The monoclonal antibody (mAb) therapies trastuzumab (T) and pertuzumab (P) are used to treat the ∼20% of BCs classed as HER2-positive (+). The major modes of action of the mAbs are abrogation of HER2-mediated intracellular signaling, and engagement of immune cells through ADCC. The HER-family targeting TKIs lapatinib (LAP) and neratinib (NER) are approved for the treatment of HER2+ BC. LAP and NER can alter the expression of HER2 and the ADCC response to mAbs in vitro. This in vitro study examines the impact of the TKIs on the levels of HER2, MHC CLASS I, mAb binding and NK cell-mediated ADCC in BC cell line models.
HER2 + (SKBR3 and HCC1954) and HER2-low (MCF-7 and T47D) cell lines were treated with LAP or NER (2µM for 48 hr). HER2 and MHC CLASS I levels were examined by Western blot. Fluorescently-tagged T and P were utilized to examine mAb binding in TKI-treated cell lines using flow-cytometry (% positive cells). A flow cytometry-based assay utilized TKI pre-treated HCC1954 and T47D and healthy volunteer CD56+ NK cells to assess T, P and T+P-mediated ADCC.
LAP-induced increases and NER-induced decreases in HER2 levels were confirmed in all four cell lines by Western blot. In SKBR3 and HCC1954, 98.8% of cells stained T+ and TKI treatment had minimal impact (<3.7% change). In a low-antigen setting, LAP increased (p < 0.05) the T+ cells by 4% (T47D) or 13.5% (MCF-7). NER decreased (p < 0.001) T+ cells by 40.5% (T47D) and 58.4% (MCF-7). 91.7 ± 4.1% SKBR3 and 77.6 ± 4.9% HCC1954 cells stained P+. LAP increased the % P+ cells for both cell lines, significantly so in HCC1954. NER reduced (p < 0.001) P+ cells in both SKBR3 (67.8% decrease) and HCC1954 (50.9% decrease). The TKIs impacted dual T/P binding in the same manner as for each individual mAb. MHC CLASS I expression was reduced (p < 0.05) by LAP in HCC1954 and by NER in T47D. Reduced MHC CLASS I levels were associated with a significant decrease in T-mediated, but not T/P-mediated, ADCC in HCC1954 and T47D.
LAP and NER alter HER2 levels, mAb binding and MHC CLASS I levels. Further investigation is warranted as antigen levels may not be the only factor associated with TKI-altered NK cell-mediated ADCC.
Molecular Therapeutics for Cancer Ireland, National Institute for Cellular Biotechnology, Dublin City University, Glasnevin, Dublin, Ireland.
The Cancer Clinical Research Trust (Charity No.: CHY12210), Science Foundation Ireland-funded Molecular Therapeutics for Cancer Ireland (08/SRC/B1410), Irish Cancer Society Research Centre Breast Predict, The Irish Research Council, Roche.
D.M. Collins: Research funding: Roche, Puma Biotechnology Inc.; Postdoctoral fellowship held for Roche. N.T. Conlon: Research funding: Puma Biotechnology, Inc. N. O’Donovan: Research funding: Roche, GSK. J.P. Crown: Advisory board meetings: Roche, Novartis, Puma Biotechnology Inc.; Travel support for conference attendance: Roche; Research funding: Roche, Puma Biotechnology Inc., GSK. All other authors have declared no conflicts of interest.