A.M. Neville

Role of immunohistochemical detection of lymph-node metastases in management of breast cancer

BACKGROUND This study was designed to ascertain whether immunohistochemical methods could improve the detection of metastases in primary breast-cancer patients whose axillary lymph nodes were classified, by conventional methods, as disease free. METHODS Ipsilateral lymph nodes (negative for metastases by routine histology) from 736 patients (participants in Trial V of the International [Ludwig] Breast Cancer Study) were examined by serial sectioning and staining with haematoxylin and eosin (two sections from each of six levels) and by immunohistochemistry of a single section (with two anticytokeratins AE-1 and CAM 5.2). After median follow-up of 12 years, disease-free and overall survival were estimated by Kaplan-Meier methods. FINDINGS Occult nodal metastases were detected by serial sectioning and haematoxylin and eosin in 52 (7%) of 736 patients and by immunohistochemistry in 148 (20%). Only two (3%) of 64 invasive lobular or mixed invasive lobular and ductal cancers had node micrometastases, detected by haematoxylin and eosin, compared with 25 (39%) by immunohistochemistry. Occult metastases, detected by either method, were associated with significantly poor disease-free and overall survival in postmenopausal but not in premenopausal patients. Immunohistochemically detected occult lymph-node metastases remained an independent and highly significant predictor of recurrence even after control for tumour grade, tumour size, oestrogen-receptor status, vascular invasion, and treatment assignment (hazard ratio 1.79 [95% CI 1.17-2.74], p=0.007). INTERPRETATION The immunohistochemical examination of ipsilateral axillary lymph nodes is a reliable, prognostically valuable, and simple method for the detection of occult nodal metastases. Immunohistochemistry is recommended as a standard method of node examination in postmenopausal patients.

Monoclonal antibodies to the human mammary gland

A range of primary and metastatic human breast carcinomas has been examined with respect to the staining by four monoclonal antibodies which were raised to the human milk fat globule membrane. Within the normal breast the luminal epithelial cells expressing the antigens detected by the monoclonal antibodies were heterogeneous in their distribution. The heterogeneity was not only confined to single cells, but also to regions within the breast. The breast carcinomas also expressed the antigens in a variable manner. Morphological differentiation and functional differentiation, defined by the monoclonal antibodies, were not invariably coincident. Lymph node metastases gave similar results to the primary carcinomas. The monoclonal antibodies have revealed a heterogeneity, with respect to surface antigenic expression, within the normal and neoplastic breast epithelium. This cellular heterogeneity of breast carcinomas, may have significant prognostic and therapeutic implications in the management of primary breast cancer.

LOCATION OF METASTATIC BREAST CARCINOMA BY A MONOCLONAL ANTIBODY CHELATE LABELLED WITH INDIUM-111

The ability of a radiolabelled monoclonal antibody, LICR-LON-M8 (M8), to locate metastatic breast carcinomas has been investigated. The scans generated by M8, either when labelled with radioiodine, or when conjugated with diethylenetriamine-pentaacetic acid (DTPA) and labelled with radioactive indium (111In), have been compared with X-rays and 99mTc-methyl diphosphonate (MDP) bone scans. All 10 patients with skeletal metastases had positive 111In-DTPA-M8 scans and the overall correlation with X-rays and MDP scans was good but varied with the region studied. By contrast, radioiodinated M8 did not detect metastases at any site. The discrepancies between 111In-DTPA-M8 images and conventional techniques may be related to the different stages in the evolution and development of the lesion at which the various techniques detect bone metastases.

Ectopic production of human chorionic gonadotrophin-like material by breast cancer.

Human chorionic gonadotrophin (hCG) is a placental protein whose ectopic secretion by nontrophoblast tumors has been claimed to be of clinical relevance. Serum levels of hCG were measured in 570 patients with breast disease. A double‐antibody radioimmunoassay (RIA) using antisera to hCG‐β was employed. Approximately 14% of patients with breast cancer were found to have elevated serum hCG levels. Such raised titers were not stage‐ or tumor‐type‐related, but occurred only in postmenopausal subjects. Further study showed that those patients with elevated hCG levels also had raised levels of human luteinizing hormone (hLH). Assay cross‐reactivity was shown to account for the “spurious” hCG elevations. An immunocytochemical study also failed to find hCG an ectopic breast tumor constituent and/or product. It is concluded that hCG is not produced by breast tumors and has no clinical utility.

ELIMINATION OF CARCINOMA CELLS FROM HUMAN BONE MARROW

The value of a monoclonal antibody, LICR-LON-Fib-75 (Fib-75), which mediates complement lysis and recognises an antigen present on all epithelial-tumour cells so far studied but not on lymphoid cells or bone-marrow stem cells, in clearing infiltrated bone marrow was assessed. Chromium-51-release and trypan-blue-exclusion assays showed that no tumour cells were viable after exposure to Fib-75 and complement except when large clumps (greater than 500 cells) were present, whereas Fib-75 had no significant effect on bone-marrow cells, as judged by colony-forming-unit and pluripotential-stem-cell assays.

Screening for metastases in breast cancer: An assessment of biochemical and physical methods

Ten tumor markers were measured in serum or urine at approximately three month intervals in patients with breast cancer following mastectomy but before development of overt metastatic disease. In 23 patients who later had metastases, only three markers, alkaline phosphatase, carcinoembryonic antigen (CEA), and γ‐glutamyl transpeptidase (γ‐GT) were consistently abnormal prior to the development of detectable metastases in more than one patient. In half the patients, a “lead interval” of three months or more was obtained using these three markers and little advantage was obtained by the addition of any other biochemical marker. The value of these three measurements was then assessed in a larger group of patients and compared with other tests for metastases. Alkaline phosphatase, CEA, γ‐GT, clinical examination, and chest x‐ray were the best indices of the metastatic state in breast cancer, being collectively abnormal in 98% of patients at first presentation with metastases. The authors recommend screening patients postoperatively with these five tests for metastases; more detailed tests should only be carried out if results of one or more of these are abnormal.

Ultrastructural identification of Ia positive dendritic cells in the lactating rat mammary gland.

Dendritic cells which express Ia antigen have been demonstrated for the first time in the lactating rat mammary gland. Ultrastructurally, the dendritic cells appear as electron-lucent pale cells interspersed among the epithelial cells of the alveoli, forming a cell population distinct from classical macrophages. They show morphological resemblance to the dendritic cells of lymphoid organs as well as the Langerhans cells of skin. The Ia antigen has been localised by electron microscopic immunocytochemistry on the cell membrane and endocytotic vesicles and tubules. Ia positive cells are also seen in the stroma of the mammary gland. It is proposed that the dendritic cells of the mammary gland belong to the lineage of epidermal Langerhans cells and lymphoid dendritic cells, subserving an immunological role in the lactating breast.

The selective culture of keratinocytes using a cytotoxic antifibroblast monoclonal antibody

A technique is described for the selective killing of fibroblasts in primary cultures and subcultures of human keratinocytes using a monoclonal antibody raised against human fibroblasts. The antibody kills fibroblasts by complement mediated cytotoxicity but is not toxic to keratinocytes.

Human Breast Carcinoma Localization with a Radiolabelled Monoclonal Antibody

Abstract A monoclonal antibody recognizing an antigen on breast epithelial cells has been labelled with radioactive isotopes of iodine and indium. The effect of the label on the localization of human breast carcinoma by the antibody both in the animal model and in man has been studied. Superior localization achieved with indium labelled antibody has been clearly demonstrated.