Francisco J. Esteva

Fatty acid synthase phosphorylation: a novel therapeutic target in HER2-overexpressing breast cancer cells

IntroductionThe human epidermal growth factor receptor 2 (HER2) is a validated therapeutic target in breast cancer. Heterodimerization of HER2 with other HER family members results in enhanced tyrosine phosphorylation and activation of signal transduction pathways. HER2 overexpression increases the translation of fatty acid synthase (FASN), and FASN overexpression markedly increases HER2 signaling, which results in enhanced cell growth. However, the molecular mechanism and regulation of HER2 and FASN interaction are not well defined. Lapatinib is a small-molecule tyrosine kinase inhibitor that blocks phosphorylation of the epidermal growth factor receptor and HER2 in breast cancer cells, resulting in apoptosis. We hypothesized that FASN is directly phosphorylated by HER2, resulting in enhanced signaling and tumor progression in breast cancer cells.MethodsUsing mass spectrometry, we identified FASN as one of the proteins that is dephosphorylated by lapatinib in SKBR3 breast cancer cells. Immunofluorescence, immunoprecipitation, Western blotting, a kinase assay, a FASN enzymatic activity assay, an invasion assay, a cell viability assay and zymography were used to determine the role of FASN phosphorylation in invasion of SKBR3 and BT474 cells. The FASN inhibitor C75 and small interfering RNA were used to downregulate FASN expression and/or activity.ResultsOur data demonstrated that FASN is phosphorylated when it is in complex with HER2. FASN phosphorylation was induced by heregulin in HER2-overexpressing SKBR3 and BT474 breast cancer cells. Heregulin-induced FASN phosphorylation resulted in increased FASN enzymatic activity, which was inhibited by lapatinib. The FASN inhibitor C75 suppressed FASN activity by directly inhibiting HER2 and FASN phosphorylation. Blocking FASN phosphorylation and activity by lapatinib or C75 suppressed the activity of matrix metallopeptidase 9 and inhibited invasion of SKBR3 and BT474 cells.ConclusionsFASN phosphorylation by HER2 plays an important role in breast cancer progression and may be a novel therapeutic target in HER2-overexpressing breast cancer cells.

Phase I/II Study of G3139 (Bcl-2 Antisense Oligonucleotide) in Combination with Doxorubicin and Docetaxel in Breast Cancer

Purpose: Preclinical data showed enhancement of breast cancer cell death when G3139 was combined with anthracyclines and taxanes. We evaluated the efficacy and safety of a Bcl-2 antisense oligonucleotide, G3139, in combination with doxorubicin (A) and docetaxel (T) in patients with locally advanced breast cancer (LABC). Experimental Design: Following a brief phase I to determine the phase II dose, patients with locally advanced breast cancer received G3139 administered by continuous i.v. infusion for 5 to 7 days with bolus A (50 mg/m2) and T (75 mg/m2) administered on either day 3 or 6 of therapy with G3139. Cycles were repeated every 21 days × 6 in the neoadjuvant setting. Serial plasma samples were obtained for pharmacokinetic analysis. Tissue samples were obtained before and after therapy for pharmacodynamic analysis of Bcl-2 expression. Results: Thirty patients (median age, 49 years; range, 24-71 years) received 160 cycles. During the phase I portion of the trial, the dose of G3139 was escalated from 3 to 7 mg/kg/d (i.v. for 5 days) in combination with AT. During the phase II portion of the trial, several doses and schedules of G3139 were evaluated. There were no pathologic complete responses. Pharmacodynamic studies showed limited Bcl-2 down-regulation in the primary tumors. Conclusions: G3139 in combination with doxorubicin and docetaxel is well tolerated. No pathologic complete response was seen and pharmacodynamic studies showed very little down-regulation of Bcl-2 in primary tumors, perhaps related to issues with insufficient drug delivery to the intact tumor.

Monoclonal antibody to Her-2 in breast cancer

The human epidermal growth factor receptor 2 (HER-2) is a tyrosine kinase that is overexpressed in 20% to 25% of invasive breast cancers. Amplification of the her-2 gene is associated with an aggressive tumor phenotype and a reduced patient survival rate. The HER-2 status of a tumor is the critical determinant of response to the HER-2–targeted monoclonal antibody trastuzumab (Herceptin; Genentech, South San Francisco, CA). Thus, accurate assessment of HER-2 expression levels is essential for identifying breast cancer patients who will benefit from trastuzumab. Trastuzumab combined with chemotherapy increases response rates, time to disease progression, and survival duration in patients with metastatic breast cancer and those with early-stage breast cancer. This chapter reviews the development of trastuzumab from a molecular breakthrough to clinical application.