ABSTRACT: Overexpression by the HER2 gene plays a significant role in breast cancer pathogenesis, and the phenomenon is commonly regarded as a predictor of a poor prognosis. HER2 overexpression has been linked to sensitivity and/or resistance to hormone therapy and chemotherapeutic regimens, including CMF (cyclophosphamide, methotrexate(Drug information on methotrexate), and fluoro-uracil) and anthracyclines. Studies of patients with advanced disease demonstrate that, despite the association of HER2 overexpression with poor prognosis, the odds of HER2-positive patients responding clinically to taxanes were greater than three times those of HER2-negative patients. Further studies in preclinical models used combination therapy for breast cancer cells that overexpress HER2, and the use of agents that interfere with HER2 function plus paclitaxel(Drug information on paclitaxel) (Taxol) resulted in significant antitumor effects. [ONCOLOGY 11(Suppl):43-48, 1997]
Introduction
The taxanes are an important new class of anticancer agents with a unique mechanism of action.[1] Paclitaxel (Taxol), the first taxane used in clinical trials, was originally isolated in 1971 from the bark of the Pacific yew, Taxus brevifolia. It was selected for clinical development based on impressive antitumor activity against the implanted B16 melanoma and the human MX-1 mammary tumor xenograft.
Since then, paclitaxel has been shown to have a high degree of antitumor activity in women with metastatic breast cancer, as well as a lack of cross-resistance with anthracyclines. Current clinical research with paclitaxel in breast cancer is focused on several aspects, including: optimal dosing and scheduling; the agent's role in the treatment of early breast cancer; its use in high-dose intensity therapy of breast cancer; and combination therapy of paclitaxel with other antineoplastic agents.
An area of increasing interest in clinical research on taxanes is the possible role of oncogenes, such as HER2, in determining clinical response to paclitaxel. Studies have examined whether strategies can be designed to increase the agent's efficacy (or curb resistance to it) in breast cancers that overexpresess HER2. Available data that will be presented in this review suggest that HER2 overexpression may influence the response to paclitaxel in patients with metastatic breast cancer and that anti-HER2 monoclonal antibodies significantly increase the antitumor activity of paclitaxel in vitro and in vivo.
HER2/c-erbB-2/neu in Breast Cancer
During the last decade, proto-oncogenes encoding growth factors and growth factor receptors have been found to play important roles in the pathogenesis of several human malignancies, including breast cancer.[2] The HER2 gene (also known as neu and as c-erbB-2) encodes a 185-kD transmembrane glycoprotein receptor (p185HER2) that has partial homology with the epidermal growth factor (EGF) receptor; the gene shares intrinsic tyrosine kinase activity with the receptor.[3-5] The EGF receptor and p185HER2 belong to a superfamily of receptors known as type I tyrosine kinase receptors, which have an extracellular ligand-binding domain, a transmembrane lipophilic segment, and an intracellular protein tyrosine kinase domain with a regulatory carboxyl terminal segment.[6]
HER2 is overexpressed in 25% to 30% of human breast cancers[7,8] and indicates a worse prognosis in patients who have positive axillary lymph nodes.[6,7,9] The observation that HER2 overexpression is associated with a poor prognosis could imply that HER2 is solely a marker event. On the other hand, HER2 could be a prognostic factor that plays a role in the pathogenesis of breast cancer.
Role of Monoclonal Antibodies
Several lines of evidence support a direct role for HER2 in the pathogenesis and clinical aggressiveness of overexpressing tumors: (1) The introduction of HER2 into nonneoplastic cells causes their malignant transformation.[10,11] (2) Transgenic mice expressing HER2 develop mammary tumors.[12] (3) HER2 overexpression is common in ductal carcinomas in situ and in their associated invasive cancers.[13,14] (4) The mechanisms responsible for this growth advantage are thought to be related to the fact that p185HER2 overexpression results in activation of a series of signaling pathways (PLC-gamma/phosphatidylinositol; PI(3)kinase; STAT91/ISGF-3; ras/raf/MAP-kinase pathway; src family), and that activation of these pathways results in gene activation that ultimately results in cell proliferation.[6] (5) Antibodies directed against p185HER2 can inhibit the growth of tumors and of transformed cells that express high levels of this receptor.[15-19]
The latter observation suggests that p185HER2 may be a potential target for the treatment of breast cancer or preinvasive breast lesions because these cells commonly overexpress HER2. The murine monoclonal antibody (MoAb) 4D5, directed against the extracellular domain of p185HER2 (ECDHER2), is a potent inhibitor of in vitro growth and, in xenograft models, of human breast cancer cells overexpressing HER2.[20-22] Murine MoAbs, however, are limited clinically because they are immunogenic. Therefore, to facilitate further clinical investigations, MoAb 4D5 was humanized. The resulting recombinant humanized anti-p185HER2 MoAb (rhuMoAb HER2) was found to be safe and to have dose-dependent pharmacokinetics in two prior phase I clinical trials.
HER2 Overexpression as a Predictor of Response to Taxanes
Since HER2 plays a role in breast cancer pathogenesis and HER2 overexpression correlates with more aggressive clinical behavior, several studies have attempted to correlate whether HER2 overexpression is a predictor for response to systemic therapy. Studies to date have shown that HER2 overexpression predicts a worse response to hormonal therapy with tamoxifen(Drug information on tamoxifen) (Nolvadex) in advanced disease[23,24] and in early-stage breast cancer patients.[25]
Dose-Response Effects to Anthracyclines
The relationship between HER2 overexpression and response to chemotherapy appears to be more complex. Data indicate that HER2-positive tumors have increased resistance to adjuvant CMF (cyclophosphamide, methotrexate, and fluorouracil(Drug information on fluorouracil))-based therapy[26,27] and, conversely, increased dose-response effects to an anthracycline-containing regimen.[28]
In Intergroup Study 0011,[26] patients with primary breast cancer tumors larger than 3 cm or with estrogen receptor-negative tumors were randomized to receive either CMF chemotherapy or observation. Patients with HER2-negative tumors who received adjuvant chemotherapy showed significantly improved disease-free survival when compared with untreated patients. In contrast, patients with HER2-positive tumors showed no benefit from adjuvant therapy.
In the International Breast Cancer Study Group trial, patients with node-positive early breast cancer were randomized to receive either one cycle of perioperative chemotherapy or prolonged adjuvant chemotherapy, defined as six cycles of CMF-based chemotherapy.[27] In this study, the effect of prolonged chemotherapy was greater in patients with HER2-negative tumors.
The possible predictive role of HER2 overexpression has also been analyzed in patients with early breast cancer treated with anthracycline-containing adjuvant therapy. In a well-known randomized study by the Cancer and Leukemia Group B (CALGB), three doses (high, moderate, and low) of cyclophosphamide(Drug information on cyclophosphamide), doxorubicin(Drug information on doxorubicin), and fluorouracil were compared in women with node-positive breast cancer.[28] Patients randomly assigned to the high-dose regimen of adjuvant chemotherapy had significantly longer disease-free and overall survival if their tumors overexpressed HER2. This dose-response effect was not observed in patients whose tumors had minimal or no HER2 expression.
Thus, findings from the CALGB study suggest that there is a significant dose-response effect of adjuvant therapy with an anthracycline-containing regimen in patients with HER2 overexpression but not in patients with no or minimal HER2 expression. The group's final conclusion was that HER2 overexpression may be a useful marker for identifying patients who are most likely to benefit from high doses of adjuvant doxorubicin-based chemotherapy.
HER2 Overexpression and Taxane Sensitivity
Because taxanes are becoming widely used in the management of advanced breast cancer, we decided to analyze whether there is a relationship between HER2 expression and clinical sensitivity to taxanes. In a study performed at Memorial Sloan-Kettering Cancer Center, the possible relationship between HER2 overexpression and response to taxanes was analyzed in patients with metastatic breast cancer.[29] HER2 expression was studied in patients treated with one of eight protocols of single-agent taxane therapy over the past 5 years. All patients had bidimensionally measurable disease and histologically confirmed metastatic breast cancer.
Archived paraffin(Drug information on paraffin)-embedded tumor tissue was available for immunohistochemistry in 122 patients out of the total of 212 patients treated; of these, 102 (84%) received paclitaxel and 20 (16%), docetaxel(Drug information on docetaxel) (Taxotere). Geographic considerations were the most frequent obstacle in lack of tissue availability for HER2 analysis.
Tumor expression of HER2 was determined by immunohistochemical analysis of a set of thin sections prepared from patients' paraffin-archived tumor blocks (as previously described[7,8]). The primary detecting antibody used was murine MoAb 4D5, directed at the extracellular domain of p185HER2. Tumors were considered to overexpress HER2 if at least 10% of the tumor cells exhibited characteristic membrane staining for p185HER2.
Seven prognostic factors were assessed for association with tumor response: HER2 overexpression, scored as positive or negative; estrogen-receptor status; extent of disease, divided into one to two involved organ systems vs three or more involved systems; extent of prior chemotherapy, categorized as one to two courses or more than two courses; presence of visceral disease; prior therapy with doxorubicin; and Karnofsky performance status, stratified as 60 to 80 or 90 to100.
In 37.7% of patients, tumors were positive by immunohistochemistry with the 4D5 antibody. The overall response to taxanes for all patients in this analysis was 46.7%. Remarkably, 65.2% of patients with HER2-positive tumors responded vs 35.5% of patients with HER2-negative tumors. Using a Mantel-Haenszel test, the P value for this difference was significant at .002. Visceral dominance (P = .011), low performance status (P = .057), and extensive prior therapy correlated with poor clinical response. Among these, HER2 overexpression was positively correlated with low performance status (P = .002), and low performance status with extensive prior therapy.
These correlations should bias against response in HER2-positive cases, which was not observed. Indeed, stratified analysis controlling for confounding variables demonstrated the value of HER2 status in predicting taxane response. The odds ratios of response for HER2-positive vs HER2-negative tumors were 3.95 after adjusting for visceral disease, 3.17 after adjusting for number of prior therapies, and 3.06 after adjusting for performance status. Thus, despite a positive correlation of HER2 expression and poor prognostic features, the odds of HER2-positive patients responding clinically to taxanes were greater than three times those of HER2-negative patients.
Tentative Results With Polyclonal Antibody
Tumor specimens were also analyzed with a rabbit polyclonal antibody directed at the cytoplasmic c-terminus epitope of p185HER2. Immunohistochemical evaluation with this antibody resulted in a higher proportion of HER2 positivity (57%). Patients who were shown to have HER2-positive tumors using this antibody were more likely to respond to taxanes, although the difference was not statistically significant (P = .3).
Results with this polyclonal antibody have to be viewed with caution because 57% of the tumors stained positive for HER2 overexpression--a higher proportion than with the MoAb, and higher than has been reported previously. Further confirmatory studies are needed to verify our results, which could have significant implications for the treatment of patients with advanced breast cancer.
Paclitaxel and the Signal Transduction Pathway
The possible mechanisms underlying the interaction between HER2 overexpression and taxane sensitivity are unknown. Paclitaxel stabilizes microtubules, prevents tubulin depolymerization, and promotes tubulin bundling. In addition to this well-documented mechanism of action, some evidence suggests that paclitaxel activates key elements of the HER2 signal transduction pathway (Figure 1).[30-32]
Interaction of Paclitaxel With p185HER2 Signal Transduction Pathway
The mitogen-stimulated protein serine/threonine kinase c-raf-1 functions as a central component of the mitogen-activated protein kinase (MAP kinase) signal transduction pathway.[33] In MCF-7 breast cancer cells, paclitaxel treatment leads to activation of c-raf-1, documented by a reduced c-raf-1 electrophoretic mobility after paclitaxel exposure.[30] Furthermore, paclitaxel therapy induces a dose- and time-dependent accumulation of the cyclin inhibitor of p21WAF1, and c-raf-1 depletion prevents this activation. In addition to c-raf-1 activation, tyrosine phosphorylation of MAP kinase is a well-documented response to paclitaxel, and it probably represents a functionally important event by the agent.[30,32]
The activation of the HER2 signal transduction pathway by paclitaxel could also result in activation of paclitaxel-induced apoptosis.[31] Thus, HER2 overexpression would provide an increased opportunity to enhance the cytotoxic effects of paclitaxel.
