REVIEW ARTICLE

Int J Clin Oncol (2003) 8:224–229 © The Japan Society of Clinical Oncology 2003DOI 10.1007/s10147-003-0334-8

Yutaka Tokuda

Antibodies as molecular target-based therapy: trastuzumab

Received: May 7, 2003

Y. Tokuda (*)Department of Surgery, Tokai University School of Medicine,Bohseidai, Isehara, Kanagawa 259-1193, JapanTel. �81-463-93-1121; Fax �81-463-95-5941e-mail: tokuda@is.icc.u-tokai.ac.jp

Abstract The HER2 gene is overexpressed or amplified inapproximately 30% of breast cancers. Breast cancer pa-tients with HER2 overexpression or amplification haveshortened disease-free and overall survivals. The HER2protein is thought to be a unique and useful target forantibody therapy of cancers overexpressing the HER2gene. The recombinant humanized anti-HER2 monoclonalantibody, trastuzumab (Herceptin) is now available forclinical use. In a large phase II trial, trastuzumab produceda favorable response rate as a single agent in patients withmetastatic breast cancer who had received one or two priorchemotherapies. Furthermore, a large phase III trialshowed that trastuzumab plus chemotherapy as a first-linetreatment for metastatic breast cancer significantly im-proved response rate, time to disease progression, and over-all survival compared with chemotherapy alone. As a singleagent and in combination with chemotherapy, trastuzumabwas generally well tolerated. Thus, trastuzumab plays animportant role in the treatment strategy of metastatic breastcancer overexpressing HER2 protein. In this article, detailsof this novel biologic agent are reviewed, in conjunctionwith an overview of clinical studies performed in Japan.

Key words HER2/neu · Humanized monoclonal antibody ·Trastuzumab · Breast cancer

Introduction

In 1998, two papers presented at the Annual Meeting ofthe American Society of Clinical Oncology (ASCO) en-couraged patients with breast cancer via the news media.Both papers concerned clinical trials with trastuzumab(Herceptin; Genentech, San Francisco, CA, USA), which isa humanized monoclonal antibody (mAb) against the

HER2/neu oncoprotein (HER2 protein). The day after pre-sentation of the papers, USA Today reported the statementof Dr. L. Norton, Memorial Sloan Kettering Cancer Center,that “it is an opening of a new cancer treatment.” In theautumn of 1998, the FDA approved trastuzumab unusuallyquickly. In this review, some basic facts about and clinicaldata for this humanized antibody against the HER2 proteinare presented.

HER2 protein as a target molecule

The HER2/neu protooncogene (HER2 gene) encodes a re-ceptor-type tyrosine kinase1 corresponding to, but distinctfrom, the epidermal growth factor recetor.2,3 The productsconsist of extracellular, transmembrane, and intracellulardomains with kinase activity and an autophosphorylationprotein on the cell surface. Appreciable amplification and/or overexpression of this gene has been found in breast,ovarian, lung, and gastric cancers.4,5 However, the expres-sion of the gene in normal adult tissues is weak.6,7 There-fore, the HER2 protein is thought to be a useful target forantibody therapy of cancers overexpressing the HER2gene. Several series of murine mAbs directed against theextracellular domain of the HER2 protein have been re-ported to have in vitro and in vivo antitumor effects.8–15

Humanized antibody against the HER2 protein

In the clinical application of murine mAbs, a humanantimouse antibody response during therapy would be amajor limitation.16,17 Carter and colleagues18 constructed ahumanized antibody (rhu4D5; trastuzumab; Herceptin)containing only the antigen-binding loops from a murinemAb against the extracellular domain of the HER2 proteinand human variable region framework residues plus IgG1constant domains. Previously, we characterized the human-ized antibody and compared it with its murine counter-

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part.19 The binding of trastuzumab to the HER2 protein,examined by flow cytometry, was similar to the HER2protein, examined by flow cytometry, was similar to thatshown in the murine model. In vitro, trastuzumab, as well asthe murine mAb, exhibit antiproliferative activity againstHER2-overexpressing human breast carcinoma cells. Thecytotoxicity of human peripheral blood mononuclear cellsagainst a variety of HER2-positive human tumor cell lineswas significantly augmented in the presence of trastuzumab.The antitumor effects of trastuzumab were demonstrated invivo in severe combined immunodeficiency (SCID) micetransplanted with a HER2-positive human gastric cancer.

Clinical trials of trastuzumab as a single agent

Based on previous research findings, clinical trials wereinitiated (Table 1). One of the trials was a single-arm,open-label, multicenter, and multinational study usingtrastuzumab alone, in which 222 patients with HER2-overexpressing metastatic breast cancer were enrolled.20

They had received one or two chemotherapy regimens formetastatic disease. The patients received a loading dose of4mg/kg intravenously, followed by a 2-mg/kg maintenancedoes at weekly intervals. As for patient characteristics, 55%were estrogen receptor-negative, 27% had ten or morepositive lymph nodes at the time of initial diagnosis, 37%had a disease-free interval of less than 12 months, 78% hadmetastatic disease at multiple sites, and 72% had liver orlung metastases. All patients had received extensive previ-ous treatment; 32% had had one prior and 68% had hadtwo or more prior chemotherapy regiments for metastaticdisease, while 26% had undergone high-dose chemotherapywith stem-cell rescue. A blind, independent response eva-luation committee identified 8 complete and 26 partialresponses, for an objective response rate of 15% in theintent-to-treat population (95% confidence interval [CI];11% to 21%). The median duration of response was 9.1months and the median duration of survival was 13 months.The most common adverse effects, seen in about 40% ofpatients, were infusion-associated fever and/or chills thatusually occurred only during the first infusion, dropping toless than 3% during the second or later infusions. The mostclinically significant adverse effect was cardiac dysfunction,

which occurred in 4.7% of the patients. Only 1% of thepatients discontinued the study because of treatment-related adverse effects. The quality-of-life (QOL) score, bythe European Organization for Research and Treatment ofCancer (EORTC) Quality of Life Questionnaire (QLQ)-C30 did not change after treatment.

In Japan, a phase I pharmacokinetic dose escalationstudy was conducted in 18 patients with metastatic breastcancer refractory of chemotherapy.21 Three or 6 patientsat each dosage level received 1, 2, 4, and 8mg/kg oftrastuzumab during 90-min intravenous infusions. The firstdose was followed 3 weeks later by nine weekly doses. Thetarget trough serum concentration had been set at 10µg/ml,based on in vitro observations. A dose of 2mg/kg was con-sidered sufficient for achieving the target trough concentra-tion via the weekly dosing regimen. At a dose of 1mg/kg,1 patient had a grade 3 fever and another had severe, grade3 fatigue. At a dose of 8mg/kg, 1 patient had severe bonepain of grade 3. No antibodies against trastuzumab weredetected in any patients, including 1 individual treated for22 months. Objective tumor responses were observed in2 patients; one who was receiving 4mg/kg had a partialresponse of lung metastases and the other patient, receiv-ing 8mg/kg, had a complete of soft-tissue metastases. Theresponding patient with lung metastases has had prior high-dose chemotherapy with stem-cell rescue. She had under-gone surgical resection of the residual tumors in her leftlung, with a 20-month response duration, which revealedthat the tumor shadows were granulomas. She is now with-out evidence of disease 50 months after surgery.22

Vogel and colleagues23 reported the results of a multi-institutional trial of trastuzumab as first-line therapy for114 patients with HER2-positive metastatic breast cancer(Table 1). They conducted a prospective randomized trialcomparing a 4-mg/kg loading and 2-mg/kg maintenancedose with an 8-mg/kg loading and 4-mg/kg maintenancedose. The response rates of the 2- and 4-mg/kg maintenancedoses were 24% (95% CI, 13%–35%) and 28% (95% CI,16%–40%), respectively. The difference was not statisti-cally significant. Median time to disease progression (TTP)was 3.5 months (95% CI, 3.3–5.1 months) for the 2-mg/kgdose and 3.8 months (95% CI, 2.4–5.5 months) for the 4-mg/kg dose. Median survival for all enrolled patients was 24.4months (95% CI, 16.9–31.7 months). Median survival was22.9 months (95% CI, 16.0–37.1 months) for the 2-mg/kg

Table 1. Clinical trials of single-agent trastuzumab

Author Dose (mg/kg per week) No. Response rate (%); TTP (months);of patients (95% CI) (95% CI)

Cobleigh20 Loading, 4; maintenance, 2 222 15% (11%–21%) 3.1

Vogel23 Loading, 4; maintenance, 2 114 24% (13%–35%) 3.5 (3.3–5.1)Loading, 8; maintenance, 4 28% (16%–40%) 3.8 (2.4–5.5)IHC 3� 84 35IHC 2� 27 0FISH � 79 34FISH � 29 7

CI, confidence interval; TTP, time to disease progression; IHC, immunohistochemistry; FISH,fluorescence in situ hybridization

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dose and 25.8 months (95% CI, 13.3–34.7 months) for the4-mg/kg dose. The response rate of the patients with 3�positivity for HER2 by immunohistochemistry was 35%,although none of the patients with 2� positivity respondedto trastuzumab.

Clinical trials of trastuzumab in combinationwith chemotherapy

Phase III trials

In cell-culture studies, cisplatin, thiotepa, and etoposidewere found to be synergistic with trastuzumab. Additiveinteractions were observed with doxorubicin, paclitaxel,methotrexate, and vinblastine.24 Thus, to determine the effi-cacy and safety of trastuzumab in combination with chemo-therapy as a first-line treatment for patients with HER2-overexpressing metastatic breast cancers, a randomized,placebo-controlled, multicenter, multinational phase IIItrial was designed25 (Table 2). Patients without prioranthracycline treatment were randomly grouped to receivetrastuzumab plus antracycline, (doxorubicin or epirubicin) /cyclophosphamide (AC), or AC alone. Patients with prioranthracycline treatment in the adjuvant setting receivedeither trastuzumab plus paclitaxel, or just paclitaxel. Cyclo-phosphamide (600mg/m2) was administered in combinationwith doxorubicin (80mg/m2) or epirubicin (75mg/m2).Paclitaxel (175mg/m2) was administered by IV infusionover a 3-h period, after premeditation. Chemotherapy wasrepeated every 3 weeks for six cycles. Trastuzumab wasadministered intravenously as a loading dose of 4mg/kgfollowed by a weekly 2-mg/kg maintenance dose. Fourhundred and sixty-nine HER2-overexpressing breastcancer patients without prior chemotherapy for metastaticdisease were enrolled.

The overall response rates were 56% for trastuzumabplus AC and 42% for AC alone (P � 0.02); and 41% fortrastuzumab plus paclitaxel and 17% for paclitaxel alone(P � 0.001). Median TTP was 7.8 months for trastuzumabplus AC and 6.1 months for AC alone (P � 0.01); and6.9 months for trastuzumab plus paclitaxel and 3.0 monthsfor paclitaxel alone (P � 0.001). Median survival was25.1 months for trastuzumab plus chemotherapy and 20.3months for chemotherapy alone (P � 0.046). The QOLscore by EORTC QLQ-C30 was not changed by the addi-tion of trastuzumab.

It is of note that the above survival advantage in thepatients receiving trastuzumab plus chemotherapy was ob-tained although 72% of the patients initially randomized tochemotherapy alone have subsequently received trastu-zumab following disease progression. Thus, it is reasonablethat trastuzumab should be used in the earlier disease state,with greater benefits.

The median survival for patients treated withtrastuzumab plus paclitaxel was 22.1 months, which wassimilar to that of patients treated with first-line trastuzumabmonotherapy, as previously described.23 Furthermore,trastuzumab monotherapy is well tolerated, with goodQOL. Thus, trastuzumab monotherapy represents animportant new option for first-line treatment of HER2-positive metastatic breast cancer, although prospectivestudies are needed.

A prospective study was conducted to comparepaclitaxel and carboplatin to paclitaxel only in combinationwith trastuzumab26 as a first-line treatment of metastaticbreast cancer (Table 2). Paclitaxel, 175mg/m2, with or with-out carboplatin (AUC 6), was administered every 3 weeksin conjunction with trastuzumab. The interim analysisshowed that patients with paclitaxel plus carboplatin hadsignificantly better response rate and TTP.

Phase I/II trials

In terms of combination with paclitaxel, Seidman and col-leagues27 reported the results of weekly paclitaxel (90mg/m2

per h) and weekly trastuzumab (Table 3). Ninety-fivepatients with HER2-positive or -negative metastatic breastcancers were enrolled. The actual dose intensity ofpaclitaxel was 82mg/m2 per week. The most commonnonhematologic toxicity was neuropathy of grade 3 in 28%and grade 4 in 1% of the patients, while 6% of the patientshad grade 3/4 neutropenia. Cardiotoxic effects includedasymptomatic drops in ejection fraction of more than 20%in 7 patients and serious complications in 3 patients. Theoverall response rate was 56.8% (95% CI, 47%–67%), with4.2% achieving a complete response and 52.6% achieving apartial response. Median duration of response was 7 months(95% CI, 2–21 months).

In terms of combination with docetaxel, a phase II studyis ongoing to evaluate the combination of trastuzumab plusdocetaxel, 75mg/m2 every 3 weeks.31 The preliminary re-sponse rate of 16 evaluable patients was 44%. Additionally,this combination regimen was well tolerated. There were

Table 2. Phase III clinical trials of trastuzumab plus chemotherapy

Author Treatment No. of Response Median Medianpatients rate (%) TTP (months) survival (months)

Slamon25 AC 143 56 7.8 26.8AC alone 138 42 6.1 21.4P 92 41 6.9 22.1P alone 96 17 3.0 18.4

Robert26 P � carboplatin 92 52 11.2P 94 36 6.9

AC, doxorubicin or epirubicin � cyclophosphamide; p, paclitaxel

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also several studies investigating the combination oftrastuzumab plus weekly docetaxel.32,33

A phase II study of trastuzumab with vinorelbine wasalso reported34 (Table 3). Forty patients with HER2-overexpressing breast cancers who had received none, one,or two prior chemotherapy regiments for advanced breastcancer were treated with trastuzumab, 4mg/kg loading dosewith 2mg/kg weekly maintenance, and with vinorelbineat 25mg/m2 per week. The vinorelbine dose was adjustedfor weekly neutrophil counts. Fifteen percent of the pati-ents had had prior anthracycline or taxane treatment.Thirty-eight percent had grade 3/4 neutropenia, and nogranulocyte-colony stimulating factor (G-CSF) was used.There were no cases of symptomatic heart failure; 8% ofthe patients had 10%–20% drops in their ejection fractions.The overall response rate was 75%, with a completeresponse rate of 5% and a partial response rate of 70%.Responses were seen despite the extensive prior therapy.A multi-institutional phase II trial using trastuzumab plusvinorelbine as a first-line treatment showed a response rateof 78%.35 The combination was also well tolerated espe-cially in that cardiotoxicity was not shown.

Clinical safety

According to the single-agent phase II trial,20 trastuzumabwas generally well tolerated, with a low incidence of adversereactions. The most common adverse reactions wereinfusion-associated fever of chills, which occurred in about40% of patients. The symptoms could be successfullytreated and did not recur with subsequent administrations.We examined the effect of pretreatment with a nonsteroidalanti-inflammatory drug on such an infusion-associated reac-tion after the loading dose of trastuzumab.38 Of 33 patientswho underwent pretreatment, 6.1% had fever or chills ofgrade 1 or more within 24h after the initiation oftrastuzumab. In contrast, 12.5% of 8 patients who did notundergo pretreatment had an infusion reaction.

The most clinically significant adverse reaction oftrastuzumab was cardiac dysfunction. In the monotherapyphase II trial,20 signs and symptoms of cardiac dysfunction

were identified in 4.7% of the patients. A syndrome ofcardiac dysfunction, similar to that observed with anthra-cyclines, was reported more commonly with trastuzumabplus AC (18% grades 3/4) than with AC alone (3%),paclitaxel alone (0%), or trastuzumab plus paclitaxel(2%).25 To date, the mechanism of the adverse effects oftrastuzumab on cardiac function has not been fully ex-plained. Thus, paclitaxel is recommended in combinationwith trastuzumab. During combination treatment, cardiacfunction should be serially monitored, particularly for thosepatients heavily pretreated with anthracyclines.

One of the most important issues during trastuzumabtreatment was central nervous system (CNS) metastasis.38

Eight of 41 patients manifested CNS metastasis duringtrastuzumab treatment. Six had other metastatic sitesthat responded to the treatment. According to a study re-garding a patient with meningeal carcinomatosis,39 onlyminimal amounts of trastuzumab were found to penetratethe CSF. Therefore, it is unlikely that intravenoustrastuzumab would be useful to treat meningeal or cerebralmetastasis of breast cancer. As a potential treatmentmethod to resolve this issue, the safety and efficacy ofthe intrathecal administration of trastuzumab should bedetermined.

Prediction of treatment effect

As described above, the effect of trastuzumab is greatlydependent on the degree of HER2 overexpression.Seidman and colleagues27 reported the results of a clinicaltrial examining weekly paclitaxel with trastuzumab. Theresponse rate was dramatically different in subsets ofpatients with HER2 overexpression tested by immuno-histochemical assays using different antibodies or byfluorescence in situ hybridization (FISH) (Table 4). For theappropriate use of trastuzumab, an optimal testing strategyshould be established. Evaluation by FISH seemed to be astandard when various methods were examined in regard towhich was a good predictive indicator, as shown in Table 5.As seen in Table 6, immunohistochemistry (IHC) is gener-ally concordant with FISH. There were small numbers of

Table 3. Phase I/II clinical trials of trastuzumab plus chemotherapy

Author Chemotherapy No. of patients Response rate (%)

Seidman27 Paclitaxel 90 mg/m2 per week 95 (HER2� & HER2�) 57Krasna28 Paclitaxel 80 mg/m2 per week 15 67Scholtz29 Paclitaxel 90 mg/m2 per week 24 50Yeung30 Paclitaxel 80 mg/m2 per week 19 27Kuzur31 Docetaxel 75 mg/m2 every 3 weeks 16 44Uber32 Docetaxel 35 mg/m2 per week 19 63Esteva33 Docetaxel 35 mg/m2 per week 30 60Burstein34 Vinorelbine 25 mg/m2 per week 40 75Jahanzeb35 Vinorelbine 30 mg/m2 per week 40 78Bangemann36 Capecitabine 2250 mg/m2 per day for 14 day 13 47

every 3 weeksO’Shaughnessy37 Gemcitabine 1200 mg/m2 on days 1 and 8 38 32

every 3 weeks

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patients who were negative by IHC, but positive by FISH.Conversely, most, but not all IHC 3� patients were FISH-positive. In all three series whose results are shown inTable 6,27,40,41 a substantial fraction of patients who were 2�by IHC lacked FISH positivity. Accordingly, FISH testingwill further define an appropriate patient subset in patientsin whom IHC testing shows a result of 2�.

Future perspectives

Although trastuzumab was approved by the FDA in 1998,approval in Japan was obtained as late as 2001. Antibody

Table 4. Response rates and methods of HER2 testing27

HER2 Tests Positivity (%) Response rate (%)

IHCHercepTest 58 69PAb1 71 67TAB250 40 83CB11 47 76FISH 45 75

PAb1, rabbit antihuman polyclonal Ab (Zymed Laboratories, SouthSan Francisco, CA, USA); TAB250, mouse antihuman mAb (ZymedLaboratories); CB11, mouse antihuman mAb (Ventana Medical Sys-tem, Tucson, AZ, USA)

Table 5. FISH positivity and response to trastuzumab40

Overall response rate

FISH� FISH�

Second/third-line monotherapyIHC 2� (n � 35) 20% 0%IHC 3� (n � 107) 20% 0%IHC 2�/3� (95% CI) 20% (12–28) 0% (0–7)

First-line monotherapyIHC 2�/3� (n � 62) (95% CI) 41% (26–56) 5% (0–14)

First-line combination (IHC 2�/3�)Chemo alone (95% CI) 27% (19–35) 39% (26–52)Chemo � Herceptin (95% CI) 54% (45–63) 41% (27–55)

Chemo, Chemotherapy

Table 6. Concordance between IHC and FISH27,40,41

IHC score No. of patients FISH positivity

HercepTest27

0 38 11%1� 9 22%2� 16 81%3� 25 84%

HercepTest41

0 7 0%1� 37 0%2� 72 31%3� 26 100%

4D5/CB1140

0 214 3%1� 30 7%2� 88 24%3� 197 89%

4D5, Genentech, San Francisco, CA, USA

therapy against the HER2 protein is promising as a newtreatment modality for breast cancer. The very importantissues of when trastuzumab should be considered in thetreatment strategy for metastatic breast cancer and whethertrastuzumab monotherapy or combination therapy shouldbe used will be answered after future clinical trials. Further-more, a number of trials of trastuzumab as adjuvant therapyare now underway.

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