The expression of leukocytic and related antigens on the cells of human breast cancer cells]

Certain expression regularities of different markers in human breast cancer cells are demonstrated. Monomorphic determinants of HLA class II antigen are detected only in case of HLA class I expression and cancer-embryonal antigen (CEA); the latter is combined with HLA class I and ICO-84 antigens at a significantly higher rate. The expression effect of CEA and HLA-DR on the metastatic spreading rate of regional lymph nodes is marked. A heterogeneity of the single tumour and in patients with malignant breast cancer is determined by the presence of the "leucocytic" antigens and CEA. In parallel studies monoclonal antibodies (MAB) HMFG-1 and a common epithelial antigen are assayed. The antigen interacting with MAB HEA-125 is characterized by the monomorphic expression.     

ICO-10 monoclonal antibodies to the Thy-1 antigen

Mouse monoclonal antibodies (MAB) ICO-10 to Thy-1 antigen were obtained. MAB ICO-10 reacted in indirect immunofluorescence test with 5.7 +/- 0.8% human thymocytes. Antibodies did not react with granulocytes, monocytes, T- and non-T cells from peripheral blood, and with marrow cells of healthy donors. MAB ICO-10 reacted with blast cells from 25 of 53 patients with T-cell acute lymphoblastic leukemia (ALL), from 2 of 5 patients with B-cell ALL. This antigen was absent on blood and marrow cells from some patients with ALL, 80 patients with chronic lymphoid leukemia, 54 patients with chronic granulocytic leukemia at the stage of blastic crisis, 128 patients with acute nonlymphoblastic leukemia. Antibodies are specifically bound to thymocytes and spleen cells of Thy 1.1 and Thy 1.2 mice. MAB ICO-10 detect Thy-1 antigen expressed on human hematopoietic cells. MAB ICO-10 may be applied for human leukemia and lymphoma immune diagnosis.     

Study of neuroblastomas in children using a complex of monoclonal antibodies

The possibility to reveal the following types of neuroblastoma is described: with c-ALL-antigen; with Thy-1-antigen (ICO-10); with granulocytic antigen (ICO-G2); with B-cell antigen (IPO-10). Immunocompetent cells in the tumour may be studied with the use of monoclonal antibodies ICO-1, ICO-11, ICO-GM-1, OKT3, IPO-3.     

ICO-35 monoclonal antibodies to the antigen of acute lymphoblastic leukemia

ICO-35 monoclonal antibodies (Mabs) were produced following BALB/c mouse immunization with peripheral blood cells from a patient with lymphoid type of chronic myeloid leukemia in blast crisis (SML BC) ICO-35. Mabs detect antigen on CD10-positive cells from patients with acute lymphoblastic leukemia, CML BC, CD10-positive cells of Reh line and are not bound to other cells. Comparative studies of reactivity of ICO-35 and K 503 Mabs to CD10 antigen revealed their similarity. However, in contrast to K503 ICO-35 Mabs do not react with granulocytes.     

ICO-1 monoclonal antibodies against the constant region of Ia-like (Dr) antigens

Mouse monoclonal antibodies ICO-1 to constant part of Ia-Like (Dr) antigens were produced. Hybridoma continuously produced IgG3 antibodies during 22 passages in vivo, more than 7 months in vitro. Antibodies specifically bound to 29.1 +/- 2.3% of peripheral blood mononuclear cells of healthy people recognized the antigen on B lymphocytes and 44.2 +/- 3.4% of monocytes. This antigen was absent on granulocytes and T lymphocytes. Using monoclonal antibodies ICO-1 antigenically positive cells were detected in 40 patients with B-CLL, in 12 of 35 patients (34.3%) with chronic granulocytic leukemia at the stage of blastic crisis, in 43 of 65 patients (66.1%) with ALL, in 14 of 38 patients (36.8%) with lymphosarcoma and in 17 of 30 patients with acute myelomonocytic leukemia. The antibodies responded to surface antigens in the reaction of indirect surface immunofluorescence, complement-dependent cytotoxic reaction and radioimmune tests.     

ICO-63 monoclonal antibodies to the differentiation antigen of hemopoietic cells suitable for immunophenotyping of human solid tumors

ICO-63, new monoclonal antibodies (MCAB), used for immunodiagnosis, were produced by the standard hybridoma technique ICO-63 MCAB reacted with 50% granulocytes, 20% thrombocytes and endothelial cells, whereas they did not react with peripheral blood mononuclear cells, thymocytes and erythrocytes. The molecular weight of ICO-63 MCAB-identified antigen was about 100 kD. ICO-63 MCAB reacted with 12 out of 15 neuroblastomas, with some solid tissue sarcomas and melanomas, but did not react with tumours of the epithelial origin.     

Block of NK cell activity by ICO-11 monoclonal antibodies

Antigen expression determined by ICO-11 monoclonal antibodies was studied on leukemia cells from bone marrow, blood cells from patients with lymphoproliferative diseases and natural killer (NK) cells responsible for natural resistance. ICO-11 monoclonal antibodies were shown to recognize the antigen expressed on NK-cells, predecessors of T-cells (thymocytes), myelomonocytes (myeloblasts, monoblasts) and to block NK-cell activity.     

Standardization of ICO-1 monoclonal antibodies against monomorphic Ia-like (Dr) antigens

ICO-1 Mab were obtained following BALB/c mouse immunization with 24-week human fetal thymocytes. Cloning for two times by the method of limited dilutions led to a hybridoma with a stable production of G3 isotype Mab. ICO-1 Mab have immunoprecipitated an antigen consisting of two polypeptide chains with molecular weight 29 and 34 kDal. The antigen expression was enhanced following PHA or alloantigen activation of blood mononuclear cells. ICO-1 Mab inhibited the alloantigenic response of blood mononuclear cells. Mab detected 29% of antigen-positive cells in the peripheral blood of healthy adults. When the reaction of ICO-1 Mab was compared with that of Mab against monomorphic Ia-like antigens OKLa, anti-Ia-BRL, BMA-021 and HLA-Dr on blood cells from healthy donors and patients with leukemia proved to be identical.     

The expression of antigens, detectable with ICO-series monoclonal antibodies, on the surfaces of cells forming granulocyte and macrophage colonies in semiliquid agar

The expression of antigens on granulocyte-macrophagal colony-forming cells of patients with nonhematological diseases was studied. Treatment of bone marrow cells with murine monoclonal antibodies ICO-1 and ICO-11 led to statistically significant inhibition of the number of growing colonies. Monoclonal antibodies ICO-02, ICO-10, ICO-GM-1 and ICO-G-2 had no such effect.     

Immunohistochemical analysis of S6K1 and S6K2 expression in human breast tumors

AIM: To express recombinant S6K2 in baculovirus expression system; to purify large quantities of recombinant S6K2 for biochemical studies; to generate and characterise specific MABs against recombinant S6K2; to study the patterns S6K1 and S6K2 expression and subcellular localization in normal, benign and malignant breast tissues. METHODS: Recombinant baculovirus, expressing wild type S6K2 was generated using Bac-to-Bac system (Invitrogen); recombinant S6K was purified from infected Sf9 cells using affinity purification approach; monoclonal antibodies against recombinant S6K2 were generated; the specificity of generated MABs towards recombinant and endogenous S6K2 were examined by ELISA, Western blotting, immunoprecipitation and immuhohistochemical staining; immunohistochemical detection of S6K1 and S6K2 in human breast tissues was performed using specific monoclonal antibodies towards S6K1 and S6K2. RESULTS: Large amounts of enzymatically active S6K2 were purified using baculovirus expression system; highly purified preparations of S6K2 were used to generate and characterize anti-S6K2 MABs; elevated levels of S6K1 and S6K2 were found in breast tumors when compared to normal breast tissues; S6K2 is frequently localized in the nuclei of adenocarcinoma tissues, but rarely in fibroadenoma or "normal" breast tissues. CONCLUSION: Production of recombinant S6K2 in large amount and generation of specific monoclonal antibodies towards S6K2 has provided us with excellent tools to study the function and regulation of this important signalling molecule in normal and cancer cells. Immunnohistochemical analysis of S6K1 and S6K2 expression in normal and malignant breast clearly indicates that both kinases are overexpressed in breast tumors, when compared to "normal" tissues. The retention of S6K2 in the nuclei of malignant cells may be caused by disregulation of nucleocytoplasmic shuttling and could subsequently affect cell growth and proliferation.     

Surface membrane differentiation antigens of the lymph node lymphocytes in nonspecific lymphoproliferative reactions and lymphogranulomatosis in children

Monoclonal antibodies ICO and OKT, OKM5, C-ALLA have been used to investigate surface lymphocyte antigens in normal (9), reactive (27) lymph nodes and in lymphogranulomatosis (9) in children. The study has shown that the number of lymphocytes with T and B markers is 45% in normal lymph nodes; OKT4/OKT8 is 0.9. In reactive lymph nodes all lymphocytes have T and B markers; OKT4/OKT8 is, 1.5. T-cell (OKT3+, OKT11+) or B-cell (sIg+, ICO-1+) type differentiation is preferable. B-cell type proliferation with a high number of ICO-1+ lymphocytes appeared in lymph nodes of 4 patients with lymphogranulomatosis. Polyclonal proliferation and differentiation were observed in other 5 children. No correlation between immunology and histologic types of the disease was revealed.     

IPO-10 monoclonal antibodies to the human B-lymphocyte differentiation antigen

Monoclonal antibodies IPO-10 were generated following immunization of a BALB/c mice with human cell line Daudi. Reactivity of this mABs was studied by indirect immunofluorescence technique with 10 human cell lines, blood cells of healthy persons and of patients with the malignant lymphoproliferative diseases. Studies on normal and neoplastic B cell suggest that mABs IPO-10 recognizing antigen is B lineage restricted. The antigen defined by the mAb IPO-10 appear to include most but not all stages of B cell differentiation.     

The expression of lymphocytic function-associated antigen determined by IKO-11 monoclonal antibodies in patients with nephroblastoma and neuroblastoma

The importance of NK-cells was shown in 12 patients with nephroblastoma and 10 patients with neuroblastoma using ICO-11 monoclonal antibodies. The numbers of circulating LFA-1+ cells were shown to be different in the untreated and treated patients. A certain regular influence of antitumour therapy on the amount of circulating LFA-1+ cells was established.     

Comparison of multiexcitation fluorescence and diffuse reflectance spectroscopy for the diagnosis of breast cancer (March 2003)

Nonmalignant (n = 36) and malignant (n = 20) tissue samples were obtained from breast cancer and breast reduction surgeries. These tissues were characterized using multiple excitation wavelength fluorescence spectroscopy and diffuse reflectance spectroscopy in the ultraviolet-visible wavelength range, immediately after excision. Spectra were then analyzed using principal component analysis (PCA) as a data reduction technique. PCA was performed on each fluorescence spectrum, as well as on the diffuse reflectance spectrum individually, to establish a set of principal components for each spectrum. A Wilcoxon rank-sum test was used to determine which principal components show statistically significant differences between malignant and nonmalignant tissues. Finally, a support vector machine (SVM) algorithm was utilized to classify the samples based on the diagnostically useful principal components. Cross-validation of this nonparametric algorithm was carried out to determine its classification accuracy in an unbiased manner. Multiexcitation fluorescence spectroscopy was successful in discriminating malignant and nonmalignant tissues, with a sensitivity and specificity of 70% and 92%, respectively. The sensitivity (30%) and specificity (78%) of diffuse reflectance spectroscopy alone was significantly lower. Combining fluorescence and diffuse reflectance spectra did not improve the classification accuracy of an algorithm based on fluorescence spectra alone. The fluorescence excitation-emission wavelengths identified as being diagnostic from the PCA-SVM algorithm suggest that the important fluorophores for breast cancer diagnosis are most likely tryptophan, NAD(P)H and flavoproteins.     

Monoclonal antibodies ICO-02 to blast cell antigens in patients with chronic myeloleukemia in blast crisis

Mice were immunized with blood cells of a patient with chronic granulocytic leukemia, and their cells were subsequently used for the preparation of hybridoma ICO-02. This hybridoma is continuously producing monoclonal antibodies which reacted with cells in 4 out of 13 patients with blastic crisis of chronic granulocytic leukemia and in 6 out of 38 patients with acute lymphoblastic leukemia. Antibodies reacted with blast cells in 2 out of 3 patients with undifferentiated blastic crisis of chronic myelocytic leukemia and in 2 out of 5 patients with lymphoid variant of blastic crisis of chronic granulocytic leukemia. Cells of 6 patients with acute lymphoblastic leukemia which reacted with the monoclonal antibodies had immunological markers of T lymphocytes bone-marrow precursors. Monoclonal antibodies did not react with cells of blood and bone marrow from healthy people and from patients with chronic lymphocytic leukemia, acute myeloblastic leukemia, acute myelomonocytic leukemia, acute monoblastic leukemia and lymphosarcoma.     

A tumor-associated antigen of cervical cancer

The tumour-associated antigen was identified with the aid of antisera obtained from rabbits immunized with 3 M KCl extract of pool human cervical carcinoma cells. The antigen was found in 92.5% specimens of human cervical squamous cell carcinoma, in 4.7% specimens of other localization of the tumours. The antigen was absent from sets of normal human tissues. The tumour-associated antigen was not identical with CEA, alpha-fetoprotein, SP1, EPA and lactoferrin and it was localized in cytoplasm of cervical carcinoma cells.     

Immunomorphological identification of an antigen associated with squamous cell carcinoma of the cervix uteri

Data on immunomorphological identification of antigen associated with cervical squamous cell carcinoma are presented. Tissue specimens prepared of cervical squamous cell carcinoma, normal cervical tissue and different other human tumours have been stained immunohistochemically for antigen using a polyclonal antiserum. Immunofluorescence staining for tumour-associated antigen was observed in all specimens of cervical squamous cell carcinoma and in 20% of specimens of the normal cervix uteri. The above antigen is a high specific tumour-associated marker and may be useful in the diagnostic studies.     

Electrical impedance spectroscopy of the breast: clinical imaging results in 26 subjects

Electrical impedance spectroscopy (EIS) is a potential, noninvasive technique to image women for breast cancer. Studies have shown characteristic frequency dispersions in the electrical conductivity and permittivity of malignant versus normal tissue. Using a multifrequency EIS system, we imaged the breasts of 26 women. All patients had mammograms ranked using the American College of Radiology (ACR) BIRADS system. Of the 51 individual breasts imaged, 38 were ACR 1 negative, six had ACR 4-5 suspicious lesions, and seven had ACR 2 benign findings such as fibroadenomas or calcifications. A radially translatable circular array of 16 Ag/AgCl electrodes was placed around the breast while the patient lay prone. We applied trigonometric voltage patterns at ten frequencies between 10 and 950 kHz. Anatomically coronal images were reconstructed from this data using nonlinear partial differential equation methods. Typically, ACR 1-rated breasts were interrogated in a single central plane whereas ACR 2-5-rated breasts were imaged in multiple planes covering the region of suspicion. In general, a characteristic homogeneous image emerged for mammographically normal cases while focal inhomogeneities were observed in images from women with malignancies. Using a specific visual criterion, EIS images identified 83% of the ACR 4-5 lesions while 67% were detected using a numerical criterion. Overall, multifrequency electrical impedance imaging appears promising for detecting breast malignancies, but improvements must be made before the method reaches its full potential.     

Localization of embryonic prealbumin-1 in the tissues of human malignant tumors

The distribution of human embryonic prealbumin-1 (EPA-1) has been studied in 52 different malignant tumours using indirect immunofluorescent technique and two types of antibodies to different antigenic determinants: "EPA-1" and "EPA-1S". In mesenchymal tumours (except for desmoids and chondrosarcomas) this glycoproteid was observed in the cytoplasm of tumour cells, but "EPA-1S"--in the connective-tissue fibers only. Localization of glycoproteid EPA-1 depends on the type of the used primary antibodies. EPA-1 is also observed in the connective-tissue stroma of most of epithelial tumours. A conclusion is made that transformation of EPA-1 glycoproteid synthesis from definitive to fetal form occurs in malignant tumours.     

Human leukemias studied with monoclonal antibodies to erythroid cells

Activity of monoclonal antibodies HAE3 and HAE9 specific for human erythroid cells to different leukemic cells is described. These monoclonals do not react with nonerythroid leukemic cells. HAE3 and HAE9 reactivities are similar to those of polyclonal monospecific antibodies against an antigen of erythroblasts--a surface antigenic marker of nucleated red cells and reticulocytes. Monoclonal antibodies HAE3 and HAE9 are proposed to be used in diagnosis of leukemias.