New immunoassays for MUC1 in breast cancer.

Eleven experimental immunofluorometric assays (IFMAs) were made using antibodies previously tested for epitope specificities. These assays were compared with six commercially available immunoassays. The clinical performance of these experimental assays was evaluated by analysing sera from 138 breast cancer patients and 105 female blood donors. The clinical performance of these assays was evaluated at a set specificity of 0.94. The highest overall sensitivity (0.56) was observed in the experimental assay with the antibody BC2 as solid phase and GP1.4 as the tracer antibody. This combination also showed the highest sensitivity in stage I/II breast cancer. The Truquant assay (Biomira) had an overall sensitivity of 0.51, and the highest sensitivity in stages III and IV at 0.65 and 0.94, respectively. The remaining commercial assays, with sensitivity ranging from 0.67 to 0.79, were below the top five experimental assays that showed sensitivity values between 0.79 and 0.85. The findings from our current study suggest that further development in MUC1 immunoassays could improve the detection of relapse in breast cancer patients.

Elevated CA125 in breast cancer--A sign of advanced disease.

The serum tumor markers CA 125, MUC1 and CEA were measured in 221 breast cancer patients over a period of 2 years. Patients examined on at least three occasions were included in the study. Thirty-three patients had increasing or continuously high concentrations of CA 125. Thirty (91%) of these had involvement of the pleura, either as pleural metastasis or metastasis in surrounding tissue i.e. bone structures in the thorax cavity or lung parenchyma. MUC1 and CEA were elevated in 27 (82%) and 24 (73%) of the 33 patients, respectively. Increased concentrations of these two markers did not relate to the site of metastasis. However, the three tumor markers complemented each other in detecting early metastases. Increased CA 125 was associated with metastasis in or near the pleura, and in stage IV breast cancer it was related to poor prognosis.

Automated immunofluorometric assay for MUC1.

The aim of the present study was to establish a robust, reliable and fully automated immunofluorometric assay for the breast cancer serum marker MUC1. This would further serve as a prototype assay for evaluation of other MUC1 assays based on new antibody combinations. Using time-resolved fluorescence as tracer signal we developed an automated immunofluorometric assay for MUC1 (MUC1 IFMA). This assay was compared with two commercial assays. The CA15-3 EIA (CanAg) which use the same antibodies as the MUC1 IFMA, and the ETI-CA-15-3 K (Sorin) which use the original antibodies defining the CA 15-3 assay. The three assays showed comparable results. The coefficient of variation was below 10% from 9 to 2,400 kU/l for the MUC1 IFMA, from 15 to 250 kU/l for the CA15-3 EIA, and from 25 to 200 kU/l for the ETI-CA-15-3 K assay. At a specificity of 0.94 the overall diagnostic sensitivities for the MUC1-IFMA, CA15-3 EIA and ETI-CA-15-3 K assays were 0.40, 0.37, and 0.38, respectively. When applied to metastatic breast cancer, all assays had sensitivities close to 0.80. There was a close correlation (Spearman rank = 0.99) between results from the new assay and the CA15-3EIA. The new automated assay was not strictly immunometric as we could not achieve conditions where solid phase or tracer antibodies were in apparent excess. However, the assay performed well at a wide range of assay conditions. The automation, which minimizes imprecision in pipetting and handling of samples, and the high capacity of the AutoDELFIA instrument enabling measurement of all samples in a single run, were important aspects for establishing a reliable assay. The principle of the new automated immunofluorometric assay will be used as a rapid and reliable evaluation of a wide range of monoclonal antibody combinations in our search for the optimal MUC1 assay. This new automated immunofluorometric assay will be useful in the rapid and reliable evaluation of a wide range of monoclonal antibody combinations in our search for the optimal MUC1 assay.

[Abdominal metastases from signet cell carcinoma of the appendix vermiformis]. / Metastaser i bukhulen fra signetcellekarsinom i appendix vermiformis.

Signet cell carcinomas are often aggressive tumours. A patient with this tumour, originally located in the appendix, was diagnosed with peritoneal metastases. When she later got abdominal symptoms, computer axial tomography indicated large infiltrating tumour masses in the pelvis. However, when a laparotomy was performed, the only macroscopic tumour masses was localised to the ovaries (Krukenberg tumour). The surgical intervention gave the patient an improved quality of life, and most likely prolonged survival. We stress the importance of thinking of this possibility.

Summary report on the ISOBM TD-6 workshop: analysis of 20 monoclonal antibodies against Sialyl Lewisa and related antigens. Montreux, Switzerland, September 19-24, 1997.

The ISOBM TD-6 Workshop is the first international workshop on monoclonal antibodies against the Sialyl Lewisa (SLea) antigen. Eight research groups participated in a blind study to characterize the epitope binding, relative affinity and performance in immunoradiometric assays, of a panel of 20 monoclonal antibodies. The antibodies were tested against a diverse panel of neoglycoconjugates, purified antigens and human serum pools from gastrointestinal malignancies. Epitope specificities were determined for the majority of antibodies in the panel. Cross-reactivity with related saccharide structures was noted in several antibodies. Overall, the results of the TD-6 Workshop show further development of SLea immunoassays may yield yet more specific assays for the detection and management of gastrointestinal and other malignancies.

Carcinoma-associated MUC1 detected by immunoradiometric assays.

Fifty-four anti-MUC1 antibodies submitted to the International Society for Oncodevelopmental Biology and Medicine (ISOBM) Workshop (TD-4) were evaluated in immunoradiometric assays, using sera from carcinoma patients and healthy donors. The carcinoma serum pool contained sera from 30 patients with advanced cancer (10 breast, 10 colon, and 10 ovarian). This serum pool contained 696 kU/l MUC1, 770 micrograms/l CEA, and 3,700 kU/l CA 125. The reference serum pool was obtained from 10 healthy women combined with 20 sera from pregnant women, of which half had elevated CA 125 (range 82-254 kU/l). The reference serum pool contained 13 kU/l MUC1, 2 micrograms/l CEA, and 65 kU/l CA 125. The Workshop antibodies were tested both as solid-phase antibodies and as tracer antibodies with the carcinoma serum pool. Twenty-two tracer antibodies and 38 solid-phase antibodies gave at least one combination with > 10% binding of the tracer antibody for a total of 836 combinations. These were tested further with the reference serum pool. Antibodies used as tracers could be separated into three categories: Group 1 antibodies, MF06, MF11, B27.29, MF30, and Ma552, gave mainly 'carcinoma-specific' assays in combinations with the solid-phase antibodies, i.e. binding ratio between carcinoma MUC1 and reference MUC1 > 10. Group 2 antibodies, DF3, 7540MR, A76-A/C7, BC4N154, M38, 7539MR, B12, GP1.4, 232A1, Mc5, and Ma695 gave both 'specific' and 'nonspecific' binding ratios depending on the solid-phase antibody used. Group 3 antibodies, 214D4, BC4E549, E29, BCP8, BC3, and 3E1.2, gave mainly 'nonspecific' combinations, i.e. ratios < or = 10. All antibodies used to capture MUC1 on the solid phase gave both 'specific' and 'nonspecific' combinations depending on the tracer antibody used. Ten antibodies were clearly more efficient as solid-phase capture antibodies; Ma695, B12, M38, GP1.4, 214D4, MF06, B27.29, A76-A/C7, BC3, and KC4. Our findings indicate that the ability to detect 'carcinoma-specific MUC1' cannot be deduced from epitope specificity alone.

Brain metastasis model in athymic nude mice using a novel MUC1-secreting human breast-cancer cell line, MA11.

The MA11 human breast-cancer cell line was established with cells isolated from a bone-marrow sample using immunomagnetic beads conjugated to the anti-MUC1 antibody BM-2. The cell line showed a selective preference for metastasising to the brain in athymic nude mice. Following injection of MA11 cells into the left ventricle of the heart, brain metastases developed in 87% (20/23) animals, with a mean latency until development of neurological symptoms of 65 days. Necropsy and histological examination revealed tumour nodules of varying sizes throughout the brain, invading both grey and white matter of both hemispheres, and with extensive involvement of the cerebellum. MRI spin-echo images indicated brain lesions in some animals that were subsequently confirmed by histology. Three mice showed small tumour nodules (1-2 mm) in the lung, and 2 had solitary lesions (< 1 mm) within the spinal cord. Metastases were not detected in bone, liver, adrenal gland, kidney, spleen or heart. The human MUC1 mucin, as determined by a europium-based immunoradiometric assay, was detected in the serum of 9/11 animals that showed histological evidence of brain metastases. The mucin could not be found in mouse serum samples taken before day 46. The concentration range of MUC1 observed was from <1 to >50 U/ml, and did not appear to correlate with the size or number of tumours as determined from histological sections. This new model provides an opportunity to study the mechanisms of clinically relevant organ-selective metastases and may be of use in evaluating novel treatment for brain metastases in breast cancer.