Rafke Schoffelen

Pretargeted Immuno–Positron Emission Tomography Imaging of Carcinoembryonic Antigen–Expressing Tumors with a Bispecific Antibody and a 68Ga- and 18F-Labeled Hapten Peptide in Mice with Human Tumor Xenografts

18F-Fluorodeoxyglucose (18F-FDG) is the most common molecular imaging agent in oncology, with a high sensitivity and specificity for detecting several cancers. Antibodies could enhance specificity; therefore, procedures were developed for radiolabeling a small (∼1451 Da) hapten peptide with 68Ga or 18F to compare their specificity with 18F-FDG for detecting tumors using a pretargeting procedure. Mice were implanted with carcinoembryonic antigen (CEA; CEACAM5)–expressing LS174T human colonic tumors and a CEA-negative tumor, or an inflammation was induced in thigh muscle. A bispecific monoclonal anti-CEA × anti-hapten antibody was given to mice, and 16 hours later, 5 MBq of 68Ga- or 18F-labeled hapten peptides were administered intravenously. Within 1 hour, tissues showed high and specific targeting of 68Ga-IMP-288, with 10.7 ± 3.6% ID/g uptake in the tumor and very low uptake in normal tissues (e.g., tumor-to-blood ratio of 69.9 ± 32.3), in a CEA-negative tumor (0.35 ± 0.35% ID/g), and inflamed muscle (0.72 ± 0.20% ID/g). 18F-FDG localized efficiently in the tumor (7.42 ± 0.20% ID/g) but also in the inflamed muscle (4.07 ± 1.13% ID/g) and in several normal tissues; thus, pretargeted 68Ga-IMP-288 provided better specificity and sensitivity. Positron emission tomography (PET)/computed tomography images reinforced the improved specificity of the pretargeting method. 18F-labeled IMP-449 distributed similarly in the tumor and normal tissues as the 68Ga-labeled IMP-288, indicating that either radiolabeled hapten peptide could be used. Thus, pretargeted immuno-PET does exceptionally well with short-lived radionuclides and is a highly sensitive procedure that is more specific than 18F-FDG-PET. Mol Cancer Ther; 9(4); 1019–27. ©2010 AACR.

Development of an imaging-guided CEA-pretargeted radionuclide treatment of advanced colorectal cancer: first clinical results

Background:Radiolabelled antibody targeting of cancer is limited by slow blood clearance. Pretargeting with a non-radiolabelled bispecific monoclonal antibody (bsMAb) followed by a rapidly clearing radiolabelled hapten peptide improves tumour localisation. The primary goals of this first pretargeting study in patients with the anti-CEACAM5 × anti-hapten (HSG) bsMAb, TF2, and the radiolabelled hapten-peptide, IMP288, were to assess optimal pretargeting conditions and safety in patients with metastatic colorectal cancer (CRC).Methods:Different dose schedules were studied in four cohorts of five patients: (1) shortening the interval between the bsMAb and peptide administration (5 days vs 1 day), (2) escalating the TF2 dose (from 75 to 150 mg), and (3) reducing the peptide dose (from 100 to 25 μg). After confirmation of tumour targeting by 111In-IMP288, patients were treated with a bsMAb/177Lu-IMP288 cycle.Results:Rapid and selective tumour targeting of the radiolabelled peptide was visualised within 1 h, with high tumour-to-tissue ratios (>20 at 24 h). Improved tumour targeting was achieved with a 1-day interval between the administration of the bsMAb and the peptide and with the 25-μg peptide dose. High 177Lu-IMP288 doses (2.5–7.4 GBq) were well tolerated, with some manageable TF2 infusion reactions, and transient grades 3–4 thrombocytopaenia in 10% of the patients who received 177Lu-IMP288.Conclusion:This phase I study demonstrates for the first time that pretargeting with bsMAb TF2 and radiolabelled IMP288 in patients with CEA-expressing CRC is feasible and safe. With this pretargeting method, tumours are specifically and rapidly targeted.

Pretargeted immuno-PET of CEA-expressing intraperitoneal human colonic tumor xenografts: a new sensitive detection method

BackgroundIn this study, pretargeted immuno-positron-emission tomography [PET] with a bispecific monoclonal anti-carcinoembryonic antigen [CEA] (CEACAM5) × anti-hapten antibody (bispecific monoclonal antibody [bsmAb]) and a small (1.5 kD) peptide labeled with 68Ga was compared to fludeoxyglucose [18F-FDG]-PET for detecting intraperitoneal [i.p.] CEA-expressing human colonic tumor xenografts in nude mice.MethodsTwo groups of female BALB/c nude mice were inoculated with LS174T human colonic tumor cells i.p. One group received 5 MBq 18F-FDG, and the other received intravenous injections of the bsmAb, followed 16 h later with 5 MBq of 68Ga-labeled peptide. One hour after the radiolabeled peptide or FDG was given, micro-PET/computed tomography images were acquired. Thereafter, the uptake of the 68Ga or 18F in dissected tissue was determined.ResultsWithin 1 h, high uptake of the 68Ga-labeled peptide in the tumor lesions (23.4 ± 7.2% ID/g) and low background activity levels were observed (e.g., tumor-to-intestine ratio, 58 ± 22). This resulted in a clear visualization of all intra-abdominal tumor lesions ≥ 10 μL and even some tumors as small as 5 μL (2 mm diameter). 18F-FDG efficiently localized in the tumors (8.7 ± 3.1% ID/g) but also showed physiological uptake in various normal tissues (e.g., tumor-to-intestine ratio, 3.9 ± 1.1).ConclusionsPretargeted immuno-PET with bsmAb and a 68Ga-labeled peptide could be a very sensitive imaging method for imaging colonic cancer, disclosing occult lesions.

Quantitative Immuno-SPECT Monitoring of Pretargeted Radioimmunotherapy with a Bispecific Antibody in an Intraperitoneal Nude Mouse Model of Human Colon Cancer

The prospects for using pretargeted immuno-SPECT to monitor the response to pretargeted radioimmunotherapy were examined. In this study, a bispecific anticarcinoembryonic antigen (CEACAM5; CD66e) × antihapten monoclonal antibody, TF2, was used in combination with a small (1.5 kD) peptide, IMP288, labeled with 111In and 177Lu. Methods: First, tumor uptake of 111In-IMP288 and 177Lu-IMP288, as determined by immuno-SPECT, was validated by ex vivo counting. Two groups of female BALB/c nude mice had LS174T tumors implanted in the peritoneal cavity. They received intravenous injections of TF2, followed by 10 MBq of 111In-IMP288 or 90 MBq of 177Lu-IMP288. A control group of non–tumor-bearing mice received TF2 and 111In-IMP288. One hour after the radiolabeled IMP288 was given, small-animal SPECT/CT images were acquired, and subsequently animals were dissected. Furthermore, a survival study was performed in 3 groups of 10 mice with intraperitoneal tumors: mice received TF2 and 177Lu-IMP288 (60 MBq), nonpretargeted 177Lu-IMP288 (60 MBq), or phosphate-buffered saline. Immuno-SPECT scans were acquired directly after therapy and at 14 and 45 d after therapy. Tumor growth was analyzed in the successive scans in each animal. Results: 111In- and 177Lu-labeled IMP288 had similar in vivo distribution. The activity measured in the pretargeted immuno-SPECT images correlated well with the uptake measured in the dissected tumors (Pearson r = 0.99, P < 0.05). In the therapy study, the SPECT images showed rapid and selective tumor targeting with high tumor-to-background contrast (30 ± 12) as early as 1 h after injection. The successive images of the treated mice showed delayed tumor growth in the pretargeted radioimmunotherapy group, corresponding with their prolonged survival. Conclusion: Pretargeted immuno-SPECT with TF2 and 111In- or 177Lu-IMP288 can be used to predict and confirm tumor targeting and monitor the therapeutic effect of pretargeted radioimmunotherapy.

Abstract 1755: Pretargeted radioimmunotherapy of an anti-CEA bispecific antibody and 177Lu-labeled peptide: a phase I study in patients with advanced colorectal cancer

Table of Contents * Pretargeted radioimmunotherapy of an anti-CEA bispecific antibody and 177Lu-labeled peptide: a phase I study in patients with advanced colorectal cancer * Abstract Objectives: Pretargeted radioimmunotherapy (PRIT) with bispecific monoclonal antibodies (bsMAb) in combination with a radiolabeled peptide reduces the radiation dose of conventional RIT to normal tissues by rapid tumor targeting and fast blood clearance of the radiolabeled molecule. In this phase I trial, PRIT was investigated in 20 patients with advanced colorectal cancer (CRC), using a humanized recombinant bsMAb, TF2, targeting carcinoembryonic antigen (CEACAM5) and the hapten-peptide, IMP288, which was radiolabeled with177Lu. The safety, pharmacokinetics, and tumor targeting of these agents were studied, by optimizing the dose schedule of the pretargeting system. Methods: We included four cohorts with five patients per cohort. We studied the interval between the bsMAb and peptide administration and its consequences for the blood clearance of the radiolabeled peptide (cohort 1: 5 days interval vs next cohorts: 1 day). We escalated TF2 dose to saturate tumor antigens (cohort 3: 150 mg TF2, vs other cohorts: 75 mg). Furthermore, we investigated the effect of labelling the same radioactivity doses to a lower peptide dose, which ideally would increase the fractional targeting to the tumors (cohort 4: 25 µg IMP288 vs other cohorts: 100 µg) A pre-therapy cycle with 111In-labeled IMP288 was used to predict the pharmacokinetics and tumor targeting of 177Lu-IMP288, and to assure safety of high 177Lu activity doses (3.7 GBq in cohort 1 and ≤ 7.4 GBq in cohorts 2-4). Toxicity was determined by NCI-CTC v3.0. Whole-body planar and SPECT images were acquired. Results: Toxicity was generally very mild, with only two patients showing grade 3-4 thrombocytopenia, and one with grade 3 dyspnea. Dose escalation did not increase toxicity. Rapid localization of the radiolabeled peptide in the tumor lesions was clearly visualized with scintigraphy within 1 h of injection, with excellent tumor-to-normal tissue ratios (>20 at 24 h after injection in all cohorts). Low absorbed radiation doses to the red bone marrow ( Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1755. doi:1538-7445.AM2012-1755

Abstract 5282: Pretargeted immunoPET for imaging human colonic cancer in a mouse model

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Objective: Molecular imaging with pretargeting bispecific antibodies allows rapid and specific targeting of tumors for PET imaging. In this study, pretargeted immunoPET with a bispecific monoclonal anti-carcinoembryonic antigen (CEA; CEACAM5) × anti-hapten antibody (bsMAb) and a small (1.5 kD) peptide labeled with a short-lived positron-emitter 68Ga was compared to FDG for detection of intraperitoneal CEA-expressing human colonic tumor xenografts in nude mice. Methods: In BALB/c nude mice LS174T human colonic tumor cells were injected intraperitoneally. One group (n=5) received 5 MBq FDG i.v., and another group (n=5) received the bsMAb i.v. and 5 MBq of 68Ga-labeled peptide i.v. 16 h later. One hour after the injection of the peptide or FDG, PET/CT images were acquired. One day later the same procedure was repeated vice versa, and after microPET imaging the uptake of the 68Ga or 18F in dissected tissue was determined. Results: Within one hour, specific tumor targeting of 68Ga-labeled peptide (23.4 ± 7.2% ID/g) combined with the low background activity levels resulted in the clear visualization of the small intra-abdominal tumor lesions (1.5-5 mm). High tumor/background radioactivity concentration ratios were obtained (e.g. tumor/intestines 57.5 ± 22.4). FDG localized efficiently in the tumors (8.7 ± 3.1 % ID/g), but with physiological uptake in various normal tissues (ratio tumor/intestines 4.0 ± 0.9). Conclusion: Pretargeted immunoPET performs exceptionally well with short-lived radionuclides, and is a highly sensitive procedure for dectection of small tumor deposits that is also more specific than FDG-PET. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5282. doi:10.1158/1538-7445.AM2011-5282

Abstract A32: Pretargeted immunoPET of CEA-expressing in intraperitoneal human colonic tumor xenografts in nude mice

Objective: Radiolabeled antibodies were being developed commercially for the detection of several cancer types. However, imaging with radiolabeled antibodies require a relatively long interval between injection and imaging acquisition for adequate contrast to develop. The pretargeting method is a 2-step method: a unlabeled bsMAb with affinity for the tumor and a small radiolabeled molecule is injected, followed by the a radiolabeled compound. This aims to optimize the imaging system by specific tumor accumulation and fast blood clearance of the radiopharmacon, which allows imaging within one hour. Short-lived positron emitting radionuclides, e.g. 68Ga (half life: 68 min) match the pharmacokinetics of the small molecule. In this study, pretargeted immunoPET with a bispecific monoclonal anti-carcinoembryonic antigen (CEA) x anti-histamine-succinyl-glycine (HSG) antibody, TF2, and a radiolabeled peptide, IMP288, was compared to 18F-FDG in detecting CEA-expressing tumors in nude mice. Methods: Groups of five BALB/c nude mice were inoculated with CEA-expressing human colonic tumor cells intraperitoneally. One group received 5 MBq 18F-FDG intravenously (i.v.). Another group received TF2 and 5 MBq 68Ga-IMP288 i.v. 16 h later. After one hour PET/CT images were acquired. One day later this was repeated vice versa. Uptake of 68Ga-IMP288 or 18F-FDG in the tumors and other dissected tissues was determined. Results: High, specific tumor uptake of 68Ga-IMP288 (23.4 ± 7.2% ID/g) was observed, with very low accretion in the normal tissues (intestines: 0.50 ± 0.19 % ID/g; liver: 1.83 ± 0.45 % ID/g). This resulted in high tumor-to-background ratios (e.g. tumor-to-intestines 57.5 ± 22.4) and in the clear visualization of the small intra-abdominal tumor lesions (1.5–5 mm), as soon as one hour after the injection of 68Ga-IMP288. 18F-FDG localized efficiently in the tumors (8.7 ± 3.1 % ID/g), but with physiological uptake in various normal tissues (intestines: 2.15 ± 0.61 % ID/g; liver: 2.78 ± 0.52 % ID/g). The low tumor-to-background ratio (e.g. tumor-to-intestines 4.0 ± 0.9) complicated the detection of the tumors. Conclusions: This study indicated that pretargeted immunoPET with TF2 and 68Ga-IMP288 is a specific and sensitive method for detecting colon cancer. Citation Information: Clin Cancer Res 2010;16(7 Suppl):A32