A Versatile Bifunctional Chelate for Radiolabeling Humanized Anti-CEA Antibody with In-111 and Cu-64 at Either Thiol or Amino Groups: PET Imaging Of CEA-Positive Tumors with Whole Antibodies

Abstract

Radiolabeled anti-carcinoembryonic antigen (CEA) antibodies have the potential to give excellent images of a wide variety of human tumors, including tumors of the colon, breast, lung, and medullar thyroid. In order to realize the goals of routine and repetitive clinical imaging with anti-CEA antibodies, it is necessary that the antibodies have high affinity for CEA, low cross reactivity and uptake in normal tissues, and low immunogenicity. The humanized anti-CEA antibody hT84.66-M5A (M5A) fulfills these criteria with an affinity constant >10 10 M −1 , no reactivity with CEA crossreacting antigens found in normal tissues, and >90% human protein sequence. A further requirement for routine clinical use of radiolabeled antibodies is a versatile method of radiolabeling that allows the use of multiple radionuclides that differ in their radioemissions and half-lives. We describe a versatile bifunctional chelator, DO3A-VS (1, 4, 7-tris(carboxymethyl)-10-(vinylsulfone)-1, 4, 7, 10-tetraazacyclododecane) that binds a range of radiometals including 111 In for gamma-ray imaging and 64 Cu for Positron Emission Tomography (PET), and which can be conjugated with negligible loss of immunoreactivity either to sulfhydryls (SH) in the hinge region of lightly reduced immunoglobulins or surface lysines (NH) of immunoglobulins.Methods-Athymic mice peripherally xenografted with CEA-positive human colon tumors (LS-174T) were injected with 111 In-labeled or 64 Cu-labeled SH-DO3A-VS-M5A, NH-DO3A-VS-M5A, or DOTA-M5A and sacrificed at various time points for biodistribution measurements. Other mice injected with 64 Cu-labeled SH-DO3A-VS-M5A or NH-DO3A-VS-M5A were imaged serially with small animal PET from 1 to 48 h post injection and then sacrificed for biodistribution measurements.Results-Virtually identical biodistributions were obtained for SH-and NH-DO3A-VS-M5A or DOTA-M5A whether radiolabeled with 111 In or 64 Cu. Rapid tumor uptake of radiolabel was observed, reaching 40% injected dose/gram or more by 48 h. Importantly, excellent PET images of tumor were obtained as early as 22 h after injection of 64 Cu-labeled SH-or NH-DO3A-VS-M5A. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptConclusions-Based on our correlative studies comparing the kinetics of radiolabeled anti-CEA antibodies in murine models with those in man, we predict that 64 Cu-labeled intact, humanized antibodies can be used to image CEA positive tumors in the clinic.Keywords carcinoembryonic antigen; bifunctional chelate; radioimmunoimaging; positron emission tomographyIn order to be effective tumor imaging agents, radiolabeled anti-tumor antibodies must demonstrate both high tumor uptake and high tumor to normal tissue ratios (1). Monoclonal antibodies (150 kDa) have a relatively slow blood clearance (second phase half-life t 1/2 β = 48-72 h), allowing ample time for high accumulation in tumor, but suffer from low tumor to normal tissue contrast due to slow clearance from the vasculature. In order to improve the dynamic biodistribut...

Radioimmunoimaging of Liver Metastases with PET Using a 64Cu-Labeled CEA Antibody in Transgenic Mice

“…The uptake in C15A.3 lesions was lower than in subcutaneous LS-174T tumors as reported by Li et al [11]. The differences in the cell type and tumor location could both contribute to a different tumor microenvironment, explaining the discrepancy in the results.…”

“…It has been previously reported that the M5A antibody shows outstanding binding characteristics in subcutaneous CEA tumor models [11]. Therefore, we decided to use this promising antibody in a more challenging and novel model of syngeneic liver metastases.…”

Quantitative Serial Imaging of an 124I Anti-CEA Monoclonal Antibody in Tumor-Bearing Mice

“…It has been shown in studies with 64 Cu-labeled anti-CEA mAbs that TACs obtained with microPET and this calibration method produce good agreement with TACs measured for tumor, liver, and blood in separate, parallel groups of mice by direct tissue assay. 24 Of the two anticipated problems for 124 I, high ␤ ϩ energy, and true-coincidence ␥-ray background, the former appears to be the more significant. The positron-range effect can be modeled in iterative reconstruction, and some work has been done on this for MAP.…”

Positive Progress in ImmunoPET—Not Just a Coincidence

“…In a study by Li et al, 26 a humanized anti-CEA intact antibody, hT84.66-M5A (M5A), was labeled with 64 Cu using the chelator 1,4,7-tris(carboxymethyl)-10-(vinylsulfone)-1,4,7,10-tetraazacy clododecane (DO3A-VS). The 64 Cu-labeled conjugate (NH-DO3A-VS-M5A or SH-DO3A-VS-M5A) was then injected into mice bearing CEA-expressing LS-174T tumors.…”