The Relationship of Kynurenine and Neopterin Levels and Their Association with a Selection of Other Immune Markers in Chronic Myeloid Leukaemia Patients.

Among malignant diseases, chronic myeloid leukaemia (CML) is one of the best suited candidates for immunotherapy. For this purpose it is necessary to broaden the present knowledge on the immunology of this disease. As a part of such a project, the levels of kynurenine (KYN) and neopterin (NPT) were studied in 28 CML patients and in the same number of healthy subjects. At diagnosis, both KYN and NPT levels were found to be elevated in a significant portion of the patients and dependent on their leukocyte count. As in the case of KYN, increased NPT levels dropped after achieving remission. When correlating KYN and NPT levels with a selection of other markers tested, significant association was revealed only in the case of CRP and IL-6. However, there were several patients with increased KYN levels in whom NPT was not detected, and vice versa. The relapse of the disease observed in two patients was accompanied by an increased level of NPT in both cases, but by an increased level of KYN in only one of them. No significant correlation was found between KYN and NPT levels in sera taken at diagnosis. However, when the whole set of sera was taken into consideration, the association became statistically significant. Although the data obtained revealed a number of similarities between KYN and NPT production in CML patients, it also suggested a difference in the kinetics of these two biomarkers' production.

Isolation and properties of gene-modified mouse bcr-abl-transformed cells expressing various immunostimulatory factors.

UNLABELLED: B210 cells are murine (BALB/c) cells transformed by bcr-abl fusion gene. After intravenous administration they are capable of inducing leukaemia-like disease in syngeneic mice. From these cells a thymidine-kinase less subline was derived. It was significantly less pathogenic than the parental cells. However, a highly pathogenic clone denoted B210cTK-/cl-2 was isolated from its population. As determined by Western blotting, these cells produced more p210 protein than the parental B210 cells. To successfully transfect these cells a modified electroporation method was introduced. Bicistronic plasmids carrying gene for herpes simplex thymidine kinase (HSV TK) and the gene for either granulocyte-monocyte colony stimulation factor (GM-CSF), interleukin-2 (IL-2) or interleukin 12 (IL-12) were used for the transfection experiments. Gradually, cell lines producing these cytokines were isolated in media supplemented with hypoxantin, aminopterin and thymidine (HAT). All of them were highly sensitive to ganciclovir in vitro confirming that the cells produced HSV TK. The genetic modification of B210cTK-/cl-2 was associated neither with the alteration of p210 bcr-abl production nor with any changes in expression of MHC class I molecules. From populations of each of the three lines several cell clones were isolated and tested for the production of the respective cytokines. The original uncloned population and several clones differing in the cytokine production were administered intravenously into mice. All animals survived without symptoms of the disease suggesting that the gene-modification was associated with the loss of pathogenicity. KEYWORDS: CML, Bcr-Abl, HSV TK, cytokines, gene-modified tumour cells, pathogenicity.

Characteristic of two mouse bcr-abl-transformed cell lines: I. General properties of the cells.

In an effort to develop an experimental system suitable for immunological studies in which Bcr-Abl-positive cells are to be used as antigens, we examined the properties of two mouse (Balb/c) established cell lines that express the Bcr-Abl protein and are oncogenic for syngeneic animals. Under standard conditions the two cell lines, viz. Ba-p210 (B210) and 12B1, expressed comparable amounts of the Bcr-Abl protein. However, they differed in a number of characteristics. From the morphological point of view, B210 cells were the more homogeneous, being mainly represented by leukaemic blastic cells with a large number of AgNORs as markers indicating a high proliferative activity. 12B1 cells were more polymorphic and giant cells were detected within their populations. Many 12B1 cells exhibited nuclear segmentation and "band-like" structures. Markers of proliferation were less frequent in 12B1 and the tendency for aging was more pronounced in these cells. The 12B1 cells were slightly more sensitive to imatinib mesylate than B210 cells. In B210 cells, the expression of MHC class I was downregulated, which was not the case with 12B1 cells. Both cell lines induced leukaemia-like disease in mice after intravenous application but, as compared with B210, 12B1 cells were about 100 times more oncogenic and the disease they induced was more aggressive. Moreover, 12B1, but not B210, induced tumours after subcutaneous or intraperitoneal inoculation.