Purpose: A subgroup of human epidermal growth factor receptor 2 (HER2)-overexpressing breast tumors coexpresses p95HER2, a truncated HER2 receptor that retains a highly functional HER2 kinase domain but lacks the extracellular domain and results in intrinsic trastuzumab resistance. We hypothesized that lapatinib, a HER2 tyrosine kinase inhibitor, would be active in these tumors. We have studied the correlation between p95HER2 expression and response to lapatinib, both in preclinical models and in the clinical setting. Experimental Design: Two different p95HER2 animal models were used for preclinical studies. Expression of p95HER2 was analyzed in HER2-overexpressing breast primary tumors from a firstline lapatinib monotherapy study (EGF20009) and a second-line lapatinib in combination with capecitabine study (EGF100151). p95HER2 expression was correlated with overall response rate (complete + partial response), clinical benefit rate (complete response + partial response + stable disease ≥24 wk), and progression-free survival using logistic regression and Cox proportional hazard models. Results: Lapatinib inhibited tumor growth and the HER2 downstream signaling of p95HER2-expressing tumors. A total of 68 and 156 tumors from studies EGF20009 and EGF100151 were evaluable, respectively, for p95HER2 detection. The percentage of p95HER2-positive patients was 20.5% in the EGF20009 study and 28.5% in the EGF100151 study. In both studies, there was no statistically significant difference in progression-free survival, clinical benefit rate, and overall response rate between p95HER2-positive and p95HER2-negative tumors. Conclusions: Lapatinib as a monotherapy or in combination with capecitabine seems to be equally effective in patients with p95HER2-positive and p95HER2-negative HER2-positive breast tumors. ©2010 AACR.