{'fraction_non_alphanumeric': 0.029047415634344296, 'fraction_numerical': 0.023067065356685176, 'mean_word_length': 4.855, 'pattern_counts': {'":': 0, '<': 0, '<?xml version=': 0, '>': 0, 'https://': 0, 'lorem ipsum': 0, 'www.': 0, 'xml': 0}, 'pii_count': 0, 'substrings_counts': 0, 'word_list_counts': {'cursed_substrings.json': 0, 'profanity_word_list.json': 0, 'sexual_word_list.json': 0, 'zh_pornsignals.json': 0}}
Lapatinib-induced senescent-like phenotype in HER2-positive breast cancer cells.

583 Background: Resistance to HER2 targeted therapies is a significant clinical problem. To study acquired resistance to the dual EGFR/HER2 tyrosine kinase inhibitor lapatinib (L) in breast cancer, we conditioned HER2 positive breast cancer cell lines with L for 6 months. In HCC1419 cells, L conditioning caused growth arrest which was only reversed when L was removed. We hypothesize that L treatment may induce senescence in HCC1419 cells. Cellular senescence is defined by loss of proliferative potential, altered morphology (large, flattened cells), and expression of specific biomarkers such as senescence-associated β-galactosidase (β-Gal).


METHODS
Following treatment with 250 nM L, cells were fixed and stained for β-gal activity using the Senescence β-gal Staining Kit (Cell Signaling Technology). β-gal activity, which stains the perinuclear compartment blue, is the most widely used marker of senescence. Proliferation assays were performed in 96-well plates, cells were treated with L (0 - 10 µM) and incubated for 5 days. Proliferation was measured using the acid phosphatase assay.


RESULTS
After 6 months conditioning in 250 nM L, HCC1419 cells were no longer proliferating, displayed an altered morphology, and showed significant induction of β-gal activity suggesting L-induced senescence. When L was removed from the media the cells began actively proliferating, and demonstrated similar sensitivity to HCC1419 cells prior to L treatment (12.6% versus 11.0% growth in 1 µM L). We performed a time course assay and found that 1 week of L treatment is sufficient to induce the senescent phenotype in HCC1419 cells. Removal of L from the media for 1 week reverses this phenotype and returns the β-gal staining to pre-treatment levels. This indicates that the L-induced senescence is reversible in the absence of L.


CONCLUSIONS
L induces a senescent-like phenotype in HCC1419 cells and removal of L from the cells results in the reversal of senescence and active proliferation of the cells. This suggests a mechanism by which breast cancer cells can survive long term L treatment and lead to tumor recurrence when L is discontinued. Breast tumors which undergo this dormant senescent-like phenotype may be sensitive to re-treatment with L.