentrations of both drugs. NER plus DAS had a greater long term in vitro anti-proliferative effect over NER in HCC1954, HCC1569, and JIMT1 cells, preventing or delaying the development of NER resistance. NER plus DAS was well tolerated in mice for over 60 days. The combination was better than single agents and showed prolonged anti-tumor effect against HCC1954 xenografts in vivo (p=0.02).

Conclusions: This study suggests that the addition of DAS to NER may have potential as a new treatment strategy for BC, particularly for HER2+ BC.

Citation Format: Neil T. Conlon, Sandra Roche, Fiona O'Neill, Justine Meiller, John Crown, Denis M. Collins. Pre-clinical investigation of neratinib plus dasatinib in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3333.