Trastuzamab antibody drug conjugates demonstrate robust in vitro antitumor activity in HER2 amplified breast cancer cell lines

Breast cancer is a heterogeneous disease with molecular subtypes characterized by distinct histopathology and gene expression signatures. HER2 oncogene amplification and over-expression is a defining feature of the HER2-amplified subtype of breast cancer. Currently, assays for detecting gene amplification and protein over-expression are used as diagnostic assays for the selection of patients for HER2-targeted therapies. In the present study, we compared the sensitivity of several HER2-targeted therapies across a panel of 31 breast cancer cell lines and correlated the sensitivity of the cell lines with HER2 positivity, as determined by IHC, qPCR and FISH. The agents utilized for the cell line screening were trastuzamab (Herceptin), two trastuzamab antibody-drug conjugates (ADCs) that employ an anti-mitotic agent, and a small molecule inhibitor of HER2 kinase activity. HER2 testing revealed 32% of the cell lines to be positive for HER2 by qPCR and FISH, and 90% of these demonstrated gene amplification in the form of clusters, as detected by FISH, or more than 10 copies as detected by qPCR. Several of the cell lines exhibited polysomy for chromosome 17 with accompanying extra copies of HER2 but did not meet the criteria for HER2 amplification. The HER2 IHC assay revealed 10 cell lines to be positive (2+, 3+) by IHC, with 8 exhibiting high-level expression (3+). The agents tested were more efficacious in the HER2-amplified cell lines, with wideranging activity observed in response to the ADCs. Cell lines exhibiting polysomy for HER2 and 1+ expression by IHC were less responsive to any of the agents tested. Consistent with previous reports, many of the cell lines resistant to trastuzamab or the small molecule HER2 inhibitor exhibited deregulation of the PI-3 kinase pathway illustrated by mutations in the PIK3CA gene or loss of PTEN protein expression. Interestingly, the majority of these resistant cell lines were sensitive to ADCs. In summary, our study demonstrates a high sensitivity to ADCs in breast cancer cell lines harboring high-level amplification of the HER2 gene and suggests these agents may overcome in vitro resistance conferred by activation of the PI-3 kinase pathway.