Isoenzyme studies in human leukemia — III. β-hexosaminidase (E.C. 3.2.1.30)

Enzymologic profiles of β-hexosaminidase (N-acetyl-β-d-glucosaminidase, E.C.3.2.1.30) were studied in cells from childhood acute leukemias and lymphomas. By analytical isoelectric focusing or disc electrophoresis the β-hexosaminidase activity was separated into its components A, B, I and C. The isoenzyme patterns were correlated with immunologic cell surface marker characteristics found on the investigated leukemic cells. In all cases of T-ALL the β-hexosaminidase forms A and B were observed, an enzyme pattern similar to that found in normal lymphocytes. Seven out of 11 cases with cALL, three of six cases with AML and one case of AUL displayed the intermediate component (Hex I). Marked heterogeneity within the immunologically classified subgroup cALL was reflected in different enzyme patterns of the cALL samples. These biochemical phenotypes may indicate the different maturation and differentiation status of cells expressing the same immunologic surface markers.     

The use of enzyme marker analysis for subclassification of acute lymphocytic leukemia in childhood

Subclassification of acute lymphocytic leukemias in childhood by multiple marker analysis has proven the heterogeneity of this disease and this methodology has led to a better understanding of the cell-biological basis of ALL. Enzyme markers have become important tools in multiple marker analysis. This is especially true for TdT, purine metabolic enzymes, hexosaminidase I, acid phosphatase and carboxylic esterases. In spite of sophisticated methods and encouraging results multiple marker analysis has not been totally satisfactory in defining patients at risk. The same is true for a risk score established by clinical data. More efforts in the future are necessary for combining multiple marker analysis, cytogenetics, proliferation characteristics, basic clinical findings and the final outcome of the disease in these patients. Beyond that this kind of leukemia research will help to clarify the pathobiological basis of human leukemia and to develop new specific therapeutic modalities.     

Ultrastructural analysis of human plasmacytoma cells prepared on affinity beads after exposure to anti-idiotype antibodies

Mononuclear cells from a bone marrow infiltrated by plasmacytoma cells were examined by electron microscopy. An analysis was performed according to different cytoarchitectural forms of the endoplasmic reticulum (ER). Six types of plasma cells could be distinguished. The bone marrow cells were treated with an anti-idiotype antiserum from a guinea pig prepared against the patient's monoclonal serum protein and with a FITC conjugated anti-guinea pig antiserum from the rabbit as second layer. Then the cells were passed through an affinity gel column with anti-FITC antibodies. The original preparation and the cells separated on the affinity gel were analysed by electron microscopy. It was found that an ultrastructurally distinct type of plasma cell was enriched 3.5-fold over the original sample by the separation procedure.     

Chronic lymphocytic leukemia: A proliferative or accumulative disorder?

In two untreated patients with progressive CLL, quantitative ¹⁴C autoradiography of lymph nodes and, subsequently, continuous infusion of [³H] thymidine over eight and nine days, respectively, were performed in order to analyse the lymph node cell kinetics. Simultaneously, the turnover of labelled lymphocytes in the peripheral blood was evaluated. From another CLL patient a regional lymph node was removed 6 h after an intralymphatic flash injection of [³H] thymidine and sectioned for autoradiographic study of the distribution of labelled cells within the lymph node tissue. While the durations of DNA synthesis were found to be normal, the labelling indices were reduced. The relative cell production rate was far lower than normal. Very small growth fractions were calculated, amounting to less than 1% in one patient, and to 2.4% in the other. The distribution of labelled cells in the lymph nodes was focal, which supports the finding of low growth fractions. According to the present data, CLL is a disorder in which a very small number of cells cycle at a roughly normal rate. A kinetic definition of accumulative and proliferative tumour growth is introduced. Tumour growth is termed accumulative if growth appears to result from a decreased relative cell loss rate rather than an increased relative cell production rate. According to this definition, the kinetics in CLL may be classified as accumulative. In absolute terms, however, the number of lymphoid cells produced per unit of time was found to be far higher in CLL than in the healthy state.     

A monoclonal antibody (SJ-9A4) to P24 present on common alls, neuroblastomas and platelets - II. Characterization of P24 and shedding in vitro and in vivo

The release of soluble P24 antigen into culture medium by common acute lymphoblastic leukemia (C-ALL) and neuroblastoma (NB) cell lines was studied. P24 release by C-ALL cells was detected using a solid phase indirect radioimmunometric assay (IRA) which combines the specificity of lectins and monoclonal antibodies (MoAb) and using immunoadsorption of labeled P24 in spent medium from cells incubated with 35S-methionine (met). No P24 was present in the medium of cells pulse labeled at 37 degrees C when they were placed at 4 degrees C, thus this is an active process. P24 release by NB cells could not be detected by IRA, but could be detected by immunoadsorption of spent medium of metabolically-labeled cells. The absence of IRA activity of P24 from NB spent medium was due to decreased glycosylation and thus no binding to the lectins employed in the IRA was observed. This was confirmed by lectin affinity chromatography which showed that P24 in the spent medium from C-ALL cells bound Ricinus communis agglutinin (RCA1), wheat germ agglutinin (WGA), concanavalin A (Con A), and lentil lectin (LcH), but not peanut agglutinin (PNA). P24 from NB cell spent medium did not bind to any of these lectins. The lectin affinity of P24 derived from lymphoblasts is consistent with the presence of N-linked oligosaccharide chains having N-acetyl glucosamine residues, a mannose core, and a terminal D-galactose. P24 from C-ALL cell spent medium was present in the 35-45% fraction of a saturated ammonium sulfate (SAS) partition of spent medium. The P24 antigen was detected in the fractionated plasma of five patients with C-ALL at the time of diagnosis and was undetectable when the patients had achieved a complete remission. Plasma from 2 patients with P24 negative ALL, normal human plasma, and normal human serum had no detectable activity.