Originally published as JCO Early Release 10.1200/JCO.2007.13.5202 on May 19 2008

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Phase I Dose Escalation and Pharmacokinetic Study of Lapatinib in Combination With Trastuzumab in Patients With Advanced ErbB2-Positive Breast Cancer

Anna Maria Storniolo, Mark D. Pegram, Beth Overmoyer, Paula Silverman, Nancy W. Peacock, Suzanne F. Jones, Jill Loftiss, Nikita Arya, Kevin M. Koch, Elaine Paul, Lini Pandite, Ronald A. Fleming, Peter F. Lebowitz, Peter T.C. Ho, Howard A. Burris, III

From the Indiana University Melvin and Bren Simon Cancer Center, Indianapolis, IN; Sarah Cannon Research Institute, Nashville, TN; UCLA Center for the Health Sciences, Los Angeles, CA; Northwestern CT Oncology/Hematology Associates, Torrington, CT; Ireland Cancer Center, University Hospitals Case Medical Center, Case Western Reserve University, Cleveland, OH; and GlaxoSmithKline, Research Triangle Park, NC

Corresponding author: Anna Maria Storniolo, MD, Indiana University Melvin and Bren Simon Cancer Center, 535 Barnhill Dr, Room 473, Indianapolis, IN 46202; e-mail: astornio{at}iupui.edu

	ABSTRACT

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Purpose The combination of lapatinib and trastuzumab has been observed to have a synergistic, antiproliferative effect against ErbB2-positive breast cancer cells in vitro. This phase I study assessed the safety, clinical feasibility, optimally tolerated regimen (OTR), pharmacokinetics (PK), and preliminary clinical activity of this combination in patients with ErbB2-positive advanced breast cancer.

Patients and Methods Cohorts of three patients with ErbB2-positive advanced breast cancer were treated with escalating doses of lapatinib (750 to 1,500 mg) administered once daily (continuous) in combination with trastuzumab (4 mg/kg loading dose then 2 mg/kg weekly) to determine the OTR. Once the OTR was determined, additional patients were enrolled to provide the PK profile of both agents alone and in combination.

Results A total of 54 patients were treated: 27 in the dose-escalation group and 27 in the PK group. Overall, adverse events were mild to moderate in severity, with no drug-related grade 4 events. The most frequent drug-related grade 3 events included diarrhea (17%), fatigue (11%), and rash (6%). The OTR was 1,000 mg lapatinib with standard weekly trastuzumab. One patient had a complete response and seven patients had partial responses. The PK parameters (maximum concentration in plasma and area under the curve) of lapatinib and trastuzumab in combination were not significantly different than when either was administered alone.

Conclusion The OTR of the lapatinib/trastuzumab combination was lapatinib 1,000 mg per day with standard weekly trastuzumab. At these doses, the regimen was well tolerated and clinically active in this heavily pretreated ErbB2-positive breast cancer population.

	INTRODUCTION

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Identification of the mechanisms of cellular growth and survival in normal and malignant cells has led to the development of targeted therapies that hold much promise in the treatment of breast cancer. The ErbB or HER family of transmembrane tyrosine kinase receptors, specifically epidermal growth factor receptor (ErbB1) and human epidermal growth factor receptor 2 (ErbB2), are implicated in the pathogenesis of breast cancer. Overexpression of ErbB1 is reported in 14% to 91% of all primary breast cancers.¹ Overexpression of ErbB2 is reported in approximately 20% of human breast cancers and is associated with a poor prognosis.^2,3 ErbB receptor tyrosine kinase activation requires dimerization of the receptors. The ErbB2 receptor is the preferred dimerization partner for all other members of the ErbB family.⁴ Because ErbB receptor heterodimers generate potent tumor growth and survival signals, simultaneous inhibition of both ErbB1 and ErbB2 is an appealing therapeutic strategy.

Lapatinib (Tykerb; GlaxoSmithKline, Research Triangle Park, NC) is an oral, dual-tyrosine kinase inhibitor with specificity for both the ErbB1 and ErbB2 receptors.^5,6 Lapatinib acts intracellularly by reversibly binding to the cytoplasmic adenosine triphosphate–binding site, thereby preventing receptor phosphorylation and activation.⁷ Thus, lapatinib simultaneously blocks two of the major downstream pathways that are activated after ErbB receptor stimulation: the ERK1/2 pathway, which modulates cell proliferation, and the PI3K/Akt pathway, which regulates cell survival.^5,8

Trastuzumab, a humanized monoclonal antibody, targets the extracellular domain of ErbB2, with subsequent inhibition of receptor signaling.⁹ Preclinical data indicate that lapatinib has a synergistic effect with trastuzumab in ErbB2-positive breast cancer cells.¹⁰ Simultaneous inhibition of the ErbB2 receptor by two different modalities may have increased antitumor activity when compared with each agent alone. The objectives of this phase I dose-escalation study were to determine the safety, optimally tolerated regimen (OTR), pharmacokinetics, and preliminary anticancer activity of this combination in patients with advanced ErbB2-positive breast cancer.

	PATIENTS AND METHODS

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Eligibility Criteria
The study was conducted at four sites in the United States, was approved by the institutional review boards of participating institutions, and was conducted in accordance with the 1996 Declaration of Helsinki. Signed informed consent was obtained from all patients. Men or women 18 years of age with advanced or metastatic, histologically/cytologically confirmed breast cancer that overexpressed the ErbB2 protein (immunohistochemistry 2+ or 3+) or ERBB2 gene amplification by fluorescence in situ hybridization were eligible (ErbB2 assays were conducted locally and not confirmed centrally). Prior treatment with trastuzumab was permitted but not required. Other eligibility criteria included: life expectancy 12 weeks; Karnofsky performance status of 70; adequate bone marrow, liver and kidney function; and baseline left ventricular ejection fraction (LVEF) higher than 50%. Patients who had received prior maximum cumulative doxorubicin doses higher than 400 mg/m², those with uncontrolled brain or leptomeningeal metastases, pre-existing cardiac dysfunction, symptomatic intrinsic pulmonary disease, or extensive tumor involvement of the lungs were ineligible.

Pretreatment Evaluations and Definitions and Treatment Schemes
Screening assessments consisted of a complete physical examination, medical history, medication history, disease assessment, atrial natriuretic factor (ANF), multigated angiogram (MUGA) or echocardiogram (ECHO), 12-lead ECG, CBC, clinical chemistry, and urinalysis.

Patients (in cohorts of at least three during dose escalation) received daily oral lapatinib concurrently with a standard dose of trastuzumab given intravenously (IV): 4 mg/kg loading dose then 2 mg/kg weekly (Fig 1). Lapatinib was supplied as 250-mg tablets for daily oral administration. Commercially available trastuzumab (Herceptin; Genentech, South San Francisco, CA) was used. Lapatinib could be taken with a light, low-fat breakfast except on pharmacokinetic sampling days where patients fasted 4 hours before lapatinib dosing and 2 hours afterward.

View larger version (24K): [in this window] [in a new window] [PowerPoint Slide for Teaching]   Fig 1. Study design. Lapatinib pharmacokinetics (PK; at steady-state on seventh day of dosing): sequence 1: day 7 alone, day 15 with trastuzumab; sequence 2: day 7 alone, day 8 with trastuzumab. Trastuzumab PK: sequence 1: day 15 alone, day 22 with lapatinib; sequence 2: day 8 with lapatinib, day 22 alone.

DLTs were defined as grade 3 or 4 nonhematologic toxicity (excluding grade 3 nausea); grade 4 granulocytopenia lasting 5 days, with or without fever (> 38.5°C); or thrombocytopenia 25,000/mm³. Any grade 2 toxicity which persisted beyond the initial 4-week treatment period and was considered dose-limiting by the clinical investigator was considered a DLT.

The OTR was defined as the dose of lapatinib and trastuzumab at which no more than one of six patients experienced a DLT. Once the OTR was determined, additional patients (up to 10) were to be treated at the OTR to further evaluate the safety and tolerability of the combination.

Evaluation During Therapy
Before each treatment cycle, blood for hematology and clinical chemistry analyses was obtained and results reviewed before continued dosing with lapatinib and trastuzumab. Vital signs, weight, and performance status evaluation were obtained before starting each treatment cycle. Cardiac function was monitored closely; a MUGA or ECHO (investigators were requested to use the same method throughout study), ANF, and 12-lead ECG were performed at the end of treatment cycles 1 and 2 (week 4 and 8), at every two treatment cycles after that, and at discontinuation. All patients continued on study until disease progression, unacceptable toxicity, or withdrawal of consent.

Pharmacokinetic Study
Additional patients were enrolled at the OTR dose to characterize the pharmacokinetics of lapatinib and trastuzumab administered alone and in combination and randomly assigned into two treatment sequences (Fig 1). A trastuzumab washout period of 5 weeks was required for patients in the pharmacokinetic group who had received trastuzumab within 5 weeks of study entry. These patients received lapatinib monotherapy during the trastuzumab washout period. Serial blood samples were collected over a 24-hour period after ingestion of the lapatinib dose and/or the initiation of the 0.5-hour trastuzumab infusion. After the completion of the PK phase, patients still receiving clinical benefit could remain on study receiving the OTR.

Lapatinib was analyzed by a previously published liquid chromatography with tandem mass spectrometry method with a sensitivity of 5 ng/mL.¹¹ Trastuzumab was analyzed by an enzyme immunoassay method with a sensitivity of 10 µg/mL. Plasma concentration data were analyzed by standard noncompartmental methods using WINNonlin (Scientific Consultant, Apex, NC) professional software version 4.1 (Pharsight Corp, Mountain View, CA). Area under the plasma concentration-time curve (AUC) and maximum concentration in plasma (C_max) were compared using analysis of variance.

Response and Toxicity Criteria
Tumor response was assessed at the end of treatment cycle 2 (8 weeks from the beginning of drug therapy including the start of monotherapy lapatinib for patients in the PK group requiring a washout period) and then every 8 weeks by x-ray, computed tomography scan, magnetic resonance imaging, or clinical examination using the Response Evaluation Criteria In Solid Tumors guidelines. All adverse events (AE) were graded by the National Cancer Institute Common Toxicity Criteria (NCI-CTC), version 2.

	RESULTS

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Patient Population
A total of 54 patients were enrolled; 27 in the dose-escalation group and 27 in the pharmacokinetic group. The median age was 53 years (range, 30 to 80). Additional patient characteristics are provided in Table 1. The majority of patients were heavily pretreated (median prior cytotoxic regimens = 4; range, 1 to 9) and 50 patients had received prior trastuzumab treatment (median prior trastuzumab-containing regimens = 2; range, 0 to 7). The number of patients receiving lapatinib starting doses of 750, 1,000, 1,250, and 1,500 mg/d were three, 38, 10, and three, respectively.

Dose level –1 (lapatinib 750 mg) enrolled three patients. No DLTs were reported. The dose level was well tolerated and based on the absence of DLTs at the current dose level and the previous level (1,000 mg), the investigators recommended increasing the dose level to lapatinib 1,250 mg.

Dose level +1 (lapatinib 1,250 mg) enrolled five patients. No patients experienced a DLT and the investigators recommended increasing the dose level to lapatinib 1,500 mg.

Dose level +2 (lapatinib 1,500 mg) enrolled three patients. Two patients experienced DLTs of grade 3 fatigue and grade 3 diarrhea, nausea, and vomiting, respectively. Because two of three patients experienced a DLT, the decision was made to explore a lower lapatinib dose with the standard dose of trastuzumab. Given no clear differentiation between dose level 1 and 2, the decision was made to sequentially enroll patients at the 1,000-mg and 1,250-mg dose levels to further explore safety and tolerability.

Dose level 0 (lapatinib 1,000 mg) enrolled seven additional patients. The last patient enrolled experienced DLTs of grade 3 weakness and fatigue. No other patient experienced a DLT. In an effort to differentiate the toxicity at lapatinib dose levels 1,000 mg and 1,250 mg, the 1,250 mg lapatinib/standard trastuzumab cohort was expanded.

Dose level +1 (lapatinib 1,250 mg) enrolled five additional patients (total 10). One patient experienced grade 1 alopecia, grade 2 rash on scalp, grade 2 pruritus, and grade 3 pain related to rash. All events occurred in one incident, and the cluster was deemed dose limiting. Another patient experienced a DLT of grade 3 diarrhea. Based on the composite DLTs (two) at 1,250 mg lapatinib with trastuzumab and a higher frequency of fatigue which the investigators felt impacted tolerability, the combination doses of lapatinib 1,000 mg and standard dose trastuzumab were selected as the OTR.

Safety and Tolerability
All 54 patients were assessable for toxicity, and 53 experienced at least one drug-related AE during the study. Drug-related AEs reported by more than 25% of patients were: diarrhea, rash, fatigue, nausea, anorexia, and vomiting (Fig 2). No patients experienced grade 3 AEs in the 750-mg lapatinib group. Fourteen grade 3 drug-related AEs occurred in 11 patients (of a total of 38 patients) treated at the 1,000-mg lapatinib group. Eight grade 3 drug-related AEs were reported in six patients (of a total of 10 patients) in the 1,250-mg lapatinib group. Six grade 3 drug-related AEs were reported in three patients (of a total of three patients) in the 1,500-mg lapatinib group. The most common grade 3 drug-related AEs were diarrhea, fatigue, and rash. No grade 4 drug-related AEs were reported. One patient in the 1,500-mg lapatinib cohort experienced an asymptomatic decline in LVEF (NCI grade 2 cardiac toxicity) to 45% (ECHO) from a baseline value of 58% by MUGA (> 20% decline relative to baseline assessment), after approximately 7 months of treatment. Because the patient was receiving benefit from treatment (a confirmed complete response [CR]), the patient remained on continuous lapatinib and trastuzumab treatment and subsequent LVEF values by MUGA were 50%, 60%, and 55% after approximately eight, 11, and 13 months of treatment.

View larger version (16K): [in this window] [in a new window] [PowerPoint Slide for Teaching]   Fig 2. Frequency of drug-related adverse events in more than 25% of all patients by maximum toxicity grade. Number of patients by lapatinib starting dose: 750 mg (n = 3), 1,000 mg (n = 38), 1,250 mg (n = 10), and 1,500 mg (n = 3).

View this table: [in this window] [in a new window]   Table 2. Summary of Confirmed Clinical Activity and SD of 6 Months Duration

View larger version (20K): [in this window] [in a new window] [PowerPoint Slide for Teaching]   Fig 3. Mean (standard deviation) plasma lapatinib concentration versus time profiles at steady-state, alone, and in combination with trastuzumab.

	DISCUSSION

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Treatment-related fatigue occurred in 52% of all patients (20% grade 1; 20% grade 2; 11% grade 3) and in 50% and 70% of patients at the 1,000- and 1,250-mg dose levels, respectively. In lapatinib phase II monotherapy studies (1,250 to 1,500 mg), fatigue was reported in 31% of patients (11% grade 1; 10% grade 2; 10% grade 3; GlaxoSmithKline internal data). In two large phase II studies of trastuzumab alone administered at the same dose and schedule as this study, fatigue was not reported as an adverse event.^12,13 The mechanism of the fatigue or the reason for increased fatigue in this study, particularly at the higher lapatinib dose levels, is not known.

Given that this study combined two agents targeting ErbB2, assessment of cardiac function was performed during the study. Of 54 patients, only one patient (the patient with the CR) had a decrease in LVEF higher than 20% relative to their baseline assessment. This reversed on subsequent LVEF assessments and the patient remained on therapy without interruption. In a recent analysis of 3,127 patients with cancer receiving lapatinib, a 1.3% incidence of decreased LVEF was reported (NCI CTC grade 3 or 4 or asymptomatic LVEF decline of 20% relative to baseline and below the institution's lower limit of normal). Only 0.1% of patients had symptomatic LVEF dysfunction and this was generally reversible or nonprogressive.¹⁴

Clinical responses were observed at all dose levels of lapatinib tested. All eight subjects with confirmed clinical responses (CR, PR) had received prior trastuzumab therapy. The response rate observed in the pharmacokinetic phase (one of 27 subjects; 4%) was lower than that observed in the dose escalation/OTR phase (seven of 27 subjects; 26%). This difference is likely attributable to treatment differences between these two groups. Nineteen patients in the pharmacokinetic group required a washout period of at least 5 weeks and received monotherapy lapatinib during this period. After the washout period, the pharmacokinetic group received various treatment sequences to evaluate the pharmacokinetics of lapatinib alone, trastuzumab alone, or each agent in combination (Fig 1). In addition to lapatinib monotherapy received during the washout period, all patients in the pharmacokinetic group received an additional 1 week of lapatinib monotherapy (Fig 1). Thus the treatment these patients received during the 3 weeks of the pharmacokinetic study differed from that of patients on the dose escalation/OTR part of the study. Furthermore, the initial disease assessment was performed at 8 weeks from the start of drug treatment, which included the start of lapatinib monotherapy for patients with a required washout period. In review of these subjects, 10 subjects in the pharmacokinetic phase had progressive disease at their first response assessment compared with three subjects in the dose escalation/OTR phase. Thus, subjects on the pharmacokinetic phase were removed from the study before having the opportunity to receive the same duration of combination treatment that the dose escalation/OTR cohort had received.

Combining trastuzumab, which binds the extracellular domain of ErbB2¹⁵ and a small molecule directed against the intracellular kinase may permit the blockade of ErbB2 signaling from both truncated ErbB2 receptors and dimerized ErbB receptors. The ErbB2 receptor, p185^HER2, undergoes proteolytic cleavage of its extracellular domain,^16,17 resulting in a membrane-associated truncated receptor (p95^HER2) that has kinase activity and is tyrosine phosphorylated.¹⁸ The truncated form of ErbB2 can heterodimerize with ErbB3 and is stimulated by heregulin but not EGF.¹⁹ The increased expression of p95^HER2 is associated with a poor prognosis in breast cancer.^18,20,21 In vitro, tumor cells with p95^HER2 are sensitive to lapatinib but not to trastuzumab.¹⁹ This dual inhibition of the full length and truncated ErbB2 receptor may explain why patients in the current study were sensitive to the combination even after multiple lines of prior trastuzumab therapy.

Continued use of trastuzumab alone after progression on trastuzumab-containing regimens is controversial.²² Moreover, the clinical benefit of lapatinib monotherapy in ErbB2-positive patients with breast cancer whose tumors progress on trastuzumab is modest.²³ However, results from this study indicate that the combination of lapatinib and trastuzumab may offer benefit to patients having received prior trastuzumab, including those receiving multiple previous trastuzumab regimens. The combined use of lapatinib and trastuzumab may result in maximal inactivation of the ErbB2 signaling pathway. Given the tolerability, lack of pharmacokinetic interaction, and encouraging clinical anticancer activity in this heavily pretreated advanced breast cancer population, further clinical studies of this combination are justified.

	AUTHORS’ DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST

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Although all authors completed the disclosure declaration, the following author(s) indicated a financial or other interest that is relevant to the subject matter under consideration in this article. Certain relationships marked with a "U" are those for which no compensation was received; those relationships marked with a "C" were compensated. For a detailed description of the disclosure categories, or for more information about ASCO's conflict of interest policy, please refer to the Author Disclosure Declaration and the Disclosures of Potential Conflicts of Interest section in Information for Contributors.

Employment or Leadership Position: Jill Loftiss, GlaxoSmithKline (C); Nikita Arya, GlaxoSmithKline (C); Kevin M. Koch, GlaxoSmithKline (C); Elaine Paul, GlaxoSmithKline (C); Lini Pandite, GlaxoSmithKline (C); Ronald A. Fleming, GlaxoSmithKline (C); Peter F. Lebowitz, GlaxoSmithKline (C); Peter T.C. Ho, GlaxoSmithKline (C) Consultant or Advisory Role: Anna Maria Storniolo, GlaxoSmithKline (C); Mark D. Pegram, Genentech (C), GlaxoSmithKline (C); Beth Overmoyer, GlaxoSmithKline (C); Howard A. Burris III, GlaxoSmithKline (C), Genentech (C) Stock Ownership: Jill Loftiss, GlaxoSmithKline; Kevin M. Koch, GlaxoSmithKline; Elaine Paul, GlaxoSmithKline; Lini Pandite, GlaxoSmithKline; Ronald A. Fleming, GlaxoSmithKline; Peter F. Lebowitz, GlaxoSmithKline; Peter T.C. Ho, GlaxoSmithKline Honoraria: Anna Maria Storniolo, GlaxoSmithKline; Mark D. Pegram, Genentech, GlaxoSmithKline; Beth Overmoyer, GlaxoSmithKline Research Funding: Anna Maria Storniolo, GlaxoSmithKline; Paula Silverman, GlaxoSmithKline, Genentech Expert Testimony: None Other Remuneration: None

	AUTHOR CONTRIBUTIONS

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION AUTHORS' DISCLOSURES OF... AUTHOR CONTRIBUTIONS Appendix REFERENCES

 
Conception and design: Anna Maria Storniolo, Mark D. Pegram, Suzanne F. Jones, Jill Loftiss, Kevin M. Koch, Elaine Paul, Lini Pandite, Peter T.C. Ho, Howard A. Burris III

Provision of study materials or patients: Anna Maria Storniolo, Mark D. Pegram, Beth Overmoyer, Paula Silverman, Nancy W. Peacock, Suzanne F. Jones, Jill Loftiss, Howard A. Burris III

Collection and assembly of data: Suzanne F. Jones, Jill Loftiss, Nikita Arya, Kevin M. Koch, Elaine Paul, Lini Pandite, Ronald A. Fleming, Peter F. Lebowitz, Howard A. Burris III

Data analysis and interpretation: Anna Maria Storniolo, Beth Overmoyer, Suzanne F. Jones, Jill Loftiss, Nikita Arya, Kevin M. Koch, Elaine Paul, Lini Pandite, Ronald A. Fleming, Peter F. Lebowitz, Howard A. Burris III

Manuscript writing: Anna Maria Storniolo, Suzanne F. Jones, Jill Loftiss, Kevin M. Koch, Lini Pandite, Ronald A. Fleming, Peter F. Lebowitz, Howard A. Burris III

Final approval of manuscript: Anna Maria Storniolo, Mark D. Pegram, Beth Overmoyer, Paula Silverman, Suzanne F. Jones, Jill Loftiss, Nikita Arya, Kevin M. Koch, Elaine Paul, Ronald A. Fleming, Peter F. Lebowitz, Peter T.C. Ho, Howard A. Burris III

	Appendix

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION AUTHORS' DISCLOSURES OF... AUTHOR CONTRIBUTIONS Appendix REFERENCES

 
Pharmacokinetic study.
Once the optimally tolerated regimen (OTR) was determined, additional patients were enrolled at this dose to characterize the pharmacokinetics (PK) of lapatinib and trastuzumab administered alone and in combination (Fig 1). A washout period of 5 weeks was required for patients in the pharmacokinetic group who had received trastuzumab within 5 weeks of study entry. These patients received lapatinib monotherapy during the trastuzumab washout period. Patients were randomly assigned into one of two treatment sequences (Fig 1). Blood samples were collected before dosing and 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 4, 5, 6, 8, 12, and 24 hours after ingestion of the lapatinib dose and/or the initiation of the 0.5-hour trastuzumab infusion. After completion of the PK phase, patients still receiving clinical benefit could remain on study receiving the OTR.

Lapatinib was analyzed by a previously published liquid chromatography with tandem mass spectrometry method with a sensitivity of 5 ng/mL.¹¹ Trastuzumab was analyzed by an enzyme immunoassay method with a sensitivity of 10 µg/mL.

Plasma concentration data were analyzed by standard noncompartmental methods using WINNonlin (Scientific Consultant, Apex, NC) professional software version 4.1 (Pharsight Corp, Mountain View, CA). Analysis of variance of both area under the plasma concentration-time curve and maximum concentration in plasma was performed using a mixed model to estimate a point and 90% CI estimate of the true difference in least squares means between test and reference treatments for each comparison.

	ACKNOWLEDGMENTS

 
We thank the patients who participated in this study and their families.

	NOTES

 
published online ahead of print at www.jco.org on May 19, 2008.

Supported by a grant from GlaxoSmithKline.

Presented in part at the 41st Annual Meeting of the American Society of Clinical Oncology, Orlando, FL, May 13-17, 2005; European Cancer Organization 13 Meeting, Paris, France, October 30 to November 3, 2005; and the 28th Annual San Antonio Breast Cancer Symposium, San Antonio, TX, December 8-11, 2005.

Authors’ disclosures of potential conflicts of interest and author contributions are found at the end of this article.

	REFERENCES

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION AUTHORS' DISCLOSURES OF... AUTHOR CONTRIBUTIONS Appendix REFERENCES

 
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2. Slamon DJ, Clark GM, Wong SG, et al: Human breast cancer: Correlation of relapse and survival with amplification of the HER-2/neu oncogene. Science 235:177-182, 1987

3. Revillion F, Bonneterre J, Peyrat JP: ERBB2 oncogene in human breast cancer and its clinical significance. Eur J Cancer 34:791-808, 1998

4. Alroy I, Yarden Y: Biochemistry of HER2 oncogenesis in breast cancer. Breast Dis 11:31-48, 2000

5. Rusnak DW, Lackey K, Affleck K, et al: The effects of the novel, reversible epidermal growth factor receptor/ErbB-2 tyrosine kinase inhibitor, GW2016, on the growth of human normal and tumor-derived cell lines in vitro and in vivo. Mol Cancer Ther 1:85-94, 2001

6. Gaul MD, Guo Y, Affleck K, et al: Discovery and biological evaluation of potent dual erbB-2/EGFR tyrosine kinase inhibitors: 6-thiazolyquinazolines. Bioorgan Med Chem Lett 13:637-640, 2003

7. Wood ER, Truesdale AT, McDonald OB, et al: A unique structure for epidermal growth factor receptor bound to GW572016 (lapatinib): Relationships among protein conformation, inhibitor off-rate, and receptor activity in tumor cells. Cancer Res 64:6652-6659, 2004

8. Xia W, Mullin RJ, Keith BR, et al: Anti-tumor activity of GW572016: A dual tyrosine kinase inhibitor blocks EGF activation of EGFR/erbB2 and downstream Erk1/2 and AKT pathways. Oncogene 21:6255-6263, 2002

9. Gee J: Chaperone-mediated destruction of erbB2: Relevance to tyrosine kinase inhibitors. Breast Cancer Res 4:205-206, 2002

10. Konecny G, Pegram MD, Venkatesan N, et al: Activity of the dual kinase inhibitor lapatinib against HER-2-overexpressing and trastuzumab-treated breast cancer cells. Cancer Res 66:1630-1639, 2006

11. Hsieh S, Tobien T, Koch K, et al: Increasing throughput of parallel on-line extraction liquid chromatography/electrospray ionization tandem mass spectrometry system for GLP quantitative bioanalysis in drug development. Rapid Commun Mass Spectrom 18:285-292, 2004

12. Cobleigh MA, Vogel CL, Tripathy D, et al: Multinational study of the efficacy and safety of humanized anti-HER2 monoclonal antibody in women who have HER2-overexpressing metastatic breast cancer that has progressed after chemotherapy for metastatic disease. J Clin Oncol 17:2639-2648, 1999

13. Vogel CL, Cobleigh MA, Tripathy D, et al: Efficacy and safety of trastuzumab as a single agent in first-line treatment of HER2-overexpressing metastatic breast cancer. J Clin Oncol 20:719-726, 2002

14. Perez EA, Byrne JA: Results of an analysis of cardiac function in 3127 patients treated with lapatinib. J Clin Oncol 24:23s, 2006 (suppl; abstr 583)

15. Carter P, Presta L, Gorman CM, et al: Humanization of an anti-p185HER2 antibody for human cancer therapy. Proc Natl Acad Sci U S A 89:4285-4289, 1992

16. Lin YL, Clinton GM: A soluble protein related to the HER-2 proto-oncogene product is released from human breast carcinoma cells. Oncogene 6:639-643, 1991

17. Pupa SM, Menard S, Morelli D, et al: The extracellular domain of the c-erbB-2 oncoprotein is released from tumor cells by proteolytic cleavage. Oncogene 8:2917-2923, 1993

18. Christianson TA, Doherty JK, Lin YJ, et al: NH₂-terminally truncated Her-2/neu protein: Relationship with shedding of the extracellular domain and with prognostic factors in breast cancer. Cancer Res 58:5123-5129, 1998

19. Xia W, Liu LH, Ho P, et al: Truncated erbB2 receptor (p95erbB2) is regulated by heregulin through heterodimer formation with erbB3 yet remains sensitive to the dual EGFR/ErbB2 kinase inhibitor GW572016. Oncogene 23:646-653, 2004

20. Saez R, Molina M, Ramsey EE, et al: P95HER-2 predicts worse outcome in patients with HER-2 positive breast cancer. Clin Cancer Res 12:424-430, 2006

21. Molina MA, Saez R, Ramsey EE, et al: NH₂-terminal truncated HER-2 protein but not full-length receptor is associated with nodal metastasis in human breast cancer. Clin Cancer Res 8:347-353, 2002

22. Pusztai L, Esteva FJ: Continued use of trastuzumab (Herceptin) after progression on prior trastuzumab therapy in HER-2-positive metastatic breast cancer. Cancer Invest 24:187-191, 2006

23. Moy B, Goss PE: Lapatinib: Current status and future directions in breast cancer. Oncologist 11:1047-1057, 2006

Submitted August 2, 2007; accepted January 14, 2008.

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