Effect of immunotherapy on lymphocyte function in acute lymphatic leukemia]

During the immunotherapy children suffered from acute leukaemias will have a significantly higher transformation rate than at the beginning of the immunotherapy. This may be explained by an increase of the immunological competence as well as by an enhanced mobilization of lymphatic cells. Leukaemic blasts used for immunoinduction-therapy will have no higher transformation rates as antigens than those cells never contacted by children. During the immunotherapy an increase of transformation rates may be observed after administering unspecific antigens and in mixed cultures. In a retrospective manner the indication for immunotherapy may be checked again in children with immunotherapy on the basis of the clinical course and evaluation of the cellular immunoreaction.     

Leukemic chromosome clone. I. Risk factor in the acute lymphatic leukemia of children]

As long as the aetiology of acute lymphatic leukaemia of children is not known its therapy is based on clinical experience. Among the values of experience those factors will play a part, the evidence of which during the ALL initial stage will be a risk for successful therapy and survival rate. This results in a choice of more aggressive variants of modern therapy schemes. In a cytogenetic study made in 35 children with ALL it was tested, whether even leukaemic chromosome clones will be a risk for the course of acute leukaemia. The duration of the first remission and survival rate were considered as criteria. The evidence of a leukaemic chromosome clone could be shown to be followed by a short survival rate, irrespective of the stage of the disease where the clone had been observed first. Thus, cytostatic therapy in those ALL patients who are affected with luekaemic chromosome aberration of stem line character should be aimed at the complete annihilation of the clone, irrespective of other remission criteria. The failure of blood and bone-marrow cultures as early as during the untreated initial stage indicated a primary cellular immuno-insufficiency. This combination of cell immuno-depression with high peripheral leukocytes connts and a primary mediastinal tumour or a generalizing lymphosarcoma respectively, was the highest risk up till now for the course of the disease. Judging from the duration of the first remission and the survival rates, the consecutive schemes of therapy did not differ in their effect on leukaemia with pathological stem lines. On the basis of the present study the impression could not be excluded that up till now long term survival rates could be attributed rather to individual manners of response than to the modern therapy scheme.     

Lymphoblastic lymphoma in a female with chronic granulocytic leukaemia

In a 18 years old, Ph1 negative woman a lymphoblastic lymphoma developed after 16 months of treatment for juvenile form of CGL. In the course of the disease we observed twice the myeloblastic transformation and once the low differentiated transformation of CGL. Based on available methods we are not in a position to exclude a particular form of lymphoblastic transformation of CGL involving lymph nodes without leukaemic blood picture.     

Studies on the effect of various kinds of cytostatic therapy on the cellular immune reaction in children with acute lymphatic leukemia]

Clinical and experimental findings on possible changes of the lymphocyte function during an immunosuppressive or cytostatic therapy respectively caused investigations to be made for explaining the connections existing between the influence of cellular immunoreaction and the use of different cytostatic regimes. Earlier findings on the influence of cellular immunoreaction after adding cytostatics to cultivated cells and investigations on the influence of the lymphocyte function in dependence on cytostatic therapy were used for comparison. Transformation and mitosis rates as well as necrosis rates and the result of macrophage migration inhibition are comparable parameters for influencing the lymphocyte function in children treated with cytostatics. Antimetabolites, vincristine, asparaginase and daunomycin will have less influence on the transformation rate as an expression of an immunosuppressive effect on only those cells responding in accordance with their kinetic phase. Cyclophosphamide will inhibit the transformation reaction more significantly. Examinations in children with different therapeutic regimes reveal a certain validity of therapy after the first statistical evaluation of the clinical material.     

Granulocytopoiesis in children with acute lymphoblastic leukaemia.

Numbers, proliferative potential, and differentiative capacity of bone marrow granulocyte-macrophage precursor cells were studied in 130 children with acute lymphoblastic leukaemia (ALL), including 77 children in an acute phase of the disease and 53 in remission. Bone marrow samples from 65 children without haematopoietic abnormalities were used as controls. The numbers of clonogenic precursors were found to be below normal in all phases of ALL, particularly during the acute period when the bone marrow was heavily infiltrated with leukaemic cells. It is shown that the decreases in the numbers and proliferative potential of the precursor cells during the acute phases was associated with the effects of leukaemic blast cells, but that in remission the observed reduction in the precursor cell pool was due to the cytostatic effect of therapy. The differentiative capacity of clonogenic granulocyte and macrophage precursors was not altered in children with ALL.     

The therapeutic potential of stem cells in heart disease

Abstract.  Coronary heart disease and chronic heart failure are common and have an increasing frequency. Although interventional and conventional drug therapy may delay ventricular remodelling, there is no basic therapeutic regime available for preventing or even reversing this process. Chronic coronary artery disease and heart failure impairs quality of life and are associated with subsequent worsening of the cardiac pump function. Numerous studies within the past few years have been demonstrated, that the intracoronary stem cell therapy has to be considered as a safe therapeutic procedure in heart disease, when destroyed and/or compromised heart muscle must be regenerated. This kind of cell therapy with autologous bone marrow cells is completely justified ethically, except for the small numbers of patients with direct or indirect bone marrow disease (e.g. myeloma, leukaemic infiltration) in whom there would be lesions of mononuclear cells. Several preclinical as well as clinical trials have shown that transplantation of autologous bone marrow cells or precursor cells improved cardiac function after myocardial infarction and in chronic coronary heart disease. The age of infarction seems to be irrelevant to regenerative potency of stem cells, since stem cells therapy in old infarctions (many years old) is almost equally effective in comparison to previous infarcts. Further indications are non-ischemic cardiomyopathy (dilative cardiomyopathy) and heart failure due to hypertensive heart disease.     

A case of erythrophagocytic T4 lymphoma. Immunological and morphological peculiarities.

One case of T-cell lymphoma with atypical malignant cells is reported. Some of the clinical features, morphological characteristics and functional activity (erythrophagocytosis) of malignant cells suggested malignant histiocytosis. The malignant disease started with splenomegaly and developed with hepatomegaly, bone marrow infiltration, discrete lymphadenopathy and leukaemic picture. Proliferated cells were characterized by ambiguity. In addition to phagocytic capability, presence of complement receptors and ultrastructural features proper to the macrophagic lineage, the cells expressed T-cell determinants (E receptors, T3, T4 and T11 antigens) and were peroxidase and esterase-negative. Erythrocytes were partially or completely dehaemoglobinized and presented the phenomenon of autolysis in different stages of development. Because this lymphoma is difficult to diagnose and apparently resistant to therapy, its recognition and further study are warranted.     

Interferon therapy in acute lymphatic leukemia and hepatitis B in childhood

1. The use of interferon in hepatitis B will bring about a favourable course of the disease, which is reflected in a normalisation of liver function tests. 2. The concentration of surface antigens is distinctly reduced, with a lower titre remaining for a longer period of time. 3. The level of the dosage chosen seems to be optimal. However, treatment has to be continued for a period of several months, probably for about 1 year. 4. In childhood the side-effects of interferon are minimal, they are restricted to febrile reactions. 5. In our investigations no sure hematological deviations could be identified in the treatment with interferon. 6. The study must be complemented by analysing further antigens, such as E antigen, allowing the course of disease to be characterised more distinctly. 7. In order to give a final assessment of the therapy the observation of patients must be continued after terminating the treatment, the histological improvement in the further course having to be confirmed by liver biopsies.     

Differences in electrophoretic mobility and cytochemistry of leukemic T-cells with identical immunologic phenotype (CD1 positive)

Leukaemic cells of acute T-lymphoblastic leukaemia and T-lymphoblastic lymphoma with secondary leukaemic course of disease which showed the same immunological phenotype (cortical thymocytes, thymocyte stage II), could be further differentiated according to their cytochemical pattern and electrophoretic mobility (EPM). Leukaemic cells of T-ALL usually reveal a high EPM, whereas EPM of leukaemic cells of T-lymphoblastic lymphoma was significantly lowered. In cytochemical respect acid phosphatase was positive in both cases of leukaemia. An activity of acid esterase, however, could only be demonstrated in leukaemic cells of T-lymphoblastic lymphoma. The findings are interpreted as manifestation of a different stage of maturity of both T-cell clones with the same immunological phenotype.     

Cell-mediated immunity in Cryptococcosis

Cell-mediated immune responses in patients who had recovered from cryptococcosis were compared to those of healthy subjects. Cryptococcal patients were mildly lymphopenic but showed no defect in percentage of thymus-derived lymphocytes. One-third had positive delayed skin test reactions to cryptococcal antigen. Their skin test reactivity to two commonly used noncryptococcal antigens was less intense than healthy control subjects. Strongly positive and specific lymphocyte transformation occurred in the presence of an extract of Cryptococcus neoformans (cryptococcin) in half of the patients. In contrast, few healthy subjects had positive transformation responses to cryptococcin. One patient who was followed sequentially through treatment of cryptococcal meningitis acquired strong cryptococcin reactivity during the course of treatment. Cellular immunologic response to cryptococcin identifies many subjects who have had C. neoformans exposure, and may be of value for assessing immunologic status of patients undergoing therapy. These studies also indicate that most patients with cryptococcosis have a degree of deficiency in cell-mediated response to fungal antigens even when a specific underlying disease process cannot be identified.     

Functional cell compartments in a rat model for human acute myelocytic leukaemia.

Functional cell compartments were studied in a rat model for human acute myelocytic leukaemia (AML). This was done by tracing the distribution of injected 51Chromium-labelled leukaemic cells in the body. It was concluded that two functional compartments can be distinguished in acute leukaemia, i.e., a rapidly exchangeable pool of cells (including the circulating blood pool, the marginal noncirculating blood pool and the rapidly exchangeable tissue pool; RETP) and a slowly exchangeable tissue pool (SETP). The sizes of these various compartments were roughly quantified at various stages of the disease by calculations based on the principle of isotope dilution and organ weight measurements. As the leukaemia progresses, the size of the SETP increased significantly relative to the size of the RETP. Simultaneously, the exchange rates of leukaemic cells between the organs and the blood decrease. The blood transit time of leukaemic cells was also significantly prolonged, as is the case in human AML.     

An in vitro testing of chemoresistance in leukaemic patients

The fact that leukaemic cells are primarily or secondarily resistant to cytostatics is a serious phenomenon, which leads to the failure of chemotherapy of malignant diseases in clinical practise. Some detoxification and transporting systems are responsible for the generation of chemoresistance on the cellular level and the decrease of effectiveness in treatment. In vitro testing of chemoresistance of leukaemic cells is presently an inseparable component of “tailoring” therapy in the developing field of predictive oncology. The aim of this work was to estimate profiles of drug resistance, based on the predictive in vitro test, and to help in choosing the most effective cytostatic. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazoline (MTT) assay was used, based on the direct effect of cytostatics on the viability of leukaemic cells in vitro. The number of living leukaemic cells was evaluated by a computer program, where LC₅₀ (concentration of cytostatics lethal to 50% of leukaemic cells) was established from the achieved dose-relation curves. Seventy-one samples of leukaemic cells isolated from the patients’ peripheral blood or bone marrow were examined. All samples were tested to 3 cytostatics minimally. It was found by the in vitro assay, that resistance to dexamethasone, prednisolone, etoposide and vincristine is increased in patients with acute myeloid leukaemia disease, compared to the acute lymphoblastic leukaemia patients. In patients with a relapsed disease population, leukaemic cells are highly heterogeneous in the MTT assay. It was concluded that the MTT assay can be used to study drug interactions in vitro in leukaemia samples. The type of interaction was highly different between patients, and depended on drug concentrations.     

Autologous responses to human leukaemic cells in mixed leucocyte culture

Summary Cryopreserved leukaemic blasts and remission non-T cells from 22 patients with acute leukaemia (15 lymphocytic, 7 non-lymphocytic) were tested as stimulators of autologous remission T cells and normal allogeneic T cells in primary and secondary MLC. In most cases the autologous response elicited by leukaemic cells was less than or equal to that elicited by remission non-T cells. However, T cells from 2 patients in long-standing first remission from ANLL displayed greater proliferation in response to leukaemic blasts than to remission non-T cells in both primary and secondary MLC. The results are suggestive of sensitization of these 2 patients to leukaemia-specific antigens, but other possible explanations are discussed. Abbreviations used: MLC, mixed leucocyte culture; ANLL, acute non-lymphocytic leukaemia; ALL, acute lymphoblastic leukaemia; AMLR, autologous mixed lymphocyte reaction; NK cells, natural killer cells; MNC, mononuclear cells     

FUNCTIONAL CELL COMPARTMENTS IN A RAT MODEL FOR HUMAN ACUTE MYELOCYTIC LEUKAEMIA

Functional cell compartments were studied in a rat model for human acute myelocytic leukaemia (AML). This was done by tracing the distribution of injected ⁵¹Chromium-labelled leukaemic cells in the body. It was concluded that two functional compartments can be distinguished in acute leukaemia, i.e., a rapidly exchangeable pool of cells (including the circulating blood pool, the marginal noncirculating blood pool and the rapidly exchangeable tissue pool; RETP) and a slowly exchangeable tissue pool (SETP). The sizes of these various compartments were roughly quantified at various stages of the disease by calculations based on the principle of isotope dilution and organ weight measurements. As the leukaemia progresses, the size of the SETP increases significantly relative to the size of the RETP. Simultaneously, the exchange rates of leukaemic cells between the organs and the blood decrease. The blood transit time of leukaemic cells was also significantly prolonged, as is the case in human AML.     

Factors in Pathogenesis of Central-nervous-system Leukaemia

Eighty-three (50%) of 165 children with acute lymphoblastic or acute stem-cell leukaemia presenting during 1958-70 developed leukaemia of the central nervous system (C.N.S.). The rate of incidence of this complication is fairly constant throughout the first two-and-a-half years of the disease, but falls thereafter. The incidence of C.N.S. leukaemia is inversely correlated with the platelet count at the time of initial diagnosis of leukaemia, and directly correlated with the total leucocyte count and the presence of lymph-node enlargement. The major effect of initial leucocyte count is on the time of onset of clinical symptoms. It is suggested that leukaemic cells usually enter the C.N.S. from the blood as a result of intracranial petechial haemorrhage occurring around the time of initial diagnosis of leukaemia, and that the time for subsequent development of symptoms of C.N.S. disease is largely determined by the number and replication rate of leukaemic cells which gain access to the C.N.S. at that time. The increasing frequency of diagnosis of C.N.S. leukaemia in recent years is not wholly explained by increasing survival, and may in part be related to changes in the pattern of antileukaemic therapy.Prophylaxis for C.N.S. leukaemia should be instituted as early as practicable after diagnosis; the identification of a high-risk group may permit this to be done selectively.     

T and B cells in B-chronic lymphocytic leukaemia: Faust, Mephistopheles and the pact with the Devil

A large number of human malignancies are associated with decreased numbers of circulating T cells. B-CLL, in this regard, represents an anomaly since there is not only high numbers of circulating B cells, characteristic of the malignancy, but also a massive expansion of both CD4 and CD8 T cells. These T cells for the most part may probably not represent a leukaemia-specific TCR-dependent expansion. On the contrary, these T cells, especially the CD4 subset, might support a "microenvironment" sustaining the growth of the leukaemic B cell clone. Conversely, the leukaemic B cells may produce membrane-bound as well as soluble factors that stimulate the proliferation of these T cells in an antigen independent manner. In addition to these T cells lacking anti-leukaemic reactivity, there exist spontaneously occurring leukaemia-specific T cells recognizing several leukaemia-associated antigens, e.g. the tumour derived idiotype, survivin and telomerase. Both CD4 and CD8 leukaemia-specific T cells have been identified using proliferation and gamma-IFN assays. These reactive T cells can lyse autologous tumour cells in an MHC class I and II restricted manner. Spontaneously occurring leukaemia-specific T cells are more frequently noted at an indolent stage rather than in progressive disease. Preliminary results from vaccination trials using whole tumour cell preparations as vaccine have demonstrated that vaccination may induce a leukaemia-specific T cell response, which might be associated with clinical benefits. Extended clinical trials are required to establish the therapeutic effects of vaccination in B-CLL. Studies in our laboratory as well as those of others indicate that whole tumour cell antigen in the form of apoptotic bodies or RNA loaded on to dendritic cells may be a suitable vaccine candidate. Patients with low stage disease may maximally benefit from this form of therapy.     

The spontaneous lymphocyte transformation rate in newborn infants at risk]

The lymphocyte transformation test was performed in 32 risk newborns and 32 comparative persons (17 adults, 15 children). In the void controls without antigen addition the newborns had a significantly higher spontaneous blastic reaction than the control group. The phenomenon may be explained by lymphatic stem cells in the blood or an ontogenetically higher content of "embryonic tissue" respectively being present as unspecific stimulant or an immunological defence reaction against maternal immunoglobulins transmitted diaplacentally (formation of antigammaglobulin factors) or against maternal lymphocytes to prevent a "runt disease".     

Sickle cell disease in the People's Republic of Angola. 2. Behavior of laboratory parameters

In 819 children the Hb concentration, its dependence on the course of disease, the impact of various crises on Hb values and the behaviour of MCHC were evaluated. In the mean there was an anemia of 4.65 +/- 0.5 Hb mmol/l. In the course of 6 years a significant decline anemia of 4.65 +/- 0.5 Hb mmol/l. In the course of 6 years a significant decline of the Hb concentration could be observed. Only those crises connected with severe clinical symptoms (third degree) coincide with a significant Hb decrease. Every second child with a mean Hb value of 2.3 mml/l was transfused. In nearly half the cases there is a normochromic anemia, in 45.7% a hypochromic one. The mean serum bilirubin concentration lay at 31.6 mumol/l and increased with growing age of disease. The anemia has negative effects on the cardio-circulating system. The thorax-heart quotient shows a positive correlation to the degree of anemia.     

Presence of herpes simplex virus-related antigens in transformed L cells.

Antiserum prepared against herpes simplex virus type 1 (HSV-1)-infected L cells, i.e., lytic antiserum, was shown by an indirect immunofluorescence test to stain 90 percent of HSV-transformed L or HeLa cells. Immunofluorescence in these cells was always most intense in the perinuclear cytoplasmic region. Similar results were obtained with antiserum prepared against HSV-transformed L cells. These data indicate that HSV-transformed cells (both L and HeLa) express HSV-related antigens. Antiserum prepared against HSV-1-transformed L cells, i.e., transformed-cell antiserum, was found to agglutinate purified HSV type 1 virions but failed to neutralize infectivity. This suggests that HSV-1 structural antigens are expressed in HSV-1-transformed L cells. Immunodiffusion studies showed that at least two HSV-related antigens could be demonstrated with antigens from HSV-1-transformed L cells and transformed-cell antiserum. These two antigens were shown to be present in all clonal lines of HSV-1-transformed cells examined, six L cell lines and one HeLa cell line. Therefore, we conclude that transformation of cells by HSV-1, which is known to be associated with acquisition of viral thymidine kinase, must also be associated with the presence of these two antigens. We performed experiments showing that there are species of HSV-related antibody in HSV-transformed cell antiserum that could not be absorbed out with antigens from HSV-infected L cells. Antibodies present in lytic antiserum were completely removed by antigen preparations from cells lytically infected with HSV-1. Also, lytic antiserum failed to block HSV-related staining of transformed L cells in a direct immunofluorescence test. These results are compatible with one of two notions: either (i) certain genes are expressed during transformation that are not expressed during lytic infection, or (ii) these genes are expressed to a much more reduced extent during lytic infection than in transformed cells.     

Specific features of diphtheria vaccinal process in children with rheumatic diseases

Clinical and immunological examination of 55 children aged 6-15 years with rheumatic diseases, immunized against diphtheria, was carried out. All children were immunized at the stage of clinical and laboratory remission and in some cases while undergoing a prolonged course of cytostatic therapy or therapy with nonsteroid anti-inflammatory remedies. This examination demonstrated that in the overwhelming majority of children with rheumatic diseases the diphtheria vaccinal process took an asymptomatic course and had no influence on the course of the main disease. Specific features, characteristic of the immune status of this group of children, were established. In the course of the vaccinal process the restoration of the initially inhibited characteristics (the production of TNF-alpha and IL-2) to normal values were shown to occur, which was indicative of the fact that the reserve capacities of immunocompetent cells were retained in these patients. This study also revealed that immunization of children with rheumatic diseases with adsorbed DT and D toxoids with reduced antigen content was not excessive antigenic stimulation for such children, as it did not lead to immunopathological shifts, but induced transient phase changes in immunological characteristics, similar to those in healthy children. Protective levels of antibodies to diphtheria were shown to retain for a long time with considerable prolongation of intervals between booster injections. The simultaneous course of immunosuppressive maintenance therapy in the average dosage used for the corresponding age group did not inhibit the production of protective antibodies.