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Chronological changes in hematological findings were analyzed in 225 patients with myelodysplastic syndromes (MDS). They were diagnosed between 1990 and 1992. Their hematological findings, i.e. hemoglobin levels, leukocyte and platelet counts, proportions of peripheral blood (PB) blasts and monocytes, and proportion of blasts in bone marrow (BM), were recorded for up to 42 months after diagnosis, when available. BM was examined regularly in only a few patients. Therefore, it was impractical to use the French-American-British Cooperative Group criteria for subtype classification during the disease course. Thus, we used the percentage of PB blasts as the only indicator of stage evolution. We classified the disease into four stages: stage 1, less than 1% PB blasts; stage 2, 1-5% PB blasts; stage 3, 5-30% PB blasts; and stage 4, 30% or more PB blasts. There were 171 patients initially in stage 1, 37 initially in stage 2, and 17 initially in stage 3. Less than half (45%) of the patients initially in stage 1 progressed to stage 2, while 91% of the patients initially in stage 2 and all of the patients initially in stage 3 showed stage evolution. Eight variables, i.e. BM blasts 5% or more, male sex, karyotypic abnormalities, micromegakaryocytes, mononuclear large megakaryocytes, platelet counts 50 x 10(9)/l or higher, abnormal nucleus of granulocytes, and abnormal granules of granulocytes, were found to be significant risk factors for evolution from stage 1 to 2. Evolution from stage 1 to a higher stage within 15 months of diagnosis was associated with impending poor prognosis in most patients. However, of the 67 patients initially in stage 1 who died, 30 did not show stage evolution. Evolution from stage 2 to a higher stage and from stage 3 to stage 4 was also associated with impending poor prognosis. Higher levels of cytopenia were not associated with poorer prognosis in the stage 1 patients. In conclusion, our grading system proved to be useful in evaluating the chronological changes in MDS patients.

The characteristics of Japanese (JPN) patients with myelodysplastic syndromes (MDS) were investigated in 838 retrospectively collected cases. The median age of the JPN patients was 60 years, about 10 years younger than that in most of the reports based on Western patients. Median survivals were 65 months for refractory anemia (RA), 58 months for RA with ring sideroblasts (RARS), 16 months for RA with an excess of blasts (RAEB), 10 months for RAEB in transformation (RAEBT), and 20 months for chronic myelomonocytic leukemia (CMML). Cumulative leukemia-free rates at final observation were 73% for RA, 79% for RARS, 24% for RAEB, 20% for RAEBT, and 53% for CMML. When low-risk (RA and RARS) patients were divided into two groups, those 40 years of age and older, and those under 40, the cumulative leukemia-free rate was 94% for the younger patients (n = 101), compared with 66% for the older patients (n = 318). The prognostic factors for survival were different from those in Western reports, i.e., variables representing quantitative abnormalities (hemoglobin levels, granulocyte, and platelet counts) were not major prognostic factors, while variables representing qualitative abnormalities (morphological abnormalities in granulocytic and megakaryocytic series cells) were highly significant. Two scoring systems for overall survival and for leukemic transformation were developed, based on multivariate prognostic factor analysis. Neither system included variables representing the degree of cytopenia. Whatever the reason for the different prognostic factors in JPN and Western MDS patients, the use of a scoring system based on Western patients for clinical decision-making in a JPN patient could be misleading, and vice versa.

The plasma soluble interleukin 2 receptor (sIL-2R) level and its relationships with haematologic and immunologic data were examined in 40 patients with myelodysplastic syndromes (MDS). The plasma sIL-2R level was significantly higher in the high-risk MDS group (refractory anaemia with excess blasts (RAEB), RAEB in transformation and chronic myelomonocytic leukaemia) than in the low-risk MDS group (refractory anaemia (RA) and RA with ringed sideroblasts) or in normal subjects, although there was considerable variation in the plasma sIL-2R level within each MDS group. The plasma sIL-2R level correlated positively with the bone marrow cellularity and bone marrow blast mass, but not with the absolute number of CD25+ lymphocytes. This may support the idea that plasma sIL-2R is derived from malignant MDS cells in the bone marrow. The plasma sIL-2R level correlated negatively with the absolute numbers of the CD8+, CD3-CD16+, and CD3-CD56+ cell populations in freshly isolated lymphocytes, the percentage of CD3-CD56+ cells in lymphokine (interleukin 2)-activated killer (LAK) cells, and the cytotoxicity of LAK cells. We conclude that MDS patients having a high plasma sIL-2R level often have a defect in natural killer and CD8+ T-cells.

We studied the type of bcr-abl mRNA for 34 patients with chronic myelogenous leukemia and analyzed for correlations among the mRNA type, the clinical outcome and the transforming activity using the tumorigenicity assay. There was no difference in the distribution of the mRNAtypes (62-a2 and b3-a2) between clinical phases. We found no correlation between the two types of bcr-abl mRNA and the chronic phase duration or survival. The DNA from 12 of 20 chronic phase patients and all five blastic phase patients showed transforming activity. Although there was no difference in the positive rate of transforming activity among the two mRNA-type groups, the blastic phase patients showed a tendency to have higher transforming activity.

To determine the risk factors of the myelodysplastic syndromes (MDS) we conducted a case-control study in Japan. One hundred and sixteen MDS patients were diagnosed from 1 September to 31 October 1992 and from 1 August to 31 October 1993 in the 32 hospitals enrolled in the idiopathic Disorders of Hematopoietic Organs Research Committee. Age, sex, and hospital-matched controls were selected for each case. Information on cigarette smoking and drinking habits, hair dye use, history of keeping pet animals, and occupational exposures to organic solvents, lead and radiation was obtained from self-administered questionnaires. Conditional logistic regression was applied to this individually matched case-control study and odds ratios (ORs) were computed to estimate association between each exposure variable and risk of MDS. Alcohol drinking was associated with increased risk of MDS (OR = 2.15; 95% confidence interval = 1.12-4.16) and there was a significant trend in risk with increasing amounts of ethanol consumed per week (P0.05). We also found elevated ORs for cigarette smokers (OR = 1.80), users of hair dye products (OR = 1.77), and workers exposed to organic solvents (OR = 1.50), although these ratios were not statistically significant. Exposure to pet animals was not associated with risk of MDS. The association observed between alcohol drinking and MDS was still eminent even after adjusted with other variables of cigarette smoking, hair dye use and occupational exposure to organic solvents, and the dose-response relationship was also confirmed.

Little is known about the transforming gene identified in the genomic DNA of chronic myelogenous leukemia (CML) by the tumorigenicity assay. To detect a new transforming gene of CML, we re-investigated the transforming activity of interferon regulatory factor (IRF)-1 and -2 genes in the tumorigenicity assay of NIH3T3 cells transfected with genomic DNA of leukemic cells from 15 patients with CML (12 patients in the chronic phase, one in the blastic phase and two in both phases). We detected the functionally active IRF-2 gene only in the tumor DNA from two CML patients in the blastic phase. We did not detect integration of the IRF-1 gene in the DNA of any tumors derived from the CML patient samples, and also we detected no expression of human IRF-1 mRNA. Thus, NIH3T3 cells may have been transformed due to integration of the functionally active IRF-2 gene from CML patients in the blastic phase. We surmise that there is a possibility that the IRF-2 gene may be involved in the evolution of the blastic phase of CML.

A new B-cell line (VL51) with cytoplasmic villi was established from a female patient with splenic lymphoma with circulating villous lymphocytes (SLVL). The patient exhibited a clinical picture characteristic of SLVL, including massive enlargement of the spleen. Tartrate-resistant acid phosphatase (TRAP)-negative villous lymphocytes were seen in the peripheral blood, bone marrow (BM) and both red and white pulps of the spleen. Monoclonality of the VL51 cell line was confirmed by clonal genotype abnormalities in the immunoglobulin heavy chain (IgH) gene and the T-cell receptor beta (TCR beta) gene. Evidence for commitment of phenotype of the VL51 cell line to the B lineage was also shown by the immunophenotype, including expression of CD10, CD19, CD20 and surface immunoglobins. The VL51 cells were positive for Epstein-Barr virus nuclear antigen (EBNA). The VL51 cell line is the first SLVL cell line to be established, and it is expected to be useful in clarifying the leukemogenesis of SLVL.

To evaluate the molecular events in the genome that are associated with myelodysplastic syndromes (MDS) and the development of leukemia, we investigated the expression of the deleted in colorectal carcinoma (DCC) gene by the reverse transcriptase-polymerase chain reaction (RT-PCR) method in 24 MDS cases and in 7 overt leukemia cases that progressed from MDS. Expression of the DCC gene was absent or extremely reduced in 2 of the 24 MDS cases, and those 2 cases developed overt leukemia within 6 months. Moreover, in 5 of the 7 cases of overt leukemia that developed from MDS, expression of the DCC gene was absent or extremely reduced. These findings suggest that inactivation of the DCC gene may be the late event that triggers the progression of MDS to leukemia.

Eighty-one of 473 patients with refractory myelodysplastic anemias registered by the Japanese Cooperative Study Group prior to January 1987 survived more than 5 years. At presentation, most patients had mild cytopenia, a less aggressive form of the disease (48 with refractory anemia, 23 with refractory anemia with ring sideroblasts, 10 with refractory anemia with excess of blasts), less blast cells in the marrow and blood, and onset at younger ages. Their clinical profiles 5 years after presentation showed no significant improvement. The results suggest that the long-term survivors were found in a subpopulation of patients with a favorable prognosis and that it is a part of the natural course rather than the results of treatment.

We determined the position of the breakpoint within the bcr gene in 22 patients with Philadelphia-positive chronic myelocytic leukemia using conventional Southern-blots and analyzed its relationship to thrombopoiesis. After DNA digestion with restriction endonucleases (Hind III, Bam HI and Bgl II), we localized the breakpoint in bcr using two genomic probes. The location of the breakpoint within the bcr was assigned to one of five zones. Breakpoints in zones 1 and 2 were grouped as "5"', and those in zones 3, 4 and 5 as "3'". Thus we subdivided patients with bcr rearrangements into those with genomic breaks at either 5' or 3' of the Bam HI site, just upstream of exon 3. Nine patients had 5' breakpoints and 13 patients had 3' breakpoints. The platelet counts of 3' patients were significantly higher than those of 5' patients (1395 vs 274 x 10(9)/l; p less than 0.03). The megakaryocyte counts from bone marrow histological sections in 3' patients (n = 12) and 5' patients (n = 7) were 63.4/mm2 and 19.5/mm2, respectively, with a significant difference of p less than 0.006. The mean number of megakaryocyte progenitor cells assayed by in vitro cloning was 128.3/2 x 10(5) bone marrow cells for 3' patients (n = 7) compared with 46.3 for 5' patients (n = 4). These results suggest that Philadelphia-positive CML patients with 3' breakpoints have higher thrombopoietic activity than patients with 5' breakpoints.