Purpose: This phase I study explored the biodistribution and pharmacokinetics of the immunoconjugate CMD-193 [a humanized anti–Lewis Y (Ley) antibody conjugated with calicheamicin in patients with advanced cancers expressing the Ley antigen.

Experimental Design: The primary objectives were to determine biodistribution and pharmacokinetics of CMD-193. Secondary objectives included response rates and change in tumor metabolism. Patients with progressive, measurable, and Ley positive malignancies were eligible for enrollment in one of two dose cohorts, 1.0 and 2.6 mg/m2. The first cycle was trace labeled with 111In for biodistribution assessment using γ camera imaging. Subsequent cycles were administered every 3 weeks up to a maximum of six cycles, depending on toxicity and response. Pharmacokinetic analysis was based on radioassay and ELISA.

Results: Nine patients were enrolled in the study. Biodistribution images showed initial blood pool activity, followed by markedly increased hepatic uptake by day 2, and fast blood clearance in all patients. There was low uptake in tumor in all patients. The overall T½β of 111In-CMD-193 was 102.88 ± 35.67 hours, with no statistically significant difference between the two dose levels. One patient had a partial metabolic response on 18F-fluorodeoxyglucose-positron emission tomography (18F-FDG PET) after four cycles, but no radiological responses were observed. Myelosuppression and effects on liver function were the most significant adverse effects.

Conclusions: CMD-193 shows rapid blood clearance and increased hepatic uptake compared with prior studies of the parental antibody hu3S193. These results highlight the importance of biodistribution and pharmacodynamic assessment in early phase studies of new biologics to assist in clinical development. (Clin Cancer Res 2009;15(21):6709–15)

Translational Relevance

This article describes a phase I trial of a novel recombinant immunoconjugate against the Lewis-y antigen, expressed on solid tumors. The results of this study showed the impact of toxin conjugation on the biodistribution and tumor uptake of the immunoconjugate, which could not be predicted based on preclinical studies, and which was not evident from pharmacokinetics or adverse event profiles. Alterations in immunoconjugate biodistribution may impact on tumor uptake and efficacy and, without detailed characterization, will ultimately lead to reduced efficacy and futile clinical trials. This trial design, and results, provides unique information on the importance of pharmacodynamic/bioimaging studies early in the clinical development of immunoconjugates in cancer patients.

The concept of antibody-targeted chemotherapy was successfully translated into the clinic in 2000, when gemtuzumab ozogamicin (Mylotarg, also known as CMA-676) was approved by the Food and Drug Administration for relapsed acute myeloid leukemia (1). This CD33-targeted immunoconjugate of calicheamicin has accelerated the investigation of this therapeutic strategy in solid tumors. CMD-193 is one such calicheamicin immunoconjugate that makes use of the same drug-linker combination as that used in Mylotarg, but the antibody to which it is conjugated targets the Lewis Y (Ley) antigen.

Ley (CD174) is a difucosylated tetrasaccharide internalized antigen displayed on both glycolipid and glycoprotein backbones of membrane surface molecules, and is involved in cellular motility and adhesion (2). Ley has restricted normal tissue expression, and is overexpressed in the majority of epithelial carcinomas (40-90%) including breast, ovary, pancreas, prostate, colon, and lung cancers (311).

The immunoconjugate CMD-193 is composed of G193, a humanized monoclonal antibody based on the anti-Ley antibody hu3S193 (12), covalently linked to NAc-γ calicheamicin DMH via an acid-labile AcBut linker with retention of Ley affinity. In prior phase I trials, hu3S193 showed specific targeting of Ley-expressing tumors, restricted normal tissue distribution, a long serum half-life, and lack of immunogenicity (13, 14). Hu3S193 is currently under development in phase II trials as a naked humanized antibody in Ley-expressing tumors. Dose-dependent regression of Ley-expressing human carcinoma xenografts by the immunoconjugate CMD-193 has been shown in preclinical studies, highlighting the potential therapeutic potential of CMD-193 in cancer patients (15). Initial clinical development of CMD-193 involved a dose escalation phase I study in patients with Ley-expressing advanced solid tumors with an expanded preliminary evaluation of efficacy in patients with non-small cell lung carcinoma.

In support of the initial clinical development of CMD-193, we conducted an additional phase I dose escalation study of CMD-193 in patients with advanced solid tumors expressing the Ley antigen. The primary objectives of this trial were to determine the biodistribution and pharmacokinetics of 111In-CMD-193. The tumor uptake of 111In-CMD-193 was based on qualitative and quantitative assessment of biodistribution images and dosimetry. Secondary objectives were to determine tumor response to CMD-193 through changes in tumor 18F-FDG PET metabolism and measurement by Response Evaluation Criteria in Solid Tumors (RECIST) criteria (16).

Patients

Eligible patients were >18 y of age, who had histologically confirmed solid malignancies with ≥20% tumor cells displaying Ley antigen positivity on immunohistochemistry of archived tumor samples (13), and who had progressed following standard therapy. Inclusion criteria included the following: measurable disease, Eastern Cooperative Oncology Group performance status of 0 to 1, and adequate renal, hepatic, and bone marrow function and ability to give informed consent. Exclusion criteria included the following: cancer therapy within 21 d of the first dose of CMD-193, clinically active central nervous system metastases, significant prior allergic reaction to recombinant human or murine proteins, and serious concurrent medical conditions including chronic liver disease.

Trial design

This trial planned to enroll into three dose cohorts: 1.0, 1.7, and 2.6 mg/m2. These dose levels were selected based on the lowest practical dose able to be trace labeled for biodistribution studies (1.0 mg/m2), and the anticipated therapeutic dose based on other calicheamicin conjugates. After a parallel phase I trial determined the maximum tolerated dose of CMD-193 to be 3.6 mg/m2 and therapeutic dose to be 2.6 mg/m2,9

9C. Zacharchuk and D. S. Sonnichsen, unpublished data.

with dose limiting toxicity related to thrombocytopenia and hepatic enzyme changes, a protocol amendment was approved to remove the 1.7 mg/m2 cohort. Following pretreatment assessments, eligible patients received a single infusion of Indium-111–labeled CMD-193 [111In-CMD-193; 3-7 mCi (120-280 MBq)] at a protein dose level of 1.0 or 2.6 mg/m2 over 1 h on day 1 of cycle 1. Subsequent cycles of unlabeled CMD-193 were administered at three weekly intervals up to a maximum of six cycles, subject to tolerability and response.

Radiolabeling of CMD-193

The immunoconjugate CMD-193 (Wyeth Pharmaceuticals, Inc.) was labeled with 111In- (MDS Nordion) via the bifunctional metal ion chelate CHX-A″-diethylenetriaminepentaacetic acid according to methods described previously (12, 17).

Assessments

Biodistribution evaluation was done by whole body γ camera scans on day 1, day 2, day 3 or 4, day 5 or 6, and day 7 or 8 following 111In-CMD-193 infusion. Pharmacokinetic sampling of 111In-CMD-193 was done during cycle 1 on day 1 (preinfusion, 1 and 4 h postinfusion commencement), day 3, day 8, and day 15. Changes in tumor metabolism were evaluated using 18F-FDG-PET, which was done at screening, between days 15 and 21 of cycles 2 and 4, and at study completion. Antitumor response was assessed using RECIST criteria (16), with computed tomography (CT) scans performed at screening, between days 15 and 21 of cycles 2 and 4, and at study completion. Safety evaluation was done weekly throughout the trial with all adverse events documented and graded according to National Cancer Institute Common Terminology Criteria for Adverse Events version 3.0. Causality was determined as “related to study drug” if the event was deemed definitely, probably, or possibly related to the administration of CMD-193 by the investigator.

Biodistribution and whole body clearance

Whole body planar γ camera images were acquired on a dual-headed γ camera (Picker International and Phillips Medical Systems). Single photon emission CT (SPECT) images of a region of the body with known tumor were also obtained on at least one occasion during this period. Biodistribution analysis was done by examination of whole body and SPECT images by experienced nuclear medicine physicians. Comparison to prior studies of parental antibody hu3S193 was also done (13). Whole body and organ clearance of 111In-CMD-193 was calculated from quantitative whole body conjugate view γ camera images obtained at the multiple time points postinfusion using a well-validated method (13, 18, 19).

Pharmacokinetics

Serum obtained from patients following infusion of 111In-CMD-193 was aliquoted and counted in a γ scintillation counter (Packard Instruments). The results were expressed as % injected dose per liter (%ID/L) and μg/mL. Estimates were determined for the following parameters: T½α and T½β (half-lives of the initial and terminal phases of disposition); V1, volume of central compartment; Cmax, maximum serum concentration; AUC, area under the serum concentration curve extrapolated to infinite time; and CL, total serum clearance. A two-compartment i.v. bolus model with macroparameters, no lag time, and first-order elimination (WNL Model 8) was fitted to individual serum 111In-CMD-193 data for each subject using unweighted nonlinear, least squares with WinNonLin version 5.2 (Pharsight Corp). Measured serum levels of CMD-193 were expressed as ng/mL. Serum samples were also obtained for measurement of total and free calicheamicin by a validated ELISA protocols (LLOQ, 2.45 ng/mL).

Tumor metabolic response assessment

Patient preparation for and acquisition of FDG-PET scans were standardized (20, 21). For each FDG-PET performed, the maximum standardized uptake value (SUVmax) corrected for body weight for all target lesions >2cm identified on CT imaging was calculated using region of interest. The region of interest was determined with the aid of the anatomic detail provided by the CT scan. SUVmax for normal lung tissue was taken as the reference region for each patient, which ensured that any SUV changes in tumors could be directly attributed to treatment response or to disease progression (16). Metabolic response was calculated using the target lesion with the greatest baseline SUVmax, and was categorized according to the European Organization for Research and Treatment of Cancer guidelines (20).

Human-anti-human antibody response

Assay of serum for anti–CMD-193 antibodies in human serum was done by Wyeth Pharmaceuticals using a validated ELISA protocol.

Statistical considerations

The independent samples t test was used to compare mean pharmacokinetic and clearance parameters across the two dose cohorts of CMD-193. It was also used to compare results with those obtained with our prior phase I trial using the unconjugated parental antibody hu3S193 (13).

Patient characteristics

Nine patients were eligible and enrolled (six patients, 1.0 mg/m2 cohort; three patients, 2.6 mg/m2 cohort). Baseline patient demographics and disease characteristics are shown in Table 1. All patients had metastatic disease at study entry and many patients had been extensively pretreated, having received one to five lines of prior chemotherapy, monoclonal antibody, or biological agent. Only one patient (patient 102) received all six cycles of CMD-193. Four patients were withdrawn due to progressive disease after two cycles of treatment (patients 101, 105, 106, and 161), and four patients were withdrawn because of toxicity (patients 103, 104, 107, and 108).

Table 1.

Patient characteristics at study entry and study outcomes

Pt IDAge (y)/SexECOG PSPrimary diagnosisSites of metastatic disease at study entryTumor Ley antigen positivityDose cohortNo. of cyclesFDG-PET response (EOS)CT response (EOS)
101 53/F Colon carcinoma Lymph nodes, liver 20-50% PMD PD 
102 49/F Cholangiocarcinoma Retro-pancreatic mass >75% SMD PD 
103 71/M Colon carcinoma Liver 51-75% PMR SD 
104 58/M Gastric carcinoma Liver 20-50% SMD SD 
105 53/M Colon carcinoma Lymph nodes liver, lung, presacrum 20-50% PMD PD 
106 46/M Gastro-osophageal junction carcinoma Lymph nodes, liver 51-75% PMD PD 
107 77/M Bronchoalveolar carcinoma Lung 20-50% PMD SD 
108 69/F Gastric carcinoma Lymph nodes, liver, lung 20-50% SMD SD 
161 46/F Pulmonary adenocarcinoma Lymph nodes liver, bone 20-50% 2* N/A PD* 
Pt IDAge (y)/SexECOG PSPrimary diagnosisSites of metastatic disease at study entryTumor Ley antigen positivityDose cohortNo. of cyclesFDG-PET response (EOS)CT response (EOS)
101 53/F Colon carcinoma Lymph nodes, liver 20-50% PMD PD 
102 49/F Cholangiocarcinoma Retro-pancreatic mass >75% SMD PD 
103 71/M Colon carcinoma Liver 51-75% PMR SD 
104 58/M Gastric carcinoma Liver 20-50% SMD SD 
105 53/M Colon carcinoma Lymph nodes liver, lung, presacrum 20-50% PMD PD 
106 46/M Gastro-osophageal junction carcinoma Lymph nodes, liver 51-75% PMD PD 
107 77/M Bronchoalveolar carcinoma Lung 20-50% PMD SD 
108 69/F Gastric carcinoma Lymph nodes, liver, lung 20-50% SMD SD 
161 46/F Pulmonary adenocarcinoma Lymph nodes liver, bone 20-50% 2* N/A PD* 

Abbreviations: ECOG, Eastern Cooperative Oncology Group; PS, performance status; EOS, end of study assessment; N/A, not assessed.

*Withdrew prematurely cycle 2 day 9. Response assessed clinically, as patient died of progressive disease before repeat CT scan.

Biodistribution

Evaluation of γ camera imaging following infusion of 111In-CMD-93 showed rapid clearing of blood pool activity, followed by markedly increased hepatic uptake by day 2, persisting to day 8 (Fig. 1). This pattern was observed for all patients in both dose levels. No significant uptake of 111In-CMD-193 in tumor was visualized in target lesions for all patients.

Fig. 1.

Representative biodistribution pattern of 111In-CMD-193. Anterior whole body γ camera images in patient 106 (1.0 mg/m2 dose cohort) following infusion are shown for day 1 (A), day 3 (B), and day 8 (C). Following infusion of 111In-CMD-193, there was initial blood pooling, followed by markedly increased hepatic uptake by day 2 that persisted to day 8. No tumor uptake was apparent in the whole body γ camera images (arrow) or SPECT (D). E, corresponding CT scan shows the large hepatic metastasis, also evident in F, coregistered SPECT/CT scan.

Fig. 1.

Representative biodistribution pattern of 111In-CMD-193. Anterior whole body γ camera images in patient 106 (1.0 mg/m2 dose cohort) following infusion are shown for day 1 (A), day 3 (B), and day 8 (C). Following infusion of 111In-CMD-193, there was initial blood pooling, followed by markedly increased hepatic uptake by day 2 that persisted to day 8. No tumor uptake was apparent in the whole body γ camera images (arrow) or SPECT (D). E, corresponding CT scan shows the large hepatic metastasis, also evident in F, coregistered SPECT/CT scan.

Close modal

Whole body clearance was (mean ± SD) 47.82 ± 3.24 hours, and there was no statistically significant difference between dose levels (P = 0.74). Quantitative analysis confirmed the high levels of hepatic uptake of 111In-CMD-193 apparent visually on biodistribution images. Compared with the parental hu3S193 antibody, hepatic uptake was significantly higher at 24 hours (time of maximal hepatic uptake; Table 2). Tumor uptake was also significantly lower for CMD-193 compared with hu3S193 (Table 2).

Table 2.

Peak tumor uptake and liver uptake of CMD-193 compared with parental antibody hu3S193

hu3S193CMD193
Mean ± SDRangeMean ± SDRange
Peak tumor uptake (μg/gm) 2.9 ± 1.7 1.2-6.3 0.15 ± 0.03* 0.11-0.19 
Liver %ID 24 p.i. 7.3 ± 1.5 4.5-9.5 33 ± 6.0 26.9-42.0 
hu3S193CMD193
Mean ± SDRangeMean ± SDRange
Peak tumor uptake (μg/gm) 2.9 ± 1.7 1.2-6.3 0.15 ± 0.03* 0.11-0.19 
Liver %ID 24 p.i. 7.3 ± 1.5 4.5-9.5 33 ± 6.0 26.9-42.0 

*Peak tumor uptake hu3S193 vs CMD-193, P = 0.0016.

Maximal liver uptake hu3S193 vs CMD-193, P < 0.0001.

Pharmacokinetics and HAHA

CMD-193 displayed a fast clearance from blood, consistent with the biodistribution findings (Table 3). Selected serum samples from patients were analyzed by fast protein liquid chromatography for immune complex or metabolite formation, and no complexes or metabolites or free 111In-chelate was observed up to 72 hours postinfusion (data not shown). No significant differences were found for T½α, T½β, clearance, or V1 between the two dose levels. The results for CMD-193 were significantly different compared with the parental antibody hu3S193 (13), with T½β for CMD-193 102.88 ± 35.67 hours versus 189.63 ± 62.17 hours for hu3S193 (P < 0.001) and CL for CMD-193 113.22 ± 56.58 mL/h versus 22.09 ± 9.87 mL/h for hu3S193, P < 0.001. No HAHA was detected in any patient. Free calicheamicin levels were at or below the limit of assay quantitation in all patients.

Table 3.

111In-CMD-193 serum pharmacokinetic analysis results

ParameterUnitsCMD-193 All (n = 9)1 mg/m2 CMD-193 (n = 6)2.6 mg/m2 CMD-193 (n = 3)t test (comparing two dose levels)
MeanSDMeanSDMeanSDP
T½αHour 4.76 2.15 5.47 1.99 3.32 2.00 0.17 
T½β Hour 102.88 35.67 104.42 37.94 99.79 38.32 0.87 
V1 mL 4,071.22 731.41 4,366.18 586.87 3,481.31 704.13 0.08 
CL mL/h 113.22 56.58 130.04 61.25 79.56 29.67 0.23 
AUC μg.h/mL 29.93 22.31 16.37 6.13 56.45 17.05 Not done 
ParameterUnitsCMD-193 All (n = 9)1 mg/m2 CMD-193 (n = 6)2.6 mg/m2 CMD-193 (n = 3)t test (comparing two dose levels)
MeanSDMeanSDMeanSDP
T½αHour 4.76 2.15 5.47 1.99 3.32 2.00 0.17 
T½β Hour 102.88 35.67 104.42 37.94 99.79 38.32 0.87 
V1 mL 4,071.22 731.41 4,366.18 586.87 3,481.31 704.13 0.08 
CL mL/h 113.22 56.58 130.04 61.25 79.56 29.67 0.23 
AUC μg.h/mL 29.93 22.31 16.37 6.13 56.45 17.05 Not done 

Response assessment

Metabolic response by FDG-PET and antitumor response according to RECIST criteria results are shown in Table 1. Response could not be formally measured in patient 161, who died of rapid progressive disease following early study withdrawal during cycle 2. Patient 103 showed a partial metabolic response (PMR), with a 41.7% reduction in SUVmax (prestudy, 10.8; restaging, 6.3) after four cycles of CMD-193 (Fig. 2). This was despite no change in CT dimensions of the target lesion. Stable metabolic disease (SMD) was observed in three patients and four had progressive metabolic disease (PMD) at end of study assessment. Patient 108 showed SMD, but had a 25% reduction in SUVmax (prestudy, 4.4; restaging, 3.3) after five cycles of treatment. Of the four patients with PMD, three were in dose cohort 1 and one was in dose cohort 2. Assessment of antitumor response by CT scanning showed four patients with stable disease (SD) and four with progressive disease (PD) at final staging.

Fig. 2.

PMR to CMD-193: assessment by 18F-FDG-PET. According to RECIST criteria, patient 103 had stable disease following four cycles of CMD-193 at a dose of 1.0 mg/m2, with a large liver lesion (arrow) remaining similar in size on CT. The patient did however show a PMR in this lesion, with a 41.7% reduction in SUVmax observed on 18F-FDG-PET. Prestudy and post–CMD-193 cycle 4 imaging is shown: A and D, CT image; B and E,18F-FDG-PET; C and F, fused PET/CT images.

Fig. 2.

PMR to CMD-193: assessment by 18F-FDG-PET. According to RECIST criteria, patient 103 had stable disease following four cycles of CMD-193 at a dose of 1.0 mg/m2, with a large liver lesion (arrow) remaining similar in size on CT. The patient did however show a PMR in this lesion, with a 41.7% reduction in SUVmax observed on 18F-FDG-PET. Prestudy and post–CMD-193 cycle 4 imaging is shown: A and D, CT image; B and E,18F-FDG-PET; C and F, fused PET/CT images.

Close modal

Adverse events

CMD-193 at doses of 1.0 and 2.6 mg/m2 was reasonably well tolerated, and no difference in toxicity was observed between patients in the two dose levels (Table 4). Four patients, two from each dose cohort, were withdrawn because of toxicity. The main adverse events with some relationship to CMD-193 were asymptomatic myelosuppression, particularly thrombocytopenia (12 grade 1-2 events, 2 grade 3 events, and 1 grade 4 event), and abnormal liver function (5 grade 3 events). All other related adverse events, including fatigue, lethargy, anorexia, and nausea, were mild (grade 1-2). There were no infusion-related reactions. There were no serious adverse events related to study drug, and no serious or severe unexpected toxicities were observed. Toxicity was consistent with that found in the parallel phase I trial of CMD-193.9

Table 4.

CMD-193 related adverse events (possibly, probably, or definitely related)

System organ class Adverse eventG1G2G3G4Total
Blood and lymphatic system disorders Anemia 
Leucopenia 
Lymphopenia 
Neutropenia 
Thrombocytopenia 11 15 
Gastrointestinal disorders Abdominal bloating 
Epigastric discomfort 
Gastresophageal reflux 
Nausea 10 
Vomiting 
General disorders and administration site conditions Flu-like symptoms 
Flushing 
Fatigue 
Lethargy 
Hepatobiliary disorders Hyperbilirubinaemia 
Investigations ALP increased 
ALT increased 
Amylase increased 
AST increased 
GGT increased 
Lipase increased 
Weight loss 
Metabolism and nutrition disorders Anorexia 
Respiratory, thoracic and mediastinal disorders Epistaxis 
Skin and subcutaneous tissue disorders Bruising 
Rash erythematous 
Total  68 28 105 
System organ class Adverse eventG1G2G3G4Total
Blood and lymphatic system disorders Anemia 
Leucopenia 
Lymphopenia 
Neutropenia 
Thrombocytopenia 11 15 
Gastrointestinal disorders Abdominal bloating 
Epigastric discomfort 
Gastresophageal reflux 
Nausea 10 
Vomiting 
General disorders and administration site conditions Flu-like symptoms 
Flushing 
Fatigue 
Lethargy 
Hepatobiliary disorders Hyperbilirubinaemia 
Investigations ALP increased 
ALT increased 
Amylase increased 
AST increased 
GGT increased 
Lipase increased 
Weight loss 
Metabolism and nutrition disorders Anorexia 
Respiratory, thoracic and mediastinal disorders Epistaxis 
Skin and subcutaneous tissue disorders Bruising 
Rash erythematous 
Total  68 28 105 

Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase.

This study showed lack of targeting of CMD-193 to known sites of metastatic disease, and marked hepatic uptake and rapid clearance from blood, consistent with the observed short T1/2 β and fast serum clearance. There were no documented objective responses seen in size of tumor, but one patient did display a PMR according to 18F-FDG-PET analysis. These bioimaging and pharmacokinetic results highlight the importance of detailed investigation of the properties of antibodies and immunoconjugates in early phase I trials, and can provide critical information impacting on subsequent clinical development.

The biodistribution, clearance, and pharmacokinetic properties of CMD-193 were found to be significantly different to the parental antibody hu3S193. Phase I studies of 111In-hu3S193 have shown prominent specific uptake in tumor, a lack of consistent normal tissue/organ uptake, and a long half-life in blood (13, 14). This is in contrast to the fast clearance from blood, rapid uptake in liver parenchyma, and lack of tumor uptake of 111In-CMD-193 observed in this study. Importantly, one patient (patient 103) participated in both clinical studies, allowing direct comparison of biodistribution, clearance, and hepatic uptake between CMD-193 and hu3S193 in the same patient (Fig. 3). The whole body clearance and terminal half-life of 111In-CMD-193 were also faster than that observed with other humanized IgG1 antibodies (19, 22). Interestingly, this difference in serum clearance of a toxin-conjugate, compared with the parental antibody (hu3S193), has also been recently reported for the Herceptin-maytansinoid conjugate T-DM1, which had a T½ of 2.1 to 3.7 days (compared with Herceptin T½ of >10 days), although biodistribution data were not reported for this conjugate (23, 24).

Fig. 3.

111In-CMD-193 biodistribution pattern compared with the biodistribution of the parental anti-Ley antibody 111In-hu3S193 observed in the prior phase I study13. Patient 103 had participated in both clinical studies, allowing direct comparison of biodistribution, clearance, and hepatic uptake between CMD-193 and hu3S193 in the same patient. A, 1 d; B, 7 d postinfusion of 111In-CMD-193 (arrow, tumor in liver); C, day 0; D, 2 d; and E, 7 d postinfusion of 111In-hu3S193. Note that in the original hu3S193 trial, a small tumor in the liver was visualized on SPECT imaging.

Fig. 3.

111In-CMD-193 biodistribution pattern compared with the biodistribution of the parental anti-Ley antibody 111In-hu3S193 observed in the prior phase I study13. Patient 103 had participated in both clinical studies, allowing direct comparison of biodistribution, clearance, and hepatic uptake between CMD-193 and hu3S193 in the same patient. A, 1 d; B, 7 d postinfusion of 111In-CMD-193 (arrow, tumor in liver); C, day 0; D, 2 d; and E, 7 d postinfusion of 111In-hu3S193. Note that in the original hu3S193 trial, a small tumor in the liver was visualized on SPECT imaging.

Close modal

The rapid clearance of CMD-193 from blood, and the liver uptake observed, was not predicted by preclinical studies. The marked difference in biodistribution of CMD-193 compared with parental antibody hu3S193 cannot be explained by antibody specificity, as retention of Ley binding by CMD-193 was confirmed before infusion. The lack of CMD-193 complexes or metabolites in blood (measured by fast protein liquid chromatography), absence of HAHA, and the lack of prominent spleen or bone marrow uptake on imaging, excludes CMD-193 complexes or free 111In-contributing to the increased liver uptake. It is possible that a physicochemical change induced by conjugation of the antibody with calicheamicin may have led to the altered biodistribution observed in this study. Size and charge are known to influence the uptake of circulating macromolecules by hepatic cells, possibly by influencing electrostatic attraction and hydrophobic interactions or specific receptor–mediated interactions with scavenger receptors, which remove acidic macromolecules (2426). Despite the low uptake of CMD-193 in tumor, the observation of a PMR in one patient highlights the potent biological effect of calicheamicin, even at low tumor concentrations.

To our knowledge, CMB-401 (hCTM01-calicheamicin) is the only calicheamicin immunoconjugate to have reached phase II trials in solid tumors. This combined N-acetyl/analogue of calicheamicin with a polymorphic epithelial mucin targeting humanized antibody hCTM01 using an amide-based linkage. An initial phase I study in patients with epithelial ovarian cancer, which included a predose of unconjugated antibody to minimize uptake in normal tissues and complex formation with circulating antigen, found it to be tolerable and defined the maximum tolerated dose (27). A subsequent phase II trial in 21 patients with recurrent ovarian cancer failed to show clinical efficacy (28). In this case, a lack of efficacy (and suspension of development) was attributed to instability of the amide linker, although free calicheamicin and measurement of serum complex formation results were not published. Biodistribution and pharmacokinetic assessment of CMB-401 in patients were also not done in this trial, and hence, it is not possible to draw direct comparisons with CMD-193 other than the common toxicity profile relating to calicheamicin (29, 30).

CMD-193 showed a similar toxicity profile to Mylotarg and CMB-401, with predominant hepatic and hematological toxicity. Hepatic toxicity seen in some patients following administration of CMD-193 may be explained by hepatic uptake and metabolism of the NAc-γ calicheamicin DMH, as Ley is not expressed by liver cells (4). With Mylotarg however, hepatotoxicity can be explained partly by sinusoidal obstruction syndrome, the mechanism of which probably involves targeting of CD33+ cells in the sinusoids of the liver (30). Although liver toxicity was also documented in phase I study of CMB-401 (but interestingly not mentioned in phase II trial), in this case, it was attributed to expression of target antigen in liver bile duct cells, rather than to uptake of calicheamicin metabolites by hepatocytes (27, 28). Myelosuppression following CMD-193 may be explained by the myelosuppressive effects of a small amount of free calicheamicin.

In summary, the detailed biodistribution and pharmacokinetic assessment performed in this trial was able to identify an unexpected in vivo fate for the novel Ley-targeting immunoconjugate CMD-193 in patients with advanced Ley-positive epithelial cancers. Although CMD-193 was generally tolerable, and hints of biological activity were shown, the marked hepatic uptake, low tumor uptake, and rapid blood clearance observed was in contrast to the parental antibody hu3S193. On the basis of clinical trial data and the biodistribution results shown in this study, the clinical development of CMD-193 has not been continued. These results highlight the importance of detailed biodistribution and pharmacodynamic assessment in early phase studies of new biologics to inform and guide clinical development.

D.S. Sonnichsen, employment, Wyeth Research. Funding for this study was provided by Wyeth Research.

1
Bross
P
,
Beitz
J
,
Chen
G
, et al
. 
Approval summary: gemtuzumab ozogamicin in relapsed acute myeloid leukemia
.
Clin Cancer Res
2001
;
7
:
1490
6
.
2
Dettke
M
,
Palfi
G
,
Loibner
H
. 
Activation-dependent expression of the blood group-related lewis Y antigen on peripheral blood granulocytes
.
J Leukoc Biol
2000
;
68
:
511
4
.
3
Zhang
S
,
Zhang
HS
,
Cordon-Cardo
C
, et al
. 
Selection of tumor antigens as targets for immune attack using immunohistochemistry: II. Blood group-related antigens
.
Int J Cancer
1997
;
73
:
50
6
.
4
Kitamura
K
,
Stockert
E
,
Garin-Chesa
P
, et al
. 
Specificity analysis of blood group Lewis-y (Le(y)) antibodies generated against synthetic and natural Le(y) determinants
.
Proc Natl Acad Sci U S A
1994
;
91
:
12957
61
.
5
Yin
BW
,
Finstad
CL
,
Kitamura
K
, et al
. 
Serological and immunochemical analysis of Lewis y (Ley) blood group antigen expression in epithelial ovarian cancer
.
Int J Cancer
1996
;
65
:
406
12
.
6
Myers
RB
,
Srivastava
S
,
Grizzle
WE
. 
Lewis Y antigen as detected by the monoclonal antibody BR96 is expressed strongly in prostatic adenocarcinoma
.
J Urol
1995
;
153
:
1572
4
.
7
Leoni
F
,
Colnaghi
M
,
Canevari
S
, et al
. 
Glycolipids carrying Ley are preferentially expressed on small-cell lung cancer cells as detected by the monoclonal antibody MLuC1
.
Int J Cancer
1992
;
51
:
225
231
.
8
Baldus
SE
,
Monig
SP
,
Zirbes
TK
, et al
. 
Lewis(y) antigen (CD174) and apoptosis in gastric and colorectal carcinomas: correlations with clinical and prognostic parameters
.
Histol Histopathol
2006
;
21
:
503
10
.
9
Zenita
K
,
Kirihata
Y
,
Kitahara
A
, et al
. 
Fucosylated type-2 chain polylactosamine antigens in human lung cancer
.
Int J Cancer
1988
;
41
:
344
9
.
10
Cordon-Cardo
C
,
Lloyd
KO
,
Sakamoto
J
, et al
. 
Immunohistologic expression of blood-group antigens in normal human gastrointestinal tract and colonic carcinoma
.
Int J Cancer
1986
;
37
:
667
76
.
11
Sakamoto
J
,
Furukawa
K
,
Cordon-Cardo
C
, et al
. 
Expression of Lewisa, Lewisb, X, and Y blood group antigens in human colonic tumors and normal tissue and in human tumor-derived cell lines
.
Cancer Res
1996
;
46
:
1553
61
.
12
Scott
AM
,
Geleick
D
,
Rubira
M
, et al
. 
Construction, production, and characterization of humanized anti-Lewis Y monoclonal antibody 3S193 for targeted immunotherapy of solid tumors
.
Cancer Res
2000
;
60
:
3254
61
.
13
Scott
AM
,
Tebbutt
N
,
Lee
FT
, et al
. 
A phase I biodistribution and pharmacokinetic trial of humanized monoclonal antibody Hu3S193 in patients with advanced epithelial cancers that express the Lewis-Y antigen
.
Clin Cancer Res
2007
;
13
:
3286
92
.
14
Krug
LM
,
Milton
D
,
Chen
L
, et al
. 
Targeting Lewis Y (LeY) in small cell lung cancer (SCLC) with a humanzied monoclonal antibody, hu3S193
.
J Clin Oncol
2006
;
24
:
7086
.
15
Boghaert
ER
,
Sridharan
L
,
Armellino
DC
, et al
. 
Antibody-targeted chemotherapy with the calicheamicin conjugate hu3S193-N-acetyl γ calicheamicin dimethyl hydrazide targets Lewisy and eliminates Lewisy-positive human carcinoma cells and xenografts
.
Clin Cancer Res
2004
;
10
:
4538
49
.
16
Therasse
P
,
Arbuck
SG
,
Eisenhauer
EA
, et al
. 
New guidelines to evaluate the response to treatment in solid tumors. European Organization for Research and Treatment of Cancer, National Cancer Institute of the United States, National Cancer Institute of Canada
.
J Natl Cancer Inst
2000
;
92
:
205
16
.
17
Scott
AM
,
Lee
F-T
,
Hopkins
W
, et al
. 
Specific targeting, biodistribution, and lack of immunogenicity of chimeric anti-GD3 monoclonal antibody KM871 in patients with metastatic melanoma: results of a phase I trial
.
J Clin Oncol
2001
;
19
:
3976
87
.
18
Liu
A
,
Williams
LE
,
Raubitschek
AA
. 
A CT assisted method for absolute quantitation of internal radioactivity
.
Med Phys
1996
;
23
:
1919
28
.
19
Scott
AM
,
Lee
F-T
,
Jones
R
, et al
. 
A phase I trial of humanized monoclonal antibody A33 in patients with colorectal carcinoma: biodistribution, pharmacokinetics, and quantitative tumor uptake
.
Clin Cancer Res
2005
;
11
:
4810
17
.
20
Young
H
,
Baum
R
,
Cremerius
U
, et al
. 
Measurement of clinical and subclinical Tumor response using [18F]-fluorodeoxyglucose and positron emission tomography: review and 1999 EORTC recommendations
.
Eur J Cancer
1999
;
35
:
1773
82
.
21
Shankar
LK
,
Hoffman
JM
,
Bacharach
S
, et al
. 
Consensus recommendations for the use of 18F-FDG PET as an indicator of therapeutic response in patients in National Cancer Institute Trials
.
J Nucl Med
2006
;
47
:
1059
66
.
22
Scott
AM
,
Lee
FT
,
Tebbutt
N
, et al
. 
A phase I clinical trial with monoclonal antibody ch806 targeting transitional state and mutant epidermal growth factor receptors
.
Proc Natl Acad Sci U S A
2007
;
104
:
4071
76
.
23
Holden
SN
,
Beeram
M
,
Krop
IE
, et al
. 
A phase I study of weekly dosing of trasutzumab-DM1 (T-DM1) in patients with advanced Her2+ breast cancer [abstract 1029]
.
J Clin Oncol
26: 
2008
(May 20 suppl; abstr 1029).
24
Terpstra
V
,
van Amersfoort
ES
,
van Velzen
AG
, et al
. 
Hepatic and extrahepatic scavenger receptors: function in relation to disease
.
Arterioscler Thromb Vasc Biol
2000
;
20
:
1860
72
.
25
Kooistra
T
,
Duursma
AM
,
Bouma
JM
, et al
. 
Effect of size and charge on endocytosis of lysozyme derivatives by sinusoidal rat liver cells in vivo
.
Biochim Biophys Acta
1980
;
631
:
439
50
.
26
Nishida
K
,
Mihara
K
,
Takino
T
, et al
. 
Hepatic disposition characteristics of electrically charged macromolecules in rat in vivo and in the perfused liver
.
Pharm Res
1991
;
8
:
437
44
.
27
Gillespie
AM
,
Broadhead
TJ
,
Chan
SY
, et al
. 
Phase I open study of the effects of ascending doses of the cytotoxic immunoconjugate CMB-401 (hCTMO1-calicheamicin) in patients with epithelial ovarian cancer
.
Ann Oncol
2000
;
11
:
735
41
.
28
Chan
SY
,
Gordon
AN
,
Coleman
RE
, et al
. 
A phase 2 study of the cytotoxic immunoconjugate CMB-401 (hCTM01-calicheamicin) in patients with platinum-sensitive recurrent epithelial ovarian carcinoma
.
Cancer Immunol Immunother
2003
;
52
:
243
8
.
29
Larson
RA
,
Sievers
EL
,
Stadtmauer
EA
, et al
. 
Final report of the efficacy and safety of gemtuzumab ozogamicin (Mylotarg) in patients with CD33-positive acute myeloid leukemia in first recurrence
.
Cancer
2005
;
104
:
1442
52
.
30
McKoy
JM
,
Angelotta
C
,
Bennett
CL
, et al
. 
Gemtuzumab ozogamicin-associated sinusoidal obstructive syndrome (SOS): an overview from the research on adverse drug events and reports (RADAR) project
.
Leuk Res
2007
;
31
:
599
604
.

Competing Interests

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.