Chantal Curtet

A convenient, novel approach for the synthesis of polyaza macrocyclic bifunctional chelating agents

Abstract The convenient, synthetically useful bifunctional chelating agents, (10-p-aminobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetate (p-ABz-DO3A) 3 and (11-p-aminobenzyl)-1,4,8,11-tetraazacyclotetradecane-1,4,8-triacetate (p-ABz-TE3A) 6 , were obtained by reaction of an ethyl bromoacetate with 1,4,7,10-tetraazacyclododecane and 1,4,8,11-tetraazacyclotetradecane followed by reaction with nitrobenzyl bromide. This method proved more efficient than any described in the literature since the overall yield in a two-step synthesis sequence starting from tetraazamacrocycles was 68%.

Pharmacokinetic study of radiolabeled anti-colorectal carcinoma monoclonal antibodies in tumor-bearing nude mice

Monoclonal antibodies (MoAbs) 17-1A and 19-9, which specifically bind human colorectal carcinoma (CRC) cells, were tested for their usefulness in localizing colorectal tumors in nude mice. One of the 131I-labeled MoAbs and an irrelevant 125I-labeled immunoglobulin of the same isotype were injected into nude mice simultaneously bearing a human CRC and a human melanoma. The percentage of the injected dose of antibody per gram of tissue, the CRC/tissue ratios of antibody distribution, and the localization indices were calculated at various time intervals (2 h to 9 days). For both MoAbs, labeling to a specific activity of 10 μCi/μg by the iodogen method gave optimum immunoreactivity. The accumulation of MoAb 17-1A in CRC reached its maximum at 5 days and remained at this level for up to 9 days postinjection. For MoAb 19-9, which detects a circulating antigen shed by the tumor into the serum, the accumulation in the CRC was maximum at 24 h, and decreased thereafter. The CRC/organ ratios and localization indices for both MoAbs increased with time in the CRC tissue, but remained low and unchanged in the melanoma and normal tissues. Using F(ab′)2 antibody fragments, faster kinetics with earlier maximum accumulation, higher tumor/organ ratios, and better localization indices were achieved than with intact MoAbs. The data obtained was useful in defining parameters which must be considered before radiolabeled MoAbs are used in cancer patients for diagnostic purposes.

Feasibility study of radioimmunoguided surgery of colorectal carcinomas using indium-111 CEA-specific monoclonal antibody

The study was undertaken to define the potential use of indium 111 carcinoembryonic antigen-specific antibody labelled [CEA F(ab′)2] for the radioimmunodetection of colorectal carcinoma using an intraoperative hand-held gamma probe. The use of a linear radioactive source allowed optimization of physical characteristics. The best results regarding sensitivity and resolution were obtained using a 5-mm thick tungsten alloy collimator. A simulation study with a liver phantom (22 MBq or 0.6 mCi) was performed to determine the effect of side scatter as opposed to direct background and showed that it is possible to detect small radioactive targets (3.7 KBq or 0.1 μCi) 4 cm from the phantom. A clinical study performed with ten patients showed that tumours with good uptake of CEA-specific antibody could be detected with sufficient contrast in two patients when the probe was used. Results of a biodistribution study performed after tumour fragment or normal tissue countings in a well counter showed high tumour uptake (above 8 x 10−3 injected dose/g) and tumour-to-normal tissue ratios (between 2.5 and 20) in five patients. Results with the probe showed markedly lower ratios. There was no correlation between absolute tumour uptake and the count rates of tumour measured intraoperatively. This can be attributed to the degradation of depth resolution resulting from the high energy photopeak of gamma-emitting111In.

Immunotherapy of gastrointestinal cancer with monoclonal antibodies

Twenty patients with widespread metastatic colorectal carcinoma were infused with 500 mg of the cytotoxic IgG2a monoclonal antibody 17-1A preincubated with autologous peripheral blood leukocytes. Ten patients showed no benefit from such therapy and ten died, with a mean survival time of 7.6±4.5 months after treatment and a median survival of 6 months. In ten additional patients, the sourse of disease was modified by antibody therapy; disease in five of these patients stabilized, while tumor size in the other five patients decreased after therapy. Median actual survival in this group of ten patients is presently 24 months; four of these patients died of disease progression within a mean of 15±5 months. Duration of response was 10.5±6.7 months after antibody treatment. Treatment tolerance for these 20 patients was excellent in all but one patient, who experienced an anaphylactic reaction during a second infusion with 17-1A.

Enzyme-linked immunosorbent assay to monitor colorectal carcinoma patients treated with a monoclonal antibody (17-1A)

An enzyme-linked immunosorbent assay (ELISA) was compared to a radioimmunoassay (RIA) for the detection and quantification of mouse monoclonal antibody MoAb 17-1A and for measurement of the host response (i.e. anti-mouse immunoglobulin in sera from patients receiving immunotherapy with MoAb 17-1A. Comparable sensitivity and reproducibility were noted with RIA and ELISA but ELISA was more rapid to perform than RIA. Thus quantitative ELISA compared favorably with the RIA for MoAb detection.

Magnetic resonance imaging studies on nude mice grafted with colorectal adenocarcinoma using gadolinium-labeled monoclonal antibody.

Saccavini JC, Curtet C, Bohy J, Tellier C, Bourgoin C, Lhoste JM. Magnetic resonance imaging studies on nude mice grafted with colorectal adenocarcinoma using gadolinium‐labeled monoclonal antibody. Invest Radiol 1988;23(Suppl 1):S292‐S293. A monoclonal antibody (Ab) 19.9 specific for colorectal carcinoma was labeled with a high number of gadolinium (Gd) atoms for its potential application as a contrast agent in magnetic resonance imaging (MRI). The DTPA was conjugated to 19.9 Ab via the bicyclic DTPA anhydride method (c.DTPA) using c.DTPA/Ab molar ratios between 5 and 150. The aggregates present in great amount at high c.DTPA/Ab ratios were systematically removed. Then the exact number of DTPA effectively conjugated, the immunoreactivity of the resulting 111In‐DTPA‐Ab were measured. The number of DTPA conjugated per antibody can be increased 20 to 25 with only a little loss of immunoreactivity. The 19.9 antibody conjugated with 16 and 25 DTPA was labeled with 153GdCl3 for pharmacokinetic studies on xenografted nude mice and with nonradioactive gadolinium to measure ex vivo the effect on the relaxation time T1 of the tumor. We found a 15 to 20% decrease of T1 on the tumor. In vivo experiments using a Bruker system and the same animal model showed a difference in the tumor contrast after the injection of 2 mg of Gd‐labeled Ab.

Proton NMR study of the binding of concanavalin A on myeloma plasma membranes

Abstract The dynamics of the lipids inside plasma membranes is investigated using proton NMR spectroscopy. Spectra are performed on two different fractions of purified membranes collected on a zonal rotor. A lectin, concanavalin A, provides a restriction of mobility of the lipids on one of these fractions. Enzymatic assays and chemical determinations are investigated in order to discuss the NMR results.

Monoclonal Antibody Labeling with Indium-111 and Gadolinium via DTPA Chelation on Selective and Nonselective Sites of the Antibody

The recent development of hybridoma technology has allowed the production of large quantities of monoclonal antibodies with predefined specificity (1); this has resulted in a renewed interest in their use as immunodiagnostic reagents. Radiolabeled monoclonal antibodies are today widely used for specific localization of tumors and metastases (2–4). The success of immunoscintigraphy has generated interest in the application of monoclonal antibodies as specific carriers of paramagnetic agents for magnetic resonance imaging (MRI). MRI has certain advantages over immunoscintigraphy — no ionizing radiation is involved, and the spatial resolution is higher and equals that of computed tomography. Early investigators felt that differences in relaxation times between malignant tumor and normal tissue made contrast agents unnecessary. However, despite the intrinsic tissue contrast, the injection of contrast agents, e.g., gadolinium-diethylenetriaminepentaacet ic acid (Gd-DTPA) increases the sensitivity and specificity of MR imaging as demonstrated by several investigators (5–7). Of the available paramagnetic ions, gadolinium has the greatest effect on proton relaxation time. Its association with DTPA produces a compound that gives a marked reduction in the proton relaxation time in vitro and in vivo and produces minimal acute toxicity with imaging doses (8,9).

Immunohistocytochemical Analysis of Gastrointestinal Carcinomas Using Immunoperoxidase and Immunogold Staining with Monoclonal Antibodies 19.9 and Anti CEA

Abstract Two monoclonal antibodies anti-CEA and 19.9, have been tested at tissue and cell levels by immunoperoxidase in light microscopy, and immunogood staining in electron microscopy on gastrointestinal adenocarcinomas. With immunoperoxidase assay, 122 primary tumors have been tested: Out of 42 colorectal carcinomas, positivity was 76% with MoAb 19.9, and 90% with MoAb anti-CEA; With other gastrointestinal carcinomas (pancreas, biliary-tract, stomach) positivity was 92% with MoAb 19.9, and 73% with MoAb anti-CEA; for non gastrointestinal adenocarcinomas (breast, parotid, bronchoalveolar, ovary) positivity 32% with MoAb 19.9, and 20% with MoAb anti-CEA; All non glandular tumors (lymphosarcoma, insulinoma, glucagonoma) were negative. Twelve primary tumors and their recurrences were tested, the two monoclonal antibodies appeared to be complementary. With immunogold staining, four colorectal adenocarcinomas have been tested. We observed heterogeneous antigenic display in malignant cells. The labelling was localized punctually, on the cytoplasm, never on the nucleus, just a little on cell membrane. The labelling was localized preferentially at the apical pole of the enterocyte.