Women with HER2–overexpressing breast cancers have poor prognosis, and many are resistant to the HER2 monoclonal antibody trastuzumab. A subgroup of HER2–overexpressing tumors also express p95HER2, an amino terminally truncated receptor that has kinase activity. Because p95HER2 cannot bind to trastuzumab but should be responsive to the HER2 tyrosine kinase inhibitor lapatinib, we compared the sensitivity of tumors expressing p95HER2 and tumors expressing the full-length HER2 receptor to these agents. MCF-7 and T47D breast cancer cells were stably transfected with either full-length HER2 or p95HER2. We studied the effects of trastuzumab and lapatinib on receptor signaling, cell proliferation, and the growth of xenograft tumors. A paraffin-based immunofluorescence assay was developed to study the association between p95HER2 expression and sensitivity to trastuzumab in patients with advanced breast cancer. All statistical tests were two-sided. Treatment of p95HER2–expressing cells with lapatinib inhibited p95HER2 phosphorylation, reduced downstream phosphorylation of Akt and mitogen-activated protein kinases, inhibited cell growth (MCF-7p95HER2 clones, lapatinib versus control, mean growth inhibition = 57.6% versus 22.6%, difference = 35%, 95% confidence interval [CI] = 22.5% to 47.3%; P <.001; T47Dp95HER2 clones, lapatinib versus control, mean growth inhibition = 36.8% versus 20%, difference = 16.8%, 95% CI = 11.3% to 22.3%, P <.001), and inhibited growth of MCF-7p95HER2 xenograft tumors (lapatinib versus control, mean = 288.8 versus 435 mm ³ , difference = 146.2 mm ³ , CI = 73.8 to 218.5 mm ³ , P = .002). By contrast, treatment with trastuzumab had no effect on any of these parameters. Of 46 patients with metastatic breast cancer who were treated with trastuzumab, only one of nine patients (11.1%) expressing p95HER2 responded to trastuzumab (with a partial response), whereas 19 of the 37 patients (51.4%) with tumors expressing full-length HER2 achieved either a complete (five patients) or a partial (14 patients) response ( P = .029). Breast tumors that express p95HER2 are resistant to trastuzumab and may require alternative or additional anti-HER2–targeting strategies.

Phosphatidylinositol-3-kinase (PI3K) pathway deregulation is a common event in human cancer, either through inactivation of the tumor suppressor phosphatase and tensin homologue deleted from chromosome 10 or activating mutations of p110-α. These hotspot mutations result in oncogenic activity of the enzyme and contribute to therapeutic resistance to the anti-HER2 antibody trastuzumab. The PI3K pathway is, therefore, an attractive target for cancer therapy. We have studied NVP-BEZ235, a dual inhibitor of the PI3K and the downstream mammalian target of rapamycin (mTOR). NVP-BEZ235 inhibited the activation of the downstream effectors Akt, S6 ribosomal protein, and 4EBP1 in breast cancer cells. The antiproliferative activity of NVP-BEZ235 was superior to the allosteric selective mTOR complex inhibitor everolimus in a panel of 21 cancer cell lines of different origin and mutation status. The described Akt activation due to mTOR inhibition was prevented by higher doses of NVP-BEZ235. NVP-BEZ235 reversed the hyperactivation of the PI3K/mTOR pathway caused by the oncogenic mutations of p110-α, E545K, and H1047R, and inhibited the proliferation of HER2-amplified BT474 cells exogenously expressing these mutations that render them resistant to trastuzumab. In trastuzumab-resistant BT474 H1047R breast cancer xenografts, NVP-BEZ235 inhibited PI3K signaling and had potent antitumor activity. In treated animals, there was complete inhibition of PI3K signaling in the skin at pharmacologically active doses, suggesting that skin may serve as surrogate tissue for pharmacodynamic studies. In summary, NVP-BEZ235 inhibits the PI3K/mTOR axis and results in antiproliferative and antitumoral activity in cancer cells with both wild-type and mutated p110-α. [Cancer Res 2008;68(19):8022–30]

Background: Lapatinib (L) plus trastuzumab (T) improves outcomes for metastatic human epidermal growth factor 2–positive breast cancer and increases the pathologic complete response in the neoadjuvant setting, but their role as adjuvant therapy remains uncertain. Methods: In the Adjuvant Lapatinib and/or Trastuzumab Treatment Optimization trial, patients with centrally confirmed human epidermal growth factor 2–positive early breast cancer were randomly assigned to 1 year of adjuvant therapy with T, L, their sequence (T→L), or their combination (L+T). The primary end point was disease-free survival (DFS), with 850 events required for 80% power to detect a hazard ratio (HR) of 0.8 for L+T versus T. Results: Between June 2007 and July 2011, 8,381 patients were enrolled. In 2011, due to futility to demonstrate noninferiority of L versus T, the L arm was closed, and patients free of disease were offered adjuvant T. A protocol modification required P ≤ .025 for the two remaining pairwise comparisons. At a protocol-specified analysis with a median follow-up of 4.5 years, a 16% reduction in the DFS hazard rate was observed with L+T compared with T (555 DFS events; HR, 0.84; 97.5% CI, 0.70 to 1.02; P = .048), and a 4% reduction was observed with T→L compared with T (HR, 0.96; 97.5% CI, 0.80 to 1.15; P = .61). L-treated patients experienced more diarrhea, cutaneous rash, and hepatic toxicity compared with T-treated patients. The incidence of cardiac toxicity was low in all treatment arms. Conclusion: Adjuvant treatment that includes L did not significantly improve DFS compared with T alone and added toxicity. One year of adjuvant T remains standard of care.

Purpose: The epidermal growth factor receptor (EGFR) is abnormally activated in cancer and two classes of anti-EGFR agents, monoclonal antibodies and low-molecular-weight tyrosine kinase inhibitors, have shown antitumor activity in patients. Because these two classes of antireceptor agents target the EGFR at different sites, we decided to explore whether the combined administration of gefitinib, a tyrosine kinase inhibitor, and cetuximab, a monoclonal antibody, had superior antitumor activity than either agent given alone. Experimental Design: We studied the effects of the combination of gefitinib and cetuximab in a panel of human cancer cell lines and in an EGFR-dependent human tumor xenograft model (A431). The effects of these two agents on EGFR signaling, proliferation, apoptosis, and vascularization were evaluated. In addition, we analyzed, with cDNA arrays, changes in gene expression profiles induced by both agents. Results: The combined treatment with gefitinib and cetuximab resulted in a synergistic effect on cell proliferation and in superior inhibition of EGFR-dependent signaling and induction of apoptosis. In a series of in vivo experiments, single-agent gefitinib or cetuximab resulted in transient complete tumor remission only at the highest doses. In contrast, suboptimal doses of gefitinib and cetuximab given together resulted in a complete and permanent regression of large tumors. In the combination-treated tumors, there was a superior inhibition of EGFR, mitogen-activated protein kinase, and Akt phosphorylation, as well as greater inhibition of cell proliferation and vascularization and enhanced apoptosis. Using cDNA arrays, we found 59 genes that were coregulated and 45 genes differentially regulated, including genes related to cell proliferation and differentiation, transcription, DNA synthesis and repair, angiogenesis, signaling molecules, cytoskeleton organization, and tumor invasion and metastasis. Conclusions: Our findings suggest both shared and complementary mechanisms of action with gefitinib and cetuximab and support combined EGFR targeting as a clinically exploitable strategy.