Cell proliferation detected by DNA polymerase alpha in acute leukemias.

Fifty-nine patients with acute leukemia were studied by flow cytometry using monoclonal antibody against DNA polymerase alpha. Since fresh and frozen cells showed identical flow cytometric histograms, 86 cryopreserved samples (39 from peripheral blood and 47 from bone marrow) were used in this study. The DNA polymerase alpha-positive population ranged from 16.3 to 84.7% in peripheral blood, and from 6.5 to 92.1% in bone marrow. A positive correlation (r = 0.80; p < 0.01) was found between DNA polymerase alpha-positive populations in peripheral blood and bone marrow from the same patient. This suggests that the DNA polymerase alpha-positive population in the bone marrow can be estimated from that in peripheral blood. A negative correlation was observed between the positive population in bone marrow samples and the time to reach a nadir (r = -0.58: p < 0.01), while a positive correlation was found between the tumor cell count in bone marrow and the DNA polymerase alpha-positive population (r = 0.64; p < 0.01) in patients who responded to chemotherapy.

[Myelodysplastic syndrome developed in a mother and her son whose bone marrow karyotype showed monosomy 7].

Two familial cases of myelodysplastic syndrome (MDS) are reported, one of whom had an abnormal karyotype of 45, XY, -7 (monosomy 7). Case 1 was a 60-year-old woman developed dizziness and nasal bleeding. She was treated with blood transfusion alone. About 11 months after diagnosis, she died of pneumonia. Case 2 was a 22 year-old man, who was the son of case 1, developed febrile disease because of recurrent skin and oral mucosa infections. He had a partial response to low-dose of cytarabine. Thirteen months after diagnosis, he died of severe pneumonia. Both cases were diagnosed as having refractory anemia with excess of blasts due to peripheral blood and bone marrow findings. Both patients had pancytopenia, erythroid hyperplasia in bone marrow, marked dyserythropoiesis, recurrent infectious diseases and severe pneumonia that resulted in death. These symptoms resembled to those reported for monosomy 7 syndrome. Familial MDS with monosomy 7 is rarely reported. These cases are of interest to investigate hereditary factors of MDS.

[Anemia and neutropenia in elderly patients caused by copper deficiency for long-term enteral nutrition].

Anemia and neutropenia caused by copper deficiency is a well-known consequence of long term total parenteral nutrition in the literature. We present 6 bed-ridden elderly patients who developed anemia and neutropenia after receiving enteral nutrition for a long time (mean: 3.3 years) In all 6 patients, serum copper and ceruloplasmin level were very low, and the mean of their hematological data were as follows: WBC 2,200/microliters, neutrophil 554/microliters, hemoglobin 8.1 g/dl, platelet 260 x 10(3)/microliters, respectively. The bone marrow examination showed cytoplasmic vacuolization of both myeloid and erythroid precursors, and maturation arrest of granulopoiesis. Then, copper sulfate was administrated by enteral tube to 6 patients, and the improvement of anemia and neutropenia was observed within a month. A 82-year-old woman who received enteral nutrition for 3.5 years with sever anemia (Hb 3.7 g/dl) and neutropenia (neutrophil 350/microliters), showed a marked improvement in hematological data (Hb 8.0 g/dl, neutrophil 4, 092/microliters, respectively) after two months by administering the copper supplementation. The exact cause of the anemia and neutropenia in copper deficiency is unclear, but it is suggested that the decreased activity of enzyme containing copper may be related. Hematological abnormalities due to copper deficiency should be cared during long term enteral nutrition with long termed bed-ridden elderly patients.

[Disseminated intravascular coagulation (DIC) and pre-DIC due to severe infection in the elderly].

Because of pre-thrombotic state frequently present in the elderly, sepsis easily progresses to pre-DIC and DIC, sometimes with a fatal outcome. We assessed 31 elderly patients who developed pre-DIC and DIC due to severe infection. They were divided into two groups, early death group: 14 elderly patients with poor prognosis died within 14 days, and long survival group: 17 patients with good prognosis lived 15 days or more. Controls consisted of 31 elderly thrombotic disease cases and 25 healthy elderly cases. The DIC score was significantly higher in the early death group than in the long survival group, and there was a correlation between DIC score and survival. Moreover, many of the early death group were long-termed bed-ridden patients, serum BUN and Cre levels were significantly increased in the early death group compared to the long survival group. While plasma TAT, PIC and D-dimer levels were increased in thrombotic disease group compared to the healthy control group, TAT and D-dimer were also increased in the pre-DIC and DIC state than in the thrombotic state. In the early death group, D-dimer was higher than in the long survival group. We suggest that early diagnosis by molecular marker is important in the DIC stage, and a high D-dimer level may be a poor prognostic factor.

Flow cytometric detection of proliferative cells in leukemias.

We studied the proliferative activity of leukemic cells obtained from the peripheral blood and bone marrow of 34 patients; 30 with acute leukemia and 4 with chronic myelogenous leukemia in blastic crisis. Flow cytometry was performed using monoclonal antibody against DNA polymerase alpha. Since fresh and frozen cells showed virtually identical DNA polymerase alpha-positive populations and flow cytometric histograms, 52 cryopreserved samples (25 from peripheral blood and 27 from bone marrow) were used in this study. The DNA polymerase alpha-positive population ranged from 20.4% to 84.7% in peripheral blood, and from 6.5% to 92.1% in bone marrow. A positive correlation (r = 0.76, P < 0.01) was found between DNA polymerase alpha-positive populations in peripheral blood and bone marrow from the same patient. This suggests that the DNA polymerase alpha-positive population in the bone marrow can be estimated from that in peripheral blood. No relationship was observed between the positive population and the response to chemotherapy. Statistical analyses for all cases showed no relationship between the DNA polymerase alpha-positive population and either the tumor cell count or time to reach a nadir. However, a negative correlation was observed between the positive population in bone marrow samples and the time to reach a nadir (r = -0.64, P < 0.05) in those patients who achieved a complete response. In addition, in the cases of acute non-lymphocytic leukemia who did not respond to chemotherapy, a positive correlation was observed between the tumor cell count in bone marrow and the DNA polymerase alpha-positive population (r = 0.93, P < 0.01). Thus, the method described here provides a simple and time-efficient means of detecting the proliferative activity of leukemic cells, which is a useful parameter in the treatment of leukemia.

Treatment of adult T-cell leukemia/lymphoma with MST-16, a new oral antitumor drug and a derivative of bis(2,6-dioxopiperazine). The MST-16 Study Group.

BACKGROUND: MST-16, a new orally administered bis(2,6-dioxopiperazine) analogue and an inhibitor of topoisomerase II, was given to 24 patients with adult T-cell leukemia-lymphoma (ATLL) in a Phase I-II multi-institutional cooperative study. METHODS: MST-16 was administered orally daily for 7 days, with courses repeated at intervals of 2-3 weeks in 24 patients. RESULTS: Two complete remissions (CR) and eight partial remissions (PR) were obtained in 23 evaluable patients who received 1200-2800 mg/day of MST-16. Among 13 acute-type ATLL, one CR and five PR were obtained. Among eight lymphoma-type ATLL, two PR were detected. Among two chronic-type ATLL, one CR and one PR occurred. Remissions were obtained at 7-232 days (median, 23 days) and lasted 43-374 days (median, 68 days). The major toxic effects were leukopenia (68%), anemia (52%), thrombocytopenia (35%), and gastrointestinal disorders (22%). CONCLUSIONS: MST-16 was shown to be effective in ATLL, which has no standard therapy. This drug deserves further clinical trials because it shows little cross resistance to currently available antitumor drugs.

[Chronic myelogenous leukemia complicated with sarcoidosis].

A 79-year-old man who had been diagnosed as having sarcoidosis when he was 63 year old, was admitted to our hospital because of marked thrombocytosis and leukocytosis in July 1991. The low neutrophil alkaline phosphatase (NAP) score, presence of Philadelphia (Ph1) chromosome in the bone marrow cells, and M-BCR rearrangement by Southern blot hybridization were observed. He was diagnosed as having chronic myelogenous leukemia complicated with sarcoidosis. The coexistence of sarcoidosis and leukemia has rarely been reported. It is difficult to discuss that there is not causal association between of them.

[Non-Hodgkin's lymphoma in a patient with sarcoidosis (the sarcoidosis-lymphoma syndrome)].

A-31-year-old man with right cervical and supraclavicular lymphadenopathy was admitted in March, 1991. He was diagnosed as having muscular sarcoidosis at the age 8 year, and was treated with corticosteroids. Since age 18, his skin was erythematous and ulcerous, and later his skin became gradually atrophic. Lymph node biopsy revealed diffused large cell non-Hodgkin's lymphoma. Lymphoma cells showed TCR-beta gene rearrangement by Southern blot hybridization. His lymphoma was refractory to CHOP and CHOP-Bleo regimens. Complete remission was achieved with cisplatin and etoposide. However, early relapse occurred, and he died of pulmonary hemorrhage 4 months after the diagnosis of non-Hodgkin's T-cell lymphoma. The so called "sarcoidosis-lymphoma syndrome" is uncommon in Japan. In 9 of 10 cases previously reported, malignant lymphoma occurred during the course of sarcoidosis. Most of the sarcoidosis cases were chronic active type, and required systemic administration of corticosteroids. Hodgkin's disease coexistent with sarcoidosis as reported in other countries, was not found in Japan. These findings suggest that the low incidence of sarcoidosis-lymphoma syndrome in our country is due to the relative rareness of Hodgkin's disease. The sarcoidosis-lymphoma syndrome possibly appears as a consequence of immunological abnormalities observed in sarcoidosis.

[Chimerism monitoring by VNTRs polymorphism analysis in a patient who received allogenic bone marrow transplantation].

A 39-year-old female diagnosed as acute myelogenous leukemia received allogenic bone marrow transplantation (BMT) pre-conditioned with busulfan and cyclophosphamide regimen from her HLA identical sibling. To distinguish donor and recipient cells, we analyzed variable numbers of tandem repeats (VNTRs) polymorphisms using a YNH-24 probe by Southern blot hybridization. VNTRs polymorphism analysis documented the engraftment of donor cells, relapse of recipient cells, and mixed hematopoietic chimerism. Assessment of the chimerism state is important for determining the prognosis of patients undergoing BMT, and VNTRs polymorphisms analysis is very useful for identifying the chimerism state.

[Neutrophilic dermatosis in a patient with refractory anemia].

A 46-year-old man diagnosed as refractory anemia was hospitalized because of high fever and extensive erythema with ulceration in the femoral region. His peripheral blood examination showed marked leukocytosis (WBC 31,500/microliter:neutrophilic 90%) and anemia (Hb 8.6 g/dl. In spite of administration of antibiotics, the cutaneous ulcer rapidly extended to the right thigh and became necrotic. The bacterial culture of the cutaneous lesion showed no growth and a skin biopsy showed infiltration of neutrophils in the dermis. He became afebrile and his cutaneous lesion improved after administration of corticosteroid. When the dose of corticosteroid was decreased, cutaneous erythema and nodules appeared at other sites repeatedly, and disappeared after the dose of corticosteroid was increased. The cutaneous lesions had characteristics of both Sweet's syndrome and pyoderma gangrenosum. Moreover, the patient had immunological abnormalities and decreased neutrophilic functions (chemotaxis and O2- generation). Thus, it was suggested that the cutaneous lesions of this patient could be diagnosed as "neutrophilic dermatosis of MDS", and corticosteroid was recognized to be very effective in treating these skin lesions.

Phase II study: treatment of non-Hodgkin's lymphoma with an oral antitumor derivative of bis(2,6-dioxopiperazine).

BACKGROUND: Although razoxane (ICRF-159), a derivative of bis(2,6-dioxopiperazine), has shown significant antitumor activity in several murine tumors, inadequate bioavailability has limited its clinical efficacy. Sobuzoxane (MST-16), another derivative of bis(2,6-dioxopiperazine), has shown activity against a broad spectrum of murine tumors and human tumor xenografts in nude mice and a lack of cross-resistance to vincristine, doxorubicin, cyclophosphamide, fluorouracil, etoposide, and mitomycin C. These findings suggest that MST-16 has a mode of cytocidal activity different from that of other antitumor agents. PURPOSE: The present late phase II study was conducted to evaluate the clinical efficacy and toxicity of MST-16 in non-Hodgkin's lymphoma (NHL). METHODS: As part of a multi-institutional cooperative study, we conducted a study of MST-16 in 27 patients with NHL who were assessable for drug efficacy and toxicity. MST-16, a bis(2,6-dioxopiperazine) analogue and an inhibitor of topoisomerase II, was administered orally at a dose of 1600 mg/m2 a day for 5-7 days at intervals of 2-3 weeks. RESULTS: Response consisted of one complete remission and seven partial remissions in 27 assessable patients. Response was achieved at a median of 13 days (range, 9-62 days) after initiation of therapy and lasted a median of 46 days (range, 29-155 days). Major toxic effects were leukopenia in 70% of the patients, thrombocytopenia in 44%, and gastrointestinal disorders in 37%. CONCLUSIONS: MST-16 was shown to be effective in NHL and deserves further clinical trial, since the drug shows little cross-resistance to available antitumor drugs. IMPLICATIONS: Phase II clinical studies of MST-16 in treatment of breast cancer, gastric cancer, and adult T-cell leukemia and lymphoma are also being conducted in Japan. Future trials of combination chemotherapy using MST-16 with other antitumor drugs are warranted in view of the additive effects observed in studies of MOLT-3 cells and studies of L1210 leukemia in mice.

[Late phase II study of MST-16 (sobuzoxane) on malignant lymphoma].

Late phase II trial of MST-16 for malignant lymphoma was conducted by the multi-institutions collaboration. Out of 34 patients entered, 29 were evaluated for efficacy as well as side effects. One complete response and 8 partial responses were achieved by the treatment of MST-16. The factors which affect the response rate were prior chemotherapies, stage of disease and performance status. The main toxicities were bone marrow suppression and G-I disorders. Leukopenia was observed in 72.4% of patients, thrombocytopenia in 44.8% and nausea/vomiting in 31.0%. Patients recovered from these side effects by discontinuation of the MST-16 therapy.

[Application of Y-chromosome specific DNA analysis for detection of mixed hematopoietic chimerism after allogeneic bone marrow transplantation].

We report a case involving mixed hematopoietic chimerism after an allogeneic bone marrow transplantation (BMT) from a sex mismatched donor. A 31 year-old-man who was diagnosed as having chronic myelogenous leukemia in the accelerated phase received an allogenic BMT from his HLA-identical sister in March, 1989. To determine the mixed chimerism we used the Y-chromosome specific repeated sequence of DNA using a specific probe (PHY 10). The donor's DNA 3.5 kb band appeared in 1-10% of male DNA by Southern blot hybridization in the peripheral blood 21 days after BMT. The Y-chromosome DNA band decreased day by day, and disappeared 110 days after BMT. The Y-chromosome DNA band could be detected, even though few metaphases were obtained immediately after BMT. Thus this method is very sensitive for determining which cells contain the Y-chromosome, and is therefore useful for detecting mixed chimerism after sex-mismatched BMT. Using this method the clinical significance of mixed chimerism can be assessed.

[Blastic form of acute erythremia: report of an autopsy case].

We report an autopsy case of acute erythremia which terminated in generalized infiltration of immature blasts similar to proerythroblasts. A 61-year-old man was admitted because of general fatigue and fever in June, 1990. Mild anemia and severe thrombocytopenia were noted. The bone marrow was hypocellular with 25.5% blasts similar to proerythroblasts and 36.5% erythroblasts, many of which were polynuclear and megaloblastoid. The blasts were cytochemically negative for POX, but positive for PAS staining. Therefore he was diagnosed as having acute erythremia. Partial remission was achieved by BHAC-EV therapy. But three months later, his marrow was replaced by 52.7% blasts as seen in admission. Those blasts were negative for lymphoid, myelocytic, megakaryocytic markers and antiglycophorin A, but positive for OKT 9. Electron microscopy revealed that some of blasts had characteristics of immature erythroblasts. In spite of low dose Ara-C therapy, he died of sudden gastrointestinal bleeding in December, 1990. The autopsy disclosed widespread infiltration of blasts, involving liver, spleen, lung, kidney and stomach. It was interesting that dysplasia had been confined to erythroid lineage throughout his clinical course. He seemed to be a rare case of blastic form of acute erythremia which should be distinguished from erythroleukemia.

[Clinical evaluation of cefbuperazone in severe infections complicated with hematological disorders].

The efficacy and the safety of an antibiotic in cephamycin group, cefbuperazone (CBPZ), were investigated in 93 patients with severe infections complicated with hematological disorders. The efficacy evaluation was made in 85 cases with underlying hematological disorders including 49 cases (57.6%) of leukemia and 18 cases (21.2%) of malignant lymphoma. The overall efficacy rate was 50.6% of the 85 evaluable cases. The clinical efficacy rate for sepsis and suspected sepsis was 53.4%. The most frequently used group of antibiotics for combination therapy was aminoglycosides in 37 cases, in which an efficacy rate of 62.2%, a higher rate than the efficacy rate of 48.5% for all the combination therapy cases, was obtained. In 16 cases in which penicillins were used as combination drug, the efficacy rate obtained was low, 31.3%. Efficacy rates obtained for cases with different neutrophil counts at the start of therapies were as follows: 52.2% in 23 cases with neutrophil counts below 100/mm3, 46.2% in 13 cases with neutrophil counts between 100 and 499/mm3 and 51.3% in 39 cases with neutrophil counts equal to or above 500/mm3, thus no significant differences in efficacy rates were observed for patients with different neutrophil counts. These results appear to suggest that CBPZ, alone or in combination with other antibiotic such as aminoglycosides, may be quite useful in the treatment of severe infections in patients with hematological disorders.

[Clinical evaluation of imipenem/cilastatin sodium in infectious complications of hematological malignancies. Tohkai Research Group on Infections in Hematological Disorders].

Imipenem/cilastatin sodium (IPM/CS), a newly developed carbapenem antibiotic, was administered to a total of 152 patients with severe infections complicating hematological disorders, of whom 138 patients are included in the present analysis of efficacy and 152 in that of safety. Most of the underlying diseases were acute leukemia (76/138), and most patients suffered from sepsis or suspicion of sepsis (84/138). Out of 138 patients in whom efficacy was evaluable, responses were excellent in 41 patients, good in 55, fair in 19, and poor in 23. The overall clinical efficacy rate was 69.6% (96/138). Prior antibiotic treatment and peripheral neutrophil count had significant effects on the clinical response. The overall eradication rate of bacteria was 76.2%. Adverse reactions were observed in 15 patients (9.9%) and abnormal laboratory test results in 19 patients (12.5%). From the above findings, IPM/CS is considered to be a useful antibiotic for the treatment of severe infections accompanying hematopoietic disorders.

A Novel Cell Line (NCU-L-1) from a Patient with Acute Lymphoblastic Leukemia, FAB, L3, Burkitt's Type.

A novel cultured cell line, NCU-L-1, was established from a 71-year-old Japanese woman with acute lymphoblastic leukemia, L3 FAB, Burkitt's type. The NCU-L-1 cells were shown to have a mature B-cell phenotype on the basis of immunologic surface marker analysis; including IgG Lambda surface immunoglobulins, CD19, CD20 and la-like antigen which were all detected on the cells. Intracytoplasmic immunoglobulin was not detected, but IgG was present in the cell culture supernatant. Cytogenetic studies revealed that the NCU-L-1 cells had t(2; 8) and an additional 14q+, a genotype which has not been identified previously in the usual Burkitt's cell lines. The NCU-L-1 cell line should prove to be useful for studying oncogenic events associated with the t(2; 8) translocation and karyotype evolution.

[T cell subsets as a prognostic factor in the chronic phase of chronic myelogenous leukemia].

A relationship between the survival time and T cell subsets in the chronic phase of chronic myelogenous leukemia (CML) studied by the method using monoclonal antibodies. No statistic difference between normal and CML was recognized in the rate of pan-T cell. The high T-Helper/Suppressor ratio (TH/TS) was revealed in the cases of the long survival time for more than six years.