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© 2001 American Society for Clinical Oncology Specific Targeting, Biodistribution, and Lack of Immunogenicity of Chimeric Anti-GD3 Monoclonal Antibody KM871 in Patients With Metastatic Melanoma: Results of a Phase I TrialByFrom the Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch, and Departments of Nuclear Medicine and Centre for Positron Emission Tomography, Surgery, and Anatomical Pathology, Austin and Repatriation Medical Centre, Melbourne, Australia; Kyowa Hakko Kogyo Co Ltd, Tokyo, Japan; Radioimmune and Inorganic Chemistry Section, National Cancer Institute, National Institutes of Health, Bethesda, MD; and Ludwig Institute for Cancer Research, New York, NY. Address reprint requests to A.M. Scott, MBBS, Tumour Targeting Program, Ludwig Institute for Cancer Research, Level 1, Harold Stokes Building, Austin and Repatriation Medical Centre, Studley Rd, Heidelberg, Victoria 3084, Australia; email: ams{at}austin.unimelb.edu.au
PURPOSE: KM871 is a chimeric monoclonal antibody against the ganglioside antigen GD3, which is highly expressed on melanoma cells. We conducted an open-label, dose escalation phase I trial of KM871 in patients with metastatic melanoma. PATIENTS AND METHODS: Seventeen patients were entered onto one of five dose levels (1, 5, 10, 20, and 40 mg/m2). Patients received three infusions of KM871 at 2-week intervals, with the first infusion of KM871 trace-labeled with indium-111 (111In) to enable assessment of biodistribution in vivo. Biopsies of metastatic melanoma sites were performed on days 7 to 10. RESULTS: Fifteen of 17 patients completed a cycle of three infusions of KM871. No dose-limiting toxicity was observed during the trial; the maximum-tolerated dose was therefore not reached. Three patients (at the 1-, 5-, and 40-mg/m2 dose levels) developed pain and/or erythema at tumor sites consistent with an inflammatory response. No normal tissue uptake of 111In-KM871 was observed, and tumor uptake of 111In-KM871 was observed in all lesions greater than 1.5 cm (tumor biopsy 111KM871 uptake results: range, 0.001% to 0.026% injected dose/g). The ratio of maximum tumor to normal tissue was 15:1. Pharmacokinetic analysis revealed a 111In-KM871 terminal half-life of 7.68 ± 2.94 days. One patient had a clinical partial response that lasted 11 months. There was no serologic evidence of human antichimeric antibody in any patient, including one patient who received 16 infusions over a 12-month period. CONCLUSION: This study is the first to demonstrate the biodistribution and specific targeting of an anti-GD3 antibody to metastatic melanoma in patients. The long half-life and lack of immunogenicity of KM871 makes this antibody an attractive potential therapy for patients with metastatic melanoma.
THE DEVELOPMENT of antibody-based therapies for metastatic melanoma has been the focus of considerable interest for many years. The high expression of gangliosides on melanoma cells has led to this antigen system being selected for targeting approaches, although the expression of gangliosides in some normal tissues is well documented.1-6 The murine monoclonal antibody (mAb) R24, which targets the ganglioside antigen GD3, has been the most important antibody studied to date in this field.7 An initial phase I clinical trial with mAb R24 in patients with advanced melanoma showed marked inflammatory reactions at tumor sites in some patients with cutaneous disease, major responses in three out of 12 treated patients, and minor responses in an additional two patients.8,9 Although additional trials of mAb R24 therapy alone,10-12 R24 in combination with cytokines,13,14 and R24 and chemotherapeutic agents15 have been conducted, assessment of the potential therapeutic value of R24 has been limited because of the strong immunogenicity of R24 in humans, accompanied by the development of high titers of antimouse antibodies in treated patients. In addition, the biodistribution of R24 throughout the body or uptake in all tumor sites could not be assessed because of the inability to trace-label R24 with iodine-131 or indium-111 (111In). Thus the true targeting potential of this antibody has not been fully established. In order to target the GD3 antigen in metastatic melanoma and reduce potential immunogenicity in patients, chimeric immunoglobulin (Ig) G1 anti-GD3 mAb KM871 was developed.16 KM871 has potent immune effector function, including complement-dependent cytotoxicity, and antibody-dependent cellular cytotoxicity, and it has minimal difference in affinity or effector function compared with the parent murine IgG3 antibody, KM641.17,18 KM871 has also been shown to have a substantial antitumor effect in animal models.16,18 We have successfully labeled KM871 with 111In, with retention of binding affinity, and demonstrated successful targeting of GD3-expressing xenografts in a nude mouse model.19 In order to determine KM871’s ability to target melanoma, define its immunogenicity, and evaluate for the first time in humans the biodistribution of an anti-GD3 antibody, we conducted a phase I trial of KM871 in patients with metastatic melanoma. The results of this trial are reported here.
KM871 Production The anti-GD3 chimeric mAb KM871 was constructed from the murine anti-GD3 KM641 mAb, as previously described.16,17 A master cell bank and manufacturer’s working cell bank were established from the YB2/0 cells expressing KM871, and production of clinical grade antibody was performed by Kyowa Hakko Kogyo Co Ltd, Tokyo, Japan. Downstream processing and concentration, formulation, and packaging into vials of clinical-grade KM871 was performed by Kyowa Hakko Kogyo and in the Biologic Production Facility of the Ludwig Institute for Cancer Research, Melbourne, Australia. Biosafety testing of the master cell bank, bulk cell culture supernatant, and the purified and final vial product was performed by an accredited biosafety testing company. Quality control testing, including physicochemical characterization, affinity assays, and measurement of immune effector function, was performed on the final vial product before use in the clinical trial.
Trial Design Three infusions of KM871 were administered at 2-week intervals. Each infusion was administered in 100 mL of 5% human serum albumin/normal saline over a 1-hour period. Before each infusion, a 0.1-µg subcutaneous dose of KM871 was administered, and the patient was observed for 1 hour. If no clinical symptoms were observed and no evidence of immediate-type hypersensitivity reaction was seen, the intravenous infusion was administered. Five dose levels of KM871 were studied, 1, 5, 10, 20, and 40 mg/m2. Three patients were entered at each dose level for evaluation of toxicity, and dose escalation was performed only after all three patients had completed three infusions and had reached day 28 with no evidence of dose-limiting toxicity. Standard common toxicity criteria ([CTC] version 2.0) were used to evaluate toxicity. For each patient, the first infusion of KM871 was trace-labeled with 111In in order to allow determination of the biodistribution of KM871 and to facilitate pharmacokinetic and biopsy analyses. Whole-body gamma camera imaging was performed on the day of infusion (day 0), day 1, and on at least three further occasions up to day 7 after infusion of 111In-KM871. All patients were observed for 4 hours after each infusion for possible symptoms. Blood samples for pharmacokinetics were obtained before each infusion, immediately afterward, at 15, 30, 45, and 60 minutes after infusion, and at 2 and 4 hours after infusion. Further blood samples were obtained on three to four occasions over 1 week after each infusion of KM871. On days 7 to 10 after the first infusion of KM871, biopsies of sites of known disease were performed to allow assessment of ganglioside expression, localization of 111In-KM871 to tumor, and histologic evaluation. Patients were restaged on day 56. Patients with tumor responses or stable disease at restaging were eligible to receive further cycles of KM871 provided that they had recovered from any toxicity.
Patient Eligibility The clinical protocol and informed consent forms were approved by the Austin and Repatriation Medical Centre Human Research Ethics Committee. Informed consent was obtained from each subject.
Radiolabeling of KM871 The immunoreactivity of radiolabeled antibody KM871 was determined using the Lindmo assay using 20 ng of labeled antibodies and SK-MEL-28 as target cells.21 To demonstrate specificity of binding, 20 µg of unlabeled KM871 and class-matched control antibody KM966 (Kyowa Hakko Kogyo) were added to the assays and the extent of binding was similarly determined. The stability of radioconjugates after radiolabeling was determined by incubation in serum at 37°C and analysis with instant thin-layer chromatography on silica gel. Retention of immunoreactivity was also determined. These evaluations were repeated on radiolabeled antibody derived from blood samples taken from patients.
Gamma Camera Imaging
Pharmacokinetics Pharmacokinetic calculations were performed of serum data using a curve-fitting program (WinNonlin; Pharsight Co, Mountain View, CA). A two-compartment model was used to calculate pharmacokinetic results. Enzyme-linked immunosorbent assay. Unlabeled KM871 in patient serum was measured using an enzyme-linked immunosorbent assay (ELISA). Purified GD3 antigen (Alexis Corp, Lausen, Switzerland) was immobilized onto microtiter plates (ImmunoPlate Maxisorp; Nunc, Roskilde, Denmark), and blocked with 5% fetal calf serum (Trace Biosciences, Sydney, Australia) in PBS. Standards of KM871 and an IgG1 control antibody (KM966) were used in the assays. Serum samples from patients were diluted with PBS/1% bovine serum albumin (Sigma), standards, and control antibody, and 100-µL samples were added in triplicate to the GD3-coated wells of the microtiter plates, incubated for 1 hour at room temperature, and washed. Goat antihuman IgG horseradish peroxidase–conjugated secondary antibody (Sigma) was then added, incubated for 1 hour, and washed, and then 2,2-azino-bis(3-ethylbenzo-thiazoline 6-sulphonic acid) chromogen substrate (Sigma) was added. Optical density readouts of samples were measured at 405 nm by a spectrophotometer (Milenia Kinetic Analyzer; Diagnostic Products Corp, Gwynedd, United Kingdom). The reference optical density was determined from the linear portion of the standard curve, and the KM871 concentration of serum samples was determined from the reference titer. The lower limit of quantitation with this validated method was determined to be 100 ng/mL.
Biopsy GD3 expression in tumor was evaluated on frozen sections from biopsy specimens. Sections were fixed in acetone and washed. Biotin/avidin was added, and sections were washed again, and then the mouse anti-GD3 antibody KM641 (Kyowa Hakko Kogyo) was added. After further washing, a biotinylated secondary antibody (Dako LSAB Kit; Dako, Glostrup, Denmark) was added, followed by washing, addition of streptavidin–horseradish peroxidase (Dako LSAB Kit), further washing, and then addition of the chromogen substrate 3-amino-9-ethylcarbazole (Sigma). A murine IgG3 antibody (Southern Biotechnology Associates, Birmingham, AL) was used as a negative control, and a GD3-positive control melanoma tissue specimen was used for all analyses. Sections were evaluated for GD3 antigen expression on tumor cells by an experienced anatomic pathologist, and the results were expressed according to the criteria of less than 10%, 10% to 25%, 25% to 50%, 50% to 75%, 75% to 90%, and more than 90% of tumor cells in the biopsy section expressing GD3 antigen. Sections of tumor were also obtained for quantitation of 111In-KM871 localization. Tumor samples were weighed and then counted in a gamma scintillation counter (Packard), along with a known standard of 111In. The uptake of 111In-KM871 was calculated based on the tumor and standard counts and expressed as percent injected dose per gram (% ID/g) of tissue. Normal tissue (eg, fat) from the biopsy sample, where available, was also obtained, weighed, and counted, and the uptake of 111In-KM871 in normal tissue was then calculated.
Human Antichimeric Antibody Prestudy serum samples were used as negative controls for each analysis of each patient sample. A serum sample was considered positive for human antichimeric antibody (HACA) if the binding of a serum component to immobilized KM871, measured in response units, increased more than two-fold over binding measured in prestudy serum samples from the same patient, provided that such an increase was not observed with an isotype-matched control channel. If the control channel also showed such an increase, this was considered to be due to nonspecific binding.
Statistical Analysis
Patients A total of 17 patients (11 men and six women) entered the clinical trial. The mean age was 49.6 years (mean age for men, 52.2 years; women, 44.8 years). The patient demographics, including prior treatment and sites of disease on study entry, are detailed in Table 1.
All patients received an initial infusion of KM871 trace-labeled with 111In. No reactions were observed after the subcutaneous dose of KM871 in any patient in the trial. Two patients developed progressive disease between the first and second infusions and were taken off study before receiving the second infusion. A total of 15 patients completed one cycle of KM871 (three infusions) and were restaged. Three patients went on to further cycles: two patients completed two cycles (a total of six infusions of KM871), and one patient completed five cycles and was in the sixth cycle when she developed disease progression; she was taken off study. This patient (patient no. 4) received a total of 16 infusions of KM871 over a 12-month period. In total, 17 patients received 66 infusions of KM871.
Adverse Events
The most common toxicities observed were rash and urticaria, rigors, fever, and rhinitis, which were observed primarily in the higher dose levels of 10 mg/m2 and above (Table 2). A clinical symptom profile consisting of CTC grade 1 or 2 (one or more symptoms of) rigors, urticaria, and rhinitis was observed in most patients at the 10-mg/m2, 20-mg/m2, and 40-mg/m2 dose levels, but it was controlled with promethazine HCl, dexamethasone, and meperidine. Urticaria and rash were observed in the lower limbs as well as in the trunk and upper limbs, without specific localization in sites of cutaneous or subcutaneous metastatic disease (Fig 1). This symptom profile usually began at the end of the first KM871 infusion, or shortly afterward, and was less marked with subsequent infusions when premedication was given. In one patient, a peripheral oximeter reading showed reduced O2 saturation during a rigor, which resolved when the rigor ceased. The prevalence of this symptom profile and the number of infusions associated with symptoms are listed in Table 4.
Other adverse events related to KM871 included inflammatory reactions at tumor sites (see Induction of Tumor Inflammation, below). No significant changes were observed in blood counts, serum electrolyte levels, or creatinine levels, in any patient.
Radiolabeling of KM871 The radiochemical purity of 111In-KM871 was 97.43% ± 2.35% (mean ± SD) for all labeling, and the specific activity was 2.52 ± 0.74 mCi/mg. The immunoreactivity of 111In-KM871 was 42.28% ± 7.23%. There were no significant differences in any of these parameters between dose levels.
Gamma Camera Imaging
Pharmacokinetics Radiolabeled KM871. The serum clearance of 111In-KM871 by each patient is shown by dose level in Fig 5. The serum clearance data of 111In-KM871 was fitted to a two-compartment model, and calculated values are listed in Table 5. There was no significant difference in T1/2  between dose levels, with a mean T1/2 (± SD) of 0.40 ± 0.33 days. The T1/2ß also showed no significant differences between dose levels, with a mean T1/2ß of 7.68 ± 2.94 days. The only patient who showed a marked difference to patients at all dose levels was patient no. 16. This patient had a fast clearance of 111In-KM871 from blood, and gamma camera images showed accumulation of 111In-KM871 in the abdomen, where ascites was found to be present. This indicated that the distribution of 111In-KM871 into the third space of the abdominal ascites was most likely responsible for the fast clearance. No dose dependence of serum clearance of 111In-KM871 was found.
In the three patients who received more than one cycle of KM871, no significant change in pharmacokinetics of 111In-KM871 was found. Results (mean ± SD) were as follows: for patient no. 3 (two cycles), T1/2 , 0.41 ± 0.35 days and T1/2ß, 7.99 ± 2.91 days; for patient no. 4 (six cycles, five labeled KM871 infusions), T1/2, 0.38 ± 0.30 days and T1/2ß, 10.46 ± 5.60 days; and for patient no. 7, T1/2, 0.33 ± 0.24 days and T1/2ß, 6.82 ± 0.67 days.
ELISA.
The calculated clearance of KM871 measured by ELISA was found to be similar to 111In-KM871 and could also be optimally fitted using a two-compartment model. For the first infusion of KM871, there was no significant difference in T1/2 The peak serum concentrations of KM871 after infusion were dose-dependent, and the serum concentrations at 1 week and 2 weeks after the first infusion of KM871 (trough values just before the second KM871 infusion) are listed in Table 5. Serum concentrations of more than 1 µg/mL at 1 week after infusion of KM871 were observed in all patients at the 5-mg/m2 dose level and higher (except for patient no. 16) and increased at the higher dose levels. Similar values for serum clearance and trough serum concentrations of KM871 were observed after infusions 2 and 3 in each cycle and dose level of KM871 (data not shown).
Biopsy
HACA No patient had detectable HACA after infusion of KM871 at any dose level, with measurements performed up to 3 months after completion of KM871 infusions. This was also consistent with the lack of change in pharmacokinetics of KM871 within each cycle (data not shown). For those patients who received more than one cycle of KM871 (patients no. 3, 4, and 7), no detectable HACA was measured throughout each subsequent cycle. No symptoms of serum sickness were observed in any patient entered onto the study.
Induction of Tumor Inflammation
Tumor Responses Of the 15 assessable patients, one patient had a clinical partial response (patient no. 4) and two patients had stable disease at restaging (patients no. 3 and 7). Patient no. 4 had supraclavicular clinically palpable lymph node disease which became impalpable after the first cycle of KM871; this response was also seen on 18F-fluorodeoxyglucose positron emission tomography scans, but disease was not observed on computed tomography scans due to its location (Fig 4). This response was maintained for 11 months, and disease progression was observed only during cycle 6 of KM871. Three further patients had all measurable or assessable disease removed at the time of biopsy and could not be evaluated for response. The remaining nine patients all had progressive disease on restaging.
This study represents the first reported demonstration of the biodistribution and targeting of an anti-GD3 antibody in patients with metastatic melanoma. At doses up to 40 mg/m2 given every second week, KM871 was well tolerated, with no dose-limiting toxicity, and the maximum-tolerated dose was not reached. A clinical symptom profile of one or more of CTC grade 1 or 2 urticaria, rigors, or rhinitis, occurring during or shortly after KM871 infusion, was observed in most patients at the 10-mg/m2 dose level and higher, but it could be controlled with medication; when patients were premedicated for subsequent KM871 infusions, these symptoms were either less marked or absent. Similar symptoms have been reported after infusion with the murine anti-GD3 antibody R24.8,9,12 The symptoms of capillary leak syndrome, lymphopenia, and serum sickness at high R24 doses (up to 800 to 1,200 mg/m2 over 6 to 8 days) were not seen in our study.10 The cause of this symptom profile seen after KM871 infusion at the higher dose levels is unclear but may be due to binding of KM871 to cell populations such as mast cells or melanocytes in the skin of patients and subsequent cytokine release. Skin biopsies of patients who had these symptoms were not performed in our study, and the precise mechanism of this effect of KM871 remains to be fully defined. The ability to label KM871 with 111In via the bifunctional metal chelate CHX-A''-DTPA was critical in the evaluation of the biodistribution of KM871 in vivo. The labeling efficiency and immunoreactivity of 111In-KM871 was maintained at high levels for each patient infusion at all dose levels. The biodistribution of KM871 was shown to be restricted to blood pool for up to 7 days after infusion, with no normal tissue uptake. This observation would indicate that the expression of GD3 in normal tissues seen in immunohistochemistry studies is not easily accessible to circulating KM871 in vivo. The only exception to this was patient no. 16, who had accumulation of 111In-KM871 in the abdomen due to the presence of ascites; this third-space extravasation was probably due to the protein loss into the abdomen associated with this condition rather than to the abdomen being a normal compartment for KM871 distribution. The specific targeting of sites of metastatic melanoma was also demonstrated, with lesions greater than 1.5 cm identified on planar or single photon emission computed tomography images (Fig 3). Sites of metastatic melanoma were identified as early as 24 to 48 hours after infusion, with metastatic lesions in subcutaneous, lymph node, omental, and liver sites readily identified. Some smaller lesions (< 1.5 cm) were not clearly imaged, usually because of the size of the lesions and the resolution limitations of a gamma camera, although low GD3 expression was observed in a small number of subcutaneous lesions biopsied as part of the trial. The ability to image metastatic sites provides evidence of the high-level and consistent GD3 antigen expression in tumor and demonstrates the rapid targeting potential of KM871 in vivo. Quantitative analysis of uptake in tumor from biopsies performed on days 7 to 10 after infusion of 111In-KM871 was also performed in all patients. The measured uptake of 111In-KM871 in tumor from biopsy samples ranged from 0.001% ID/g to 0.026% ID/g, which represents excellent specific targeting. 111In was selected as the radiolabel for KM871 in view of the poor radioiodine-labeling characteristics of KM871 and the binding affinity and internalization of KM871 in tumor cells after binding to GD3 antigen, which would enhance tumor retention of KM871.19,22-24 Ratios of tumor to normal tissue reached a maximum of 15:1, and the presence of necrosis and amount of viable tumor in the section correlated with these results. The expression of GD3 in tumor biopsy specimens was high. No non–GD3-expressing lesions were seen, although three specimens showed less than 10% tumor cells expressing GD3. These data are consistent with the known expression patterns of GD3 in metastatic melanoma reported in the literature (> 90%).2,5,6 Cellular infiltrates were observed in biopsy specimens, as has been described after R24 infusions,9,12 and the characterization of these infiltrates is currently underway.
The pharmacokinetics of KM871 was measured by gamma counting of serum samples after 111In-KM871 infusion and by an ELISA method. The mean T1/2 of 111In-KM871 was 7.68 days, did not differ significantly between dose levels, and is consistent with other reports of humanized antibodies.25-29 The T1/2 values for KM871 measured by ELISA were slightly longer than those measured by gamma counting of 111In-KM871 but were not significantly different. This may be explained by the number of samples obtained at later time points (up to 2 weeks after infusion) for ELISA measurements, whereas the half-life of 111In restricted later sampling points and probably slightly underestimated the T1/2 results. KM871 serum clearance was not significantly different between dose levels or for subsequent infusions in the same cycle (measured by ELISA). In patients receiving further cycles of KM871, pharmacokinetics also did not significantly change, indicating the stability of KM871 in serum and the reproducibility of these data. Other chimeric antibodies have been reported to have T1/2 ranging from 3 to 10 days, and this would place KM871 as having a serum residence time at the upper end of the reported spectrum. This property would make KM871 ideally suited to maintain high serum concentrations for prolonged periods, allowing the immune effector function mechanisms to have optimal effect. The trough levels of KM871 in serum of more than 1 µg/mL at 1 week after infusion at the 5-mg/m2 and higher dose levels would allow KM871 levels sufficient for complement-dependent cytotoxicity and antibody-dependent cellular cytotoxicity activity to be achieved during this time.16,17 The lack of differences in pharmacokinetics and quantitative tumor uptake between dose levels, and the achievement of target trough levels of KM871 in serum at dose levels The absence of a detectable immune response to KM871 is a major finding of this study. Patients received three infusions of KM871 during a treatment cycle, and in all patients (including two patients receiving six infusions and one receiving 16 infusions over a 12-month period) a negative HACA result was obtained. This finding was also supported by the lack of change in pharmacokinetics within each cycle of KM871. The development of an immune response to an engineered antibody is a variable phenomenon, as humanized (complementarity-determining regions–grafted) antibodies may have minimal immunogenicity25-30 or demonstrate an immune response in a proportion of patients.31 Chimeric antibodies have been reported to elicit immune responses to the antibody after a single infusion at varying frequencies, and the complete lack of an immune response to a chimeric antibody is rare.30 Our findings may be the result of the human sequence homology of KM871 (91%), which approaches that reported for many humanized antibodies. This lack of immunogenicity is of importance in the development of KM871 as a therapeutic agent because it allows the repeated treatment of patients, although confirmation of this finding in a larger series is warranted. The induction of an inflammatory response at tumor sites was observed in three patients in this study. This response was observed in patients at low and high KM871 dose levels, but it was not seen in all metastatic lesions in one patient (patient no. 15). This inflammatory response may be caused by T-cell infiltrates or complement activation at tumor sites; additional analyses of our biopsy specimens are being conducted to evaluate this mechanism further. Prior studies with R24 also reported similar findings in metastatic melanoma lesions, which were also associated in some patients with tumor regression.9,12,14 We observed a partial clinical response in one patient (at the 5-mg/m2 dose level) that lasted for 11 months; however, this study was not designed to evaluate efficacy, and the true response rate of KM871 remains to be established in subsequent studies. Nevertheless, these findings do indicate evidence of a biologic effect of KM871 in metastatic melanoma, and they provide promise for future clinical trials.
Supported by Kyowa Hakko Kogyo Co Ltd, Tokyo, Japan.
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Lee FT, Rigopoulos A, Hall C, et al: Specific localisation, gamma camera imaging and intracellular trafficking of radiolabelled chimeric anti-GD3 ganglioside monoclonal antibody KM871 in SK-MEL-28 melanoma xenografts. Cancer Res 61: 4474-4482, 2001 20. Nikula TK, Curcio MJ, Brechbiel MW, et al: A rapid, single vessel method for the preparation of clinical grade ligand conjugated monoclonal antibodies. Nucl Med Biol 22: 387-390, 1995[Medline] 21. Lindmo T, Boven E, Cuttita F, et al: Determination of the immunoreactive fraction of radiolabelled monoclonal antibodies by linear extrapolation to binding at infinite antigen excess. J Immunol Methods 72: 77-89, 1984[Medline] 22. Mellman I: Endocytosis and antigen processing. Semin Immunol 2: 229-237, 1990[Medline]
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Van Hof AC, Molthoff CFM, Davies Q, et al: Biodistribution of 111indium-labeled engineered human antibody CTMO1 in ovarian cancer patients: Influence of protein dose. Cancer Res 56: 5179-5185, 1996 30. Clark M: Antibody humanization: A case of the "Emperor’s new clothes"? Immunol Today 21: 397-402, 2000[Medline] 31. Welt S, Ritter G, Cohen L, et al: Phase I study of humanized A33 (huA33) antibody in patients with advanced colorectal cancer. Proc Am Soc Clin Oncol 16: 436a, 1997 (abstr 1563) Submitted March 8, 2001; accepted June 4, 2001. This article has been cited by other articles:
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ABSTRACT
INTRODUCTION
70%, no concurrent oral corticosteroid or immune suppressive medication, adequate contraception throughout the study, expected survival of at least 3 months, and ability to provide informed consent. Exclusion criteria included active brain metastases, WBC count less than 2.0 x 109/L, platelet count less than 150 x 109/L; AST and ALT levels more than three times the upper limit of normal, prothrombin time of more than 1.3 times the upper limit of normal, serum creatinine level of more than 0.2 mmol/L, active infection or serious medical condition that would prevent study conduct, and pregnancy or lactation. 
