Phenotypic expression of histocompatibility antigens in human primary tumours and metastases.

HLA class I and II expression was studied on 244 (177 primary and 67 metastatic) solid human tumours of different origin. Alkaline immunophosphatase (APAAP) and immunoperoxidase were used on cryostatic sections to stain MHC antigens. Monomorphic MoAbs were used against class I heavy chain, beta 2-microglobulin, DR, DQ and DP molecules. Class I expression was homogeneous on colon, melanoma and epidermoidal primitive tumours. Loss of HLA class I antigens was more frequent on basal cell carcinomas and sarcomas and was related to tumour differentiation on larynx carcinoma. Class I expression was heterogeneous on breast, larynx and stomach primitive neoplasias. Class I negative tumours were more frequent on metastatic than on primitive melanomas. Divergence of class I between primary tumours and autologous metastases was observed on melanomas, larynx and colorectal carcinomas. Class II expression was heterogeneous on all tumours and in a large number of cases was associated with high intensity of leukocytic infiltrate. HLA-DR expression was higher than HLA-DP and HLA-DQ (DR greater than DP greater than DQ) and was related to tumour progression. Four human tumour cell lines were modulated with recombinant interferon-gamma for HLA class I and II antigens. Different HLA profiles were obtained: increased class I and II expression, increased class II or a low response. Finally, class I genes from 22 tumours were compared with autologous normal cells by Southern blot analysis: 12 tumours were class I positive and 10 negative. No clear differences in RFLP were observed that could be associated with class I rearrangement. The results are discussed in relation to the role that histocompatibility antigens may play in tumour progression and invasiveness.

Histocompatibility antigens in primary and metastatic squamous cell carcinoma of the larynx.

A series of 38 primary laryngeal and hypopharyngeal tumours, 15 lymph-node metastases and normal tissue were evaluated in frozen sections for the expression of MHC class I and II antigens, using monomorphic monoclonal antibodies (MAbs) to HLA-ABC, beta 2-microglobulin, DR, DP, DQ, HLA-B and polymorphic HLA-ABC antigens. Normal distant mucosa of larynx reacted to anti-class I antibodies but not to anti-class II. In 9 primary tumours (23.7%) HLA class I antigens were not observed. The remaining 29 showed a strong reaction to not observed. The remaining 29 showed a strong reaction to anti-HLA-ABC (heavy chain) and anti-beta 2-microglobulin, although in 3 cases out of 29 no staining was observed with anti-HLA-B locus-specific MAbs. These selective losses were confirmed using the corresponding anti-HLA polymorphic MAbs. For HLA class II molecules, only DR was observed in 3 of 38 cases. Defective HLA class I expression statistically correlates with high scores according to Jakobsson's criteria for histopathological tumour grading. Loss of HLA-ABC antigens was most frequent among the cases with poor differentiation (6/8 cases). On the contrary, class II antigen expression was correlated with a well differentiated pattern and a more favourable prognosis (p less than 0.001). We have found differences in HLA class I expression when comparing primary tumours and autologous metastases (3/9 cases). Immunoprecipitation and SDS-PAGE of class I antigens, Northern and Southern blot analyses of MHC class I genes were performed. We have not detected class I gene rearrangement using HLA coding and locus-specific non-coding probes. However, we have found a class I transcription defect that corresponds with a class-I-negative phenotype.

Different CD45 and CD45R epitopes involved in T cell proliferation and NK cytotoxicity.

Different monoclonal antibodies detecting the leucocyte common antigen have been obtained, for CD45 (GRT2, GRT3 and GRT4) and for CD45R (GRT22). Several epitopes have been defined with these monoclonal antibodies (MAbs). We have performed a comparative study with CD45 and CD45R MAbs on NK and T cell proliferation. Common epitopes of CD45 antigen were found to be involved in blocking of NK activity but not CD45R restricted determinants. In the cell proliferation assays, fresh human peripheral blood mononuclear cells were stimulated with PHA and soluble CD3 MAbs. CD45 and CD45R MAbs failed to demonstrate the capacity to modify the mitogenic response when optimal and suboptimal doses of PHA were used. In contrast, both (CD45 and CD45R) MAbs led to a significant rise in anti CD3 response. A CD18 (GRF1) was used as control and inhibited both PHA and CD3 T cell proliferation as well as NK activity. We think these results can be explained on the basis of different activation pathways (PHA versus anti CD3) and the accessory signals induced by these MAbs as recorded only in anti CD3 induced mitogenesis.

Studies on CD11a and CD18 molecules with two new monoclonal antibodies: differential myelomonocytic antigen expression of PMA treated HL60 and U937 cell lines.

Biochemical and functional aspects as well as features of cellular distribution of the differentiation groups CD11a and CD18 were studied in the course of a detailed characterization of two new monoclonal antibodies which recognize the alpha (GRS3) and beta (GRF1) chains of the LFA-1 antigen. Both MAbs inhibited homotypic aggregation of an EBV cell line. In contrast, only GRF1 (anti-beta chain) was able to inhibit granulocyte aggregation as well. Different myeloid-monocyte antigen modulation was noted in PMA induced macrophage differentiation of the U937 and HL60 cell lines. PMA treated HL60 cells showed increased expression of alpha M (CD11b) and alpha X (CD11c) antigens but no change in HLA-DR or CD14 antigen expression. No variation in the expression of LFA complex antigen (CD11a, b and c, or CD18) was observed on U937 cells, which on the other hand presented de novo expression of HLA-DR and CD14 antigens.

HLA class I gene expression on human primary tumours and autologous metastases: demonstration of selective losses of HLA antigens on colorectal, gastric and laryngeal carcinomas.

The expression of HLA class I antigens was studied in 99 primary tumour (colorectal, gastric and laryngeal carcinomas) and 57 autologous metastases using immunohistological techniques and monoclonal antibodies against class I monomorphic determinants, HLA-B isotypic determinants and HLA polymorphic determinants. Fourteen per cent of colorectal, 9.6% of gastric and 20% of laryngeal carcinomas completely lacked class I molecules. Selective losses of HLA-B antigens were also detected in 8.8, 3.4 and 5.8% of these tumours respectively. Taking into account complete and selective loss of HLA-B the average alteration in the class I molecules expression totalled 21%. The comparison of class I expression between primary tumours and autologous metastases showed differences in 24% of the patients. These differences consisted mainly in a decrease of class I expression by metastases. Nevertheless, four types of divergence were detected in laryngeal carcinomas, namely: +/-, +/+, -/+, -/-. In addition, a clear correlation between degree of differentiation and class I expression was observed in laryngeal tumours. Finally, when class I gene RFLPs were compared with DNA from 15 tumours and autologous normal mucosa or peripheral lymphocytes, no differences were detected between these samples.

Distribution of the CD45R antigen in the maturation of lymphoid and myeloid series: the CD45R negative phenotype is a constant finding in T CD4 positive lymphoproliferative disorders.

We have compared the expression of the common leucocyte antigen (CD45) and the restricted leucocyte antigen (CD45R) on normal haematopoietic cells, cell lines, and a total number of 136 cases of myeloid and lymphoid proliferative syndromes. CD45, the conventional leucocyte antigen, presents a generalized distribution along the lymphoid and myeloid maturation pathway with the exception of some myelomas and pre-B leukaemias. In contrast, the expression of the CD45R determinant is more limited. Although it is found in the majority of the differentiation stages of B cells and monocytes, it is present only in the early stages of myeloid differentiation. On T cells it is expressed on mature thymocytes and in the majority of CD8+ lymphocytes and a subset of CD4+ cells on peripheral blood. Finally, our results also indicated that CD4+ T lymphoproliferative syndromes are derived from the CD4+ CD45R- subset (20/20 cases).

A monoclonal antibody, GR7A4, reacting with the T10 antigen.

The monoclonal antibody GR7A4 was produced against a human lymphoblastic leukemia (ALL-B). Biochemical characterization of GR7A4 was carried out with Raji cells in SDS-PAGE studies. GR7A4 precipitated a 43 kd molecule. The tissue distribution and molecular weight were similar to the T10 antigen. Modulation and capping of GR7A4 antigen reduced its binding ability to cells with either GR7A4 or OKT10. However, the cell surface distribution pattern observed was somewhat different from other similar monoclonal antibodies. Thus GR7A4 reacted greatly with pre-B-cell lines, Burkitt cell lines, EBV cell lines and activated PHA, ConA and PWM lymphocytes, however reactivity with leukemic cells was very limited. The kinetics of appearance of GR7A4 antigen on PWM blasts show that this molecule seems to represent an intermediate stage in lymphocytic activation. The differences in comparison with other similar MAbs are discussed and correlated with the peculiar discontinous pattern of appearance of this antigen.