G. Holm

Lymphocyte abnormalities in untreated patients with Hodgkin's disease.

The study comprises 38 unselected and untreated patients with Hodgkins disease (HD) and 23 healthy persons. Highly purified blood lymphocytes were analyzed for cells forming rosettes with sheep red blood cells (T lymphocytes), and lymphocytes bearing surface immunoglobulin (B lymphocytes) and/or carrying receptors for complement. Their DNA synthesis, spontaneously, or after activation with mitogens (phytohemagglutinin, concanavallin A, poke weed mitogen) and purified protein derivative (PPD) was measured. Delayed skin hypersensitivity to PPD and mumps antigen was studied. Most HD patients had low numbers of T lymphocytes (50% of the cases below normal range) while the mean B‐lymphocyte level was normal but with a greater variation than in the control group. Lymphocytes from most patients were poorly stimulated by T‐cell mitogens. Two‐thirds of the patients and one healthy control had negative skin reaction to 2 TU PPD and the DNA synthesis of their lymphocytes after activation with PPD was low. Large lymphoid cells (> 9‐mm diameter) were commonly present in HD blood and the spontaneous DNA synthesis was high, particularly in lymphocytes from stage B patients. The percentage of T lymphocytes and the stimulation of lymphocytes by T‐cell mitogens or by PPD, a T‐lymphocyte function, did not correlate and each test only detected defects in about half the cases. Simultaneous application of all tests revealed abnormalities of blood T lymphocytes in 33 out of 38 patients. Although the defects were usually more pronounced in patients with advanced disease, the impairment of T lymphocytes and their functions is present in all stages of Hodgkins disease.

Modulation of anti-idiotypic immune response by immunization with the autologous M-component protein in multiple myeloma patients.

Multiple myeloma is characterized by a proliferation of clonal B lymphocytes and plasma cells. The idiotypic structure of clonal immunoglobulin (Ig) expressed on the tumour B‐cell surface can be regarded as a tumour‐specific antigen and, as such, a potential target for anti‐idiotypic T and B cells in an immune regulation of the tumour‐cell clone. Active immunization using the autologous monoclonal Ig as a ‘vaccine’ was shown to induce tumour‐specific immunity in murine B‐cell tumours and in human B‐cell lymphoma. With the aim to induce or amplify an anti‐idiotypic response in multiple myeloma, five stage I–III patients were repeatedly immunized with the autologous monoclonal IgG. Induction of idiotype‐specific cellular immunity was analysed in vitro by an enzyme‐linked immunospot assay (interferon‐γ and interleukin‐4 secreting cells). B cells secreting anti‐idiotypic IgM antibodies were also analysed. An anti‐idiotypic T‐cell response was amplified 1.9–5‐fold in three of the five patients during immunization. The number of B cells secreting anti‐idiotypic antibodies also increased in these three patients. In two of the patients induction of idiotype‐specific immunity was associated with a gradual decrease of blood CD19+ B cells. The induced T‐cell response was eliminated during repeated immunization. Further studies are warranted to optimize the immunization schedule in order to achieve a long‐lasting T‐cell immunity against idiotypic determinants on the tumour clone. A role for immunity in controlling the tumour clone remains to be established.

Prognostic factors in hodgkin's disease with special reference to age

A series of 182 patients with Hodgkins disease, diagnosed between January 1973 and December 1978 was used to identify prognostic factors with special reference to age. There were 118 men and 64 women (mean age, 47 years; r = 15–92). During the same period 57 elderly patients who were never referred, were reported to the Local Cancer Registry. The diagnosis had been established shortly before death or at autopsy. The 182 patients under study were evenly distributed in Stages I‐IV. Nodular sclerosis (38%) and mixed cellularity (38%) were the most common histologic subtypes. The 5‐year survival probability estimate was 28% in patients above 50 years as compared to 74% in the remainder. Survival was significantly better in patients with Stage I‐II disease and lymphocyte predominance/nodular sclerosis histopathology. Age was the main prognostic factor in the whole series as well as in patients older than 50 years. However, in young patients advanced clinical stage and B‐symptoms were related to a poor prognosis. Biologic indicators such as ESR, hemoglobin and albumin were intimately linked to the extent of disease and did not add prognostic information besides that given by the clinical stage. It is concluded that the prognosis in elderly remains poor and appears to be partly unrelated to those factors which determine the prognosis in the young, assumingly reflecting a depressed host‐response and/or a decreased tolerance to intensive treatment. Cancer 53:1202‐1208, 1984.

Idiotype‐specific T cells in multiple myeloma stage I: an evaluation by four different functional tests

Idiotype-specific T cells were characterized in patients with multiple myeloma stage I by analysing idiotype-induced DNA synthesis (3H-thymidine incorporation), IL-2 and IFN-gamma production at the single cell level (ELISPOT) (in vitro tests) and delayed type hypersensitivity (DTH) skin reaction (in vivo test). In seven out of eight patients at least one of the four tests was positive. In five patients three or more tests were positive. One patient was negative in all four tests. Six patients had both IL-2 and IFN-gamma-secreting cells and three of them also a DTH response. Furthermore, those three patients with a proliferative response also had IL-2 and IFN-gamma-secreting cells induced by the idiotype. The data indicate that part of the idiotype-specific T cell fraction belongs to the CD4 Th1 cell population. Whether CD8-specific T cells also were present could not be ruled out. The present study provides further support for the existence of idiotype-specific T cells in multiple myeloma. Such cells might be an important target for an immune-mediated therapeutic approach.

Interleukin 6, tumour necrosis factor α, interleukin 1β and interleukin 1 receptor antagonist promoter or coding gene polymorphisms in multiple myeloma

Proinflammatory cytokines such as interleukin 6 (IL‐6), tumour necrosis factor α (TNF‐α) and IL‐1β are considered to be involved in the pathogenesis of multiple myeloma (MM). In the present study, we examined a G/C polymorphism at position −174 in the promoter region of IL‐6, a biallelic polymorphism at position −308 in the promoter region of TNF‐α, the TaqI restriction fragment length polymorphism in exon 5 of IL‐1β and a variable number of identical tandem repeat polymorphisms in intron 2 of IL‐1 receptor antagonist (IL‐1Ra) genes. The alleles of these loci are known to influence the level of production of the cytokines and the IL‐1Ra. Seventy‐three patients with MM, 27 with monoclonal gammopathy of undetermined significance (MGUS) and 129 healthy individuals were included. No difference was found between patients and healthy controls or between MM and MGUS patients in the distributions of genotypes and frequencies of alleles of the IL‐6 (−174), TNF‐α (−308), IL‐1βTaqI and IL‐1Ra gene polymorphisms. No associations between the polymorphisms at the loci under study and clinical factors such as age, sex, clinical stage at onset and M‐protein type were observed. Our results indicate that the cytokine (IL‐6, TNF‐α and IL‐1β) and IL‐Ra gene polymorphisms do not confer susceptibility to the development of MM.

Idiotypic Immunoglobulin Structures on Blood Lymphocytes in Human Plasma Cell Myeloma

E*, cells forming rosettes with sheep red blood cells (SRBC) (T-lymphocytes); EA*, cells rosetting with SRBC coated with rabbit IgG antibody; EAC*, cells rosetting with SRBC coated with rabbit antibody and human complement; SIg*, cells with surface bound immunoglobulin; CIg*, cells with cytoplasmic immunoglobulin; pv, polyvalent (SIg-pv* = surface staining by polyvalent antiserum); id, idiotypic (SIg-id* = surface staining by anti-idiotypic antiserum); HP*, cells with receptors for Helix pomatia A hemagglutinin; IFL, immunofiuorescence; CLL, chronic lymphocytic leukemia.

Natural killer cell activity in monoclonal gammopathies: Relation to disease activity

Natural killer (NK) activity and NK‐related cell surface markers (CD16, CD56, CD57) of peripheral blood lymphocytes were studied in patients with multiple myeloma and MGUS (monoclonal gammopathy of undetermined significance). A strong correlation (p < 0.0001) was found between the numbers of cells positive for the different NK cell surface markers. The proportion of CD16+cells correlated highly to the lytic capability (lytic units/106cells) of K562 cells (p < 0.0001). High NK activity and high numbers of cells with NK‐related cell surface markers were found in patients with a low tumor burden compared to controls, whereas low values were seen in patients with an advanced disease. The results indicate that NK cells might be involved in the disease process in monoclonal gammopathies, perhaps by exerting a regulatory function on the proliferating B‐cell clone.

Idiotype‐specific T lymphocytes in monoclonal gammopathies: evidence for the presence of CD4+ and CD8+ subsets

Tumour‐specific CD4+ T helper (Th) and CD8+ T cytotoxic (Tc) cells may participate in the control and eradication of tumour cells. In the present study, idiotype‐specific stimulation of CD4+ and CD8+ blood T cells from patients with monoclonal gammopathy of undetermined significance and patients with untreated multiple myeloma stage I was examined. Activation was measured in the CD4+ and CD8+ subsets enriched by magnetic microbeads as the incorporation of 3H‐thymidine and the secretion of interferon (IFN)‐γ, interleukin (IL)‐2 and IL‐4 by single cells using the enzyme‐linked immunospot assay. Idiotype‐specific T cells were found in four of seven patients. Stimulation was mainly confined to the CD4+ subset in three of the four responding patients. This type of response was major histocompatibility complex (MHC) class II restricted as it could be inhibited by monoclonal antibodies against MHC class II (HLA‐DR), but not against class I (HLA‐ABC) molecules. Idiotype‐specific CD8+ T cells were also demonstrated in these patients although at a lower frequency. One patient showed a strong and dominating activation of CD8+ T cells which could be blocked by antibodies against HLA‐ABC but not against HLA‐DR. Idiotype‐specific CD4+ or CD8+ T cells were mainly of the type‐1 subsets as judged by their secretion of IFN‐γ and IL‐2. Thus, this study provides evidence for the presence of idiotype‐specific and MHC‐restricted CD4+ and CD8+ T cells of the type‐1 subsets in patients with monoclonal gammopathies. Such T cells with the potential to control the growth of tumour B cells may be a suitable target for immunotherapeutic interventions in patients.

Telomerase activity in plasma cell dyscrasias

Activation of telomerase is essential for in vitro cellular immortalization and tumorigenesis. In the present study, we investigated telomerase activation and its implications in plasma cell dyscrasias including monoclonal gammopathy of undetermined significance (MGUS), multiple myeloma (MM) and plasma cell leukaemia (PCL). All 5 patients with MGUS exhibited normal levels of telomerase activity in their plasma cells. Elevated telomerase activity was found in the samples from 21/27 patients with MM and 4/4 with PCL. In addition, 4 myeloma cell lines all expressed high levels of telomerase activity. The expression of telomerase reverse transcriptase (hTERT) and telomerase RNA template (hTER) was positively associated with the levels of telomerase activity in MM/PCL. Tankyrase expression was upregulated, concomitant with the induction of hTERT and activation of telomerase in MM/PCL. The present findings indicate that MGUS cells may not be immortalized and that activation of telomerase plays a role in the malignant transformation from MGUS to MM.

Generation of dendritic cells from peripheral blood adherent cells in medium with human serum

Dendritic cells (DC) provide an effective pathway for presenting antigens to T cells, both self‐antigens during T‐cell development and foreign antigens during immunity. As such, these cells may be promising adjuvants for immunotherapy. Thus, it is important to establish simple and fast method(s) to generate sufficient numbers of human DC in medium free of calf serum so that the cells can be used for both experimental and clinical purposes. In this report, we used peripheral blood adherent cells, without laborious cell purification or depletion, as the starting population and cultured them in medium supplemented with granulocyte/macrophage colony‐stimulating factor and interleukin‐4. Substantial numbers of cells with the phenotypical and functional characteristics of immature DC were obtained in a 7‐day culture. We then compared DC cultured in medium supplemented with either fetal calf serum or pooled human ABRh+ serum and found no difference in cell yields and in their ability to stimulate alloreactive T cells or to present soluble antigens to T cells. Irradiated cells were less efficient than non‐irradiated cells in antigen presentation and stimulation of T cells. Finally, we have examined DC with or without additional tumour necrosis factor‐α treatment and found that antigen‐pulsed mature cells could as efficiently present antigen to T cells as did immature cells. This method is suitable for the generation of DC in studies of large clinical materials.