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Summary. We examined deletion and methylation of the p15INK4B(p15) and p16INK4A(p16) genes, using Southern blotting and methylation-specific polymerase chain reaction (PCR), in 70 untreated adult patients with precursor B-cell acute lymphoblastic leukaemia (PBC-ALL) and analysed the relationship between their genetic changes and clinical outcome. Methylation and homozygous deletion of the p15 gene were detected in 30 (43%) and 18 (26%) patients, while those of the p16 gene were found in 16 (23%) and 11 (16%) patients respectively. Thirteen out of 17 patients with wild-type p15 gene showed expression of p15 mRNA, whereas 31 out of 39 patients with alteration (deletion and methylation) of the p15 gene showed no p15 mRNA expression by reverse transcription–PCR, suggesting that alterations of the p15 gene are highly associated with loss of␣p15 mRNA expression. Disease-free survival (DFS) at 4 years in patients with wild-type p15 gene is 33%, compared with 4% of those with p15 gene alterations (P = 0·049). Multivariate analysis showed that the absence of p15 gene alterations was an independent significant favourable prognostic factor for longer DFS (P = 0·0001). These results suggest that alterations in the p15 but not p16␣gene can be used as a genetic prognostic indicator in PBC-ALL.

The AML1 gene encoding the DNA-binding -subunit in the Runt domain family of heterodimeric transcription factors has been noted for its frequent involvement in chromosomal translocations associated with leukemia. Using reverse transcriptase-polymerase chain reaction (RT-PCR) combined with nonisotopic RNase cleavage assay (NIRCA), we found point mutations of the AML1 gene in 8 of 160 leukemia patients: silent mutations, heterozygous missense mutations, and biallelic nonsense or frameshift mutations in 2, 4, and 2 cases, respectively. The mutations were all clustered within the Runt domain. Missense mutations identified in 3 patients showed neither DNA binding nor transactivation, although being active in heterodimerization. These defective missense mutants may be relevant to the predisposition or progression of leukemia. On the other hand, the biallelic nonsense mutants encoding truncated AML1 proteins lost almost all functions examined and may play a role in leukemogenesis leading to acute myeloblastic leukemia.

We present a patient with refractory multiple myeloma who showed a good response to a combination therapy with oral melphalan, dexamethasone, and thalidomide (MDT). A 48-year-old woman with myeloma refractory to thalidomide, dexamethasone, and clarithromycin received 6 mg melphalan for 4 days every 6 weeks in combination with thalidomide (100 mg daily) and dexamethasone (5 mg daily for 2 days every week). Four months after the initiation of MDT therapy, a 78% reduction of monoclonal protein was achieved. Although the efficacy of oral MDT combination therapy in elderly patients with newly diagnosed myeloma has been reported, the present data demonstrate the effectiveness of MDT therapy for refractory myeloma and warrant further exploration with this MDT regimen to treat myeloma.

As recurrent chromosome abnormalities in leukemia are highly associated with particular subtypes, the genetic events of specific chromosome alteration must be associated with leukemogenesis and characteristics of the disease. The chromosomal breakpoints involved in inv(16) and t(16;16) have been shown to generate the fusion gene PEBP2beta(CBFbeta)/MYH11. The PEBP2beta/MYH11 fusion transcripts in all 8 patients with M4Eo, 2 of 18 with M4, and one CML in the blastic phase were detected by using RT-PCR and Southern blotting. We demonstrated the marked expression of CD34 and c-KIT (CD117) antigens in myelomonoblastic leukemia cells from all patients carrying this fusion gene, which was in contrast to the patients with M4 but without the fusion gene. These results indicate that immunophenotypic analysis is useful for detection of leukemia with the fusion gene, and that the PEBP2beta/MYH11 fusion gene is involved in immature cells expressing CD34 and c-KIT antigens.

A 53-year-old female infected with human T lymphotropic virus type-I (HTLV-I) suffered from chronic renal failure, aplastic anemia, myelopathy, uveitis, Sjögren's syndrome and Weber-Christian disease. Although HTLV-I antibody was negative in cerebrospinal fluid, she was diagnosed as HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP) based on clinical and histological findings. Though to date there is no direct evidence, other complications have also been reported to be HTLV-I related diseases. This case provided the unique opportunity to observe various HTLV-I related diseases.

We present four patients with adult T-cell leukemia (ATL) derived from a novel T-cell subset (CD4-, CD8- [double-negative, DN], T-cell receptor [TCR] alpha beta+). In the ATL cells of these patients, neither gene nor surface expression of CD4 and CD8 antigens was detected. Clinical and laboratory data showed no difference between DN- ATL and CD4+ATL patients. In contrast to typical CD4+ATL cells, DN-ATL cells were shown to express the protein and messenger RNA (mRNA) for S100 beta in immunocytochemical assay and the reverse-transcription polymerase chain reaction assay. The mean fluorescence intensity of the TCR/CD3 complex was extremely low in all four DN-ATL patients as well as in typical CD4+ ATL. All four patients had TCR beta and gamma chain gene rearrangements, with deletion of TCR delta chain gene and mRNA expression for TCR alpha, beta, and CD3 delta but not for TCR gamma and delta chain genes. Thus, CD4- CD8- TCR alpha beta T cells are also a target for human T-cell lymphotrophic virus type I-induced leukemogenesis. In addition, expression of the TCR alpha beta/CD3 complex on the DN-ATL cells was further diminished by the addition of anti-CD3 or anti-TCR alpha beta monoclonal antibody. These results suggest that the decreased expression of the TCR alpha beta/CD3 complex by ATL cells plays a key role in the development of ATL, irrespective of CD4 expression.

To clarify the aspects affected by the PEBP2beta/MYH11 fusion gene involved in the inv(16), we analysed immunophenotypes in myelomonoblastic leukaemias. We found high expressions of CD34 and c-KIT antigens in myelomonoblastic cells from all patients carrying this fusion gene, including two with M4 and one CML blastic phase, in contrast to those with M4 without the fusion gene. These findings indicate that immunophenotyping is useful for detecting a leukaemia with the fusion gene in myelomonoblastic leukaemias and that the PEBP2beta/MYH11 gene is involved in immature cells expressing CD34 and c-KIT antigens.

We examined deletion and methylation of the p15INK4B (p15) and p16INK4A (p16) genes, using Southern blotting and methylation-specific polymerase chain reaction (PCR), in 70 untreated adult patients with precursor B-cell acute lymphoblastic leukaemia (PBC-ALL) and analysed the relationship between their genetic changes and clinical outcome. Methylation and homozygous deletion of the p15 gene were detected in 30 (43%) and 18 (26%) patients, while those of the p16 gene were found in 16 (23%) and 11 (16%) patients respectively. Thirteen out of 17 patients with wild-type p15 gene showed expression of p15 mRNA, whereas 31 out of 39 patients with alteration (deletion and methylation) of the p15 gene showed no p15 mRNA expression by reverse transcription-PCR, suggesting that alterations of the p15 gene are highly associated with loss of p15 mRNA expression. Disease-free survival (DFS) at 4 years in patients with wild-type p15 gene is 33%, compared with 4% of those with p15 gene alterations (P = 0.049). Multivariate analysis showed that the absence of p15 gene alterations was an independent significant favourable prognostic factor for longer DFS (P = 0.0001). These results suggest that alterations in the p15 but not p16 gene can be used as a genetic prognostic indicator in PBC-ALL.

We present an elderly patient with mantle cell leukemia who was successfully treated with low-dose cyclophosphamide (CY). A 76-year-old female was diagnosed as mantle cell leukemia based on abnormal lymphocytosis and splenomegaly without lymphadenopathy. She was orally treated with 50 mg of CY daily and had continuous remission over 4 years. Rearrangements of BCL1 and immunoglobulin heavy chain genes in the peripheral blood lymphocytes were detected at diagnosis, but not 1 or 4 years later. Further studies are required to confirm the role of low-dose CY therapy for patients with mantle cell leukemia and lymphoma.

TEL is a new member of the ETS-like family on chromosome 12 and forms fusion genes with several partners in leukemia. Among these fusion genes, the TEL/AML1 translocation resulting from t(12;21) is found in approximately one quarter of the childhood B-cell lineage acute lymphoblastic leukemia (ALL) cases and its prognosis is excellent. We examined 42 adult patients with B-cell lineage ALL and 13 adult patients with lymphoblastic transformation of chronic myeloid leukemia (CML) to detect TEL/AML1 fusion genes using the reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blotting, but no translocation was detected. These findings indicate that absence of the TEL/AML1 fusion transcript partly correlates with the poorer outcome of adult B-cell lineage ALL as compared with childhood ALL and the TEL/AML1 fusion transcript is specific for pediatric B-cell lineage ALL.