Role of autotransplantation in the treatment of acute promyelocytic leukemia patients in remission: Fukuoka BMT Group observations and a literature review.

We retrospectively analyzed the outcomes of 26 patients with acute promyelocytic leukemia (APL) in the first CR (CR1) or second CR (CR2), who underwent autologous PBSCT (auto-PBSCT) between 1992 and 2008. All patients received all-trans retinoic acid-based induction therapy. After two courses of consolidation chemotherapy, upfront auto-PBSCT was performed in 20 patients in the CR1. Five patients had a high WBC count of more than 10 × 10(9)/L (high risk), while 15 patients had a count of less than 10 × 10(9)/L (low risk) at initial presentation. In addition, six patients, who were considered as low-risk patients at presentation, had a relapse after three cycles of consolidation and 2 years of maintenance therapy, but gained the molecular remission after re-induction and consolidation, and underwent auto-PBSCT in the CR2. In 26 recipients, engraftment was rapid and no TRM was documented. All 20 patients autotransplanted in CR1 were still in CR at a median of 133 months (73-193 months), and six patients who underwent auto-PBSCT in CR2 were also still in CR at a median of 41 months (2-187 months) without maintenance therapy. PML/RARα chimeric mRNA was undetectable in PBSC or BM samples examined before auto-PBSCT. Despite a small number of cases studied, our retrospective observations suggest that auto-PBSCT may be an effective treatment option to continue durable CR in the treatment of high-risk APL. We review previous reports and discuss the role of autotransplantation in the treatment of APL patients in CR.

Polyvalent horse F(Ab`)2 snake antivenom: Development of process to produce polyvalent horse F(Ab`)2 antibodies anti-african snake venom

A method to obtain polyvalent anti-Bitis and polyvalent-anti-Naja antibodies was developed by immunizing horses with B. arietans, B. nasicornis, B. rhinoceros, N. melanoleuca and N. mossambicacrude venoms. Antibody production was followed by the ELISA method during the immunization procedure. Once the desired anti-venom antibody titers were attained, horses were bled and theimmunoglobulins were separated from the sera by (NH4)2SO4 precipitation, cleaved with pepsin and filtered through a 30 kDa ultrafiltration membrane. F(ab´)2 fragments were further purified by Q-Fast Flow chromatography, concentrated by molecular ultrafiltration and sterilized by filtration through 0.22 m membranes. The resulting F(ab´)2 preparations were rich in intact L and in pieces of H IgG(T) chains, as demonstrated by electrophoresis and Western blot and exhibited high antibody titers, as assayed bythe ELISA method. In addition, the preparations possess a significant capacity to neutralize the lethalityof venoms, as estimated by ED50 determination in mouse assay and are free of toxic substances, pyrogen and bacterial or fungal contaminations.

Prognostic significance of S-phase kinase-associated protein 2 and p27kip1 in patients with diffuse large B-cell lymphoma: effects of rituximab.

BACKGROUND: The F-box protein S-phase kinase-associated protein 2 (Skp2) positively regulates the G1-S transition by promoting degradation of the cyclin-dependent kinase inhibitor p27(kip1) (p27). Recent evidence has indicated an oncogenic role of Skp2 in not only carcinogenesis but also lymphomagenesis. MATERIALS AND METHODS: Clinicopathologic features and immunohistochemical expression of Skp2 and p27 were studied retrospectively in 671 patients treated with cyclophosphamide, vincristine, doxorubicin and prednisolone (CHOP) or cyclophosphamide, vincristine, doxorubicin and prednisolone plus rituximab (R-CHOP). The median follow-up periods were 43.2 months in the CHOP group (n = 425) and 24.0 months in the R-CHOP group (n = 246). RESULTS: High Skp2 or low p27 expression correlated significantly with poor overall survival (OS) and progression-free survival (P < 0.001) in both treatment groups. The prognostic value of Skp2 or p27 expression was independent of the parameters included in the International Prognostic Index by multivariate analysis. Patients with high Skp2 expression in combination with low p27 expression showed the worst survival. CONCLUSIONS: Addition of rituximab to the CHOP regimen did not provide a beneficial outcome to patients with diffuse large B-cell lymphoma with high Skp2 expression and low p27 expression. Skp2 and p27 may be useful prognostic markers in the rituximab era.

Demonstration of reversed flow in segmental branches of the portal vein with hand-held color Doppler ultrasonography after hematopoietic stem cell transplantation.

Hepatic veno-occlusive disease (VOD) is a severe complication of hematopoietic stem cell transplantation (SCT). When monitored with hand-held color Doppler ultrasonography during day -7 to +35 around SCT, reversed blood flow in the segmental branches of the portal vein was detected in nine of 56 patients who had undergone SCT. Three of nine patients had clinical evidence of VOD, but six patients did not fulfill the criteria for diagnosis of VOD initially. Two patients progressed to clinical VOD at a later date and the reversed portal flow disappeared with or without treatment for VOD in the other four patients. Monitoring for reversed portal flow with color Doppler ultrasonography may be a useful tool for the early diagnosis of VOD, and may improve prognosis by allowing early initiation of treatment.

Efficient engraftment of primary adult T-cell leukemia cells in newborn NOD/SCID/beta2-microglobulin(null) mice.

Adult T-cell leukemia (ATL) develops via multiple oncogenic steps in human T-cell leukemia virus type I (HTLV-I) carriers. To better understand pathogenesis of ATL, we developed a novel xenogeneic engraftment model in which primary ATL cells are intravenously transplanted into neonatal nonobese diabetic (NOD)/severe-combined immunodeficiency (SCID)/beta2-microglobulin(null) (NOD/SCID/beta2m(null)) mice. Acute-type ATL cells engrafted in the peripheral blood and in the lymph nodes of recipients at a high efficiency. Engrafted ATL cells were dually positive for human CD4 and CD25, and displayed patterns of HTLV-I integration identical to those of donors by Southern blot analysis. These cells infiltrated into recipients' liver, and formed nodular lesions, recapitulating the clinical feature of each patient. In contrast, in smoldering-type ATL cases, multiple clones of ATL cells engrafted efficiently in NOD/SCID/beta2m(null) mice. When smoldering-type ATL cells were retransplanted into secondary NOD/SCID/beta2m(null) recipients, single HTLV-I-infected clones became predominant, suggesting that clones with dominant proliferative activity can be competitively selected in this xenogeneic system. Taken together, the NOD/SCID/beta2m(null) newborn system is useful to understand kinetics, metastasis, and disease progression of ATL in vivo.

Association of MBL gene polymorphisms with major bacterial infection in patients treated with high-dose chemotherapy and autologous PBSCT.

A growing body of evidence indicates that genetic factors are involved in an increased risk of infection. We investigated whether mannose-binding lectin (MBL) gene polymorphisms that cause low levels of MBL are associated with the occurrence of major infections in patients, mainly bearing hematological malignancies, after high-dose chemotherapy (HDT) rescued by autologous peripheral blood stem cell transplantation (auto-PBSCT). A retrospective evaluation of 113 patients treated with HDT and auto-PBSCT revealed that the low-producing genotypes, B/B and B/LXA, were associated with major bacterial infection (P=0.0016, OR 7.9). We next performed a nation-wide large-scale study to assess the allele frequency of the MBL coding mutation in a total of 2623 healthy individuals in Japan. The frequency of allele B was estimated to be approximately 0.2, almost the same in seven different areas of Japan. This common occurrence suggests that MBL deficiency may play an important role in the clinical settings of immunosuppression.

Cidofovir for treating adenoviral hemorrhagic cystitis in hematopoietic stem cell transplant recipients.

Adenovirus (AdV) infection is an important cause of morbidity and mortality in hematopoietic stem cell transplant (HSCT) recipients. We treated 16 patients with AdV hemorrhagic cystitis (HC) following HSCT with cidofovir (CDV; 1 mg/kg/day, three times weekly for 3 weeks). Patients included 10 males and six females with a median age of 50 years (range 10-62). Two of the 16 patients were unevaluable because of early death from nonadenoviral causes. CDV therapy cleared AdV from urine in 12 of 14 patients (86%). Of 14 patients, 10 (71%) showed clinical improvements in HC. Among 14 patients, seven (50%) had avoided renal damage, the most important CDV toxicity. One patient previously treated with foscarnet for cytomegalovirus (CMV) required hemodialysis, and CDV treatment was discontinued. In another patient, CDV treatment was discontinued because of grade 2 nephrotoxicity. Four patients became positive for CMV antigenemia while being treated with CDV, and two developed herpes simplex virus (HSV) stomatitis while being treated with CDV. CDV proved effective in treating AdV HC in transplant patients. However, CDV at 1 mg/kg/day given three times weekly failed to prevent breakthrough infection with CMV and HSV in some patients.

Aggressive natural killer-cell leukemia revisited: large granular lymphocyte leukemia of cytotoxic NK cells.

Aggressive natural killer-cell leukemia (ANKL) is a rare form of large granular lymphocyte leukemia, which is characterized by a systemic proliferation of NK cells. The clinical features of 22 ANKL cases were analyzed. Hepatomegaly (64%), splenomegaly (55%) and lymphadenopathy (41%) were also frequently observed. Leukemic cells were identified as CD1-, CD2+, surface CD3-, CD4-, CD5-, CD7+, CD8+/-, CD10-, CD11b+/-, CD13-, CD16+, CD19-, CD20-, CD25-, CD33(-), CD34-, CD38+, CD56+, CD122+, HLA-DR+ and TCR-. Two of the 16 cases examined for CD57 were positive and three of the seven cases examined for cytoplasmic CD3. Epstein-Barr virus was detected in the tumor cells of 11 of the 13 cases examined. No common cytogenetic abnormalities were identified and 6q anomaly was detected in only one. Three of 13 patients treated with chemotherapy containing anthracycline/anthraquinone attained complete remission, in contrast to none of the eight who were treated with regimens without anthracycline. Although the overall prognosis was poor with a median survival of 58 days, those who attained remission showed better prognosis (P=0.005). These findings suggest that ANKL is an entity of mature cytotoxic NK-cell neoplasms with distinct phenotype and disease presentations. Intensive treatment for ANKL may result in a better prognosis.

Clinical significance of cytomegalovirus (CMV) antigenemia in the prediction and diagnosis of CMV gastrointestinal disease after allogeneic hematopoietic stem cell transplantation.

To evaluate the clinical significance of a cytomegalovirus (CMV) antigenemia assay in the prediction and diagnosis of CMV gastrointestinal (CMV-GI) disease after hematopoietic stem cell transplantation (HSCT), 19 allogeneic HSCT recipients developing CMV-GI disease were retrospectively reviewed. All patients were monitored by a CMV antigenemia assay, at least once weekly after engraftment. The median onset of CMV-GI disease occurred 31 days post transplant (range: 19-62). Only four of 19 patients (21%) developed a positive CMV antigenemia test before developing CMV-GI diseases. Although all 19 patients subsequently developed positive CMV antigenemia tests during their clinical courses, the values remained at a low-level in nine (47%) patients. Among the 14 patients in whom results of real-time polymerase chain reaction (PCR) were available, seven (50%) yielded positive results of real-time PCR before developing CMV-GI disease. In contrast to the values of CMV antigenemia, all 14 patients exclusively yielded high viral loads (median: 2.8 x 10(4) copies/ml plasma). We conclude that CMV antigenemia testing has limited value in prediction or early diagnosis of CMV-GI disease, and that real-time PCR could have a more diagnostic significance.

Allogeneic peripheral blood stem cell transplantation from two- or three-loci-mismatched related donors in adult Japanese patients with high-risk hematologic malignancies.

With the increasing frequency of haploidentical transplantation, it is becoming more important to establish the degree of HLA mismatch that can be accepted. We retrospectively analyzed clinical data of 50 adult Japanese patients with high-risk hematologic malignancies who underwent allogeneic peripheral blood stem cell transplantation (PBSCT) from two- or three-loci-mismatched related donors with HLA class I and II gene disparities in the graft-versus-host direction. They were treated at 20 transplant centers between 1996 and 2002. In all, 18 patients received unmanipulated PBSC, while 32 received purified CD34+ blood cells. Conventional (n=31) or reduced-intensity (n=19) conditioning regimens were used. Of the 39 patients (78%) who survived for > or =28 days after transplant, 37 (95%) achieved neutrophil engraftment, while graft failure and rejection occurred in two of 39 (5%) and three of 37 (8%) patients, respectively. Stepwise Cox regression analysis revealed a significantly lower incidence of grades II-IV acute GVHD in patients receiving purified CD34+ cells (hazard ratio 0.32; 95% CI 0.12-0.84; P=0.022). By 1 year post transplant, 28 patients (56%) had died of transplant-related problems, including infectious complications (30%). Although the number of patients is small, our data suggest that transplant-related problems, particularly infectious complications, are major obstacles to the success of this therapy.

Influence of transplanted dose of CD56+ cells on development of graft-versus-host disease in patients receiving G-CSF-mobilized peripheral blood progenitor cells from HLA-identical sibling donors.

We investigated effects of variations in the cellular composition of G-CSF-mobilized peripheral blood progenitor cell (G-PBPC) allografts on clinical outcomes of allogeneic PBPC transplantation. We retrospectively analyzed transplanted doses of various immunocompetent cells from 27 HLA-identical sibling donors in relation to engraftment, incidence of graft-versus-host disease (GVHD), and survival. Significant variability was documented in both absolute numbers and relative proportions of CD34+, CD2+, CD3+, CD4(high)+, CD4+25+, CD8(high)+, CD19+, CD56+, and CD56+16+ cells contained in these allografts. Stepwise Cox regression analysis revealed that the CD56+ cell dose was significantly inversely correlated with the incidence of GVHD. Thus, there was a significantly higher incidence of grade II acute GVHD in patients receiving a lower CD56+16+ cell dose (hazard ratio (HR) 0.0090; 95% confidence interval (CI), <0.00001-3.38; P=0.031), a higher incidence of chronic GVHD in those receiving allografts with a lower CD56+16+ to CD34+ ratio (HR <0.00001; 95% CI <0.00001-0.0007; P=0.0035), and a higher incidence of extensive chronic GVHD in those receiving allografts with a lower CD56+ to CD34+ ratio (HR <0.00001; 95% CI <0.00001-0.053; P=0.0083). These results suggest that CD56+ cells in G-PBPC allografts from HLA-identical sibling donors may play an important role in preventing the development of GVHD.

Development of rheumatoid arthritis following autologous peripheral blood stem cell transplantation.

A 51-year-old man with non-Hodgkin's lymphoma (NHL) was treated with high-dose chemotherapy followed by autologous peripheral blood stem cell transplantation (PBSCT). Although he had HLA-DRB1 0405 and a positive rheumatoid factor, he was unlikely to develop rheumatoid arthritis (RA) according to diagnostic criteria. However, the patient developed RA 40 days after transplantation. Our experience suggests that the systemic autoimmune disease, RA, may occur in patients with predisposing factors after autologous PBSCT.

Genetically determined interferon-gamma production influences the histological phenotype of lupus nephritis.

OBJECTIVE: To clarify whether the interferon-gamma (IFN-gamma) gene (IFNG) is associated with the histological phenotype of lupus nephritis. METHOD: We analysed microsatellite polymorphisms located within the first intron of the IFNG gene to determine the genotypes of patients with lupus nephritis WHO class IV (n=24), patients with WHO class V (n=12) and healthy controls (n=61). We used flow cytometric detection of intracellular cytokines to identify CD4(+) T cells producing IFN-gamma. Production of IFN-gamma by peripheral blood mononuclear cells after stimulation with phytohaemagglutinin was evaluated with an enzyme-linked immunosorbent assay. RESULT: The frequency of the IFNG allele 114 was significantly greater in WHO class V patients than in WHO class IV patients. Furthermore, the IFNG 114 +/+ genotype was more frequent in WHO class V than in WHO class IV patients. The level of IFN-gamma and the percentage of IFN-gamma-producing CD4(+) T cells were lower in individuals with genotype 114 +/+ than in individuals with genotype 114 -/-. CONCLUSION: The IFN-gamma gene is associated with the histological phenotype in lupus nephritis.

Monitoring of cytomegalovirus reactivation after allogeneic stem cell transplantation: comparison of an antigenemia assay and quantitative real-time polymerase chain reaction.

Cytomegalovirus (CMV) antigenemia and quantitative real-time polymerase chain reaction (PCR) were compared for monitoring of CMV reactivation after allogeneic stem cell transplantation. The number of CMV antigen-positive cells by the antigenemia assay and the level of CMV DNA by real-time PCR correlated well. The sensitivity and specificity of the antigenemia assay was 55.4% and 95.5%, respectively, using real-time PCR as the reference standard. The probability of positive antigenemia at day 100 was 76.5%, with a median of first detection at day 37 in 51 patients, compared with a positive PCR of 84.3% and day 33, respectively. When HLA-identical sibling donor transplant recipients and other donor transplant recipients were analyzed separately, there was no difference between the two tests. However, temporal patterns of first detection of CMV antigen-positive cells and CMV DNA differed between HLA-identical and alternative recipients; patients without CMV (29%) or with sporadic positive PCR results (14%) were more common in HLA-identical sibling transplants, whereas patients with simultaneous antigenemia and positive PCR occurred more in alternative transplants (48%). Two of 51 patients (4%) developed CMV colitis despite antigenemia-guided prophylaxis, but both were successfully treated with ganciclovir. Although PCR is more sensitive than antigenemia, both tests are useful in the early detection of CMV after allogeneic stem cell transplantation.

Chronic lymphocytic leukemia (CLL) is rare, but the proportion of T-CLL is high in Japan.

Chronic lymphocytic leukemia (CLL) is a rare disease in Japan. Recent advances in molecular biology, diagnostic criteria and classification of CLL have reinforced the concept of each category of CLL as a distinct entity. Since there have been no recent studies on the incidence and prevalence of CLL in Japan, the Kyushu Hematology Organization for Treatment (K-HOT) Study Group conducted two studies of CLL. One study is a prospective registration of newly diagnosed hematological disorders, which gave us some idea of the incidence of CLL in our region (Kyushu island) where adult T-cell leukemia is endemic. A total of 677 patients with hematological disorders were registered over a 6-month period and 11 patients were diagnosed as having CLL among 182 leukemia patients. This amounts to 6% of all leukemias, which is twice as frequent as previously reported in Japan. The other study is a retrospective analysis of CLL. Eleven institutions of the K-HOT Group analysed their diagnostic records of chronic lymphoid leukemia, and 145 patients with CLL were found over a period of 3-12 yr. After the data were reviewed 11 patients were excluded through having a different type of leukemia. The proportion of chronic B-cell lymphoid leukemia was 73% (98/134), while that of T-cell leukemia was 18% (24/134). The proportion of T-cell chronic leukemia was 5-6 times higher than that in Western countries. Two institutions had a complete database on hematological disorders. From this database, the annual incidence of CLL was estimated to be 0.48 per 100 000. Thus, the incidence of CLL in Japan is at least 4-5 times lower than that in Western countries, suggesting that chronic B-cell leukemia is really rare, but chronic leukemia of T-cell lineage develops in Japan as frequently as in Western societies. Further investigation is required to delineate why the incidence of B-CLL is so low in Japan.

Donor leukocyte infusion for Japanese patients with relapsed leukemia after allogeneic bone marrow transplantation: indications and dose escalation.

To clarify the role of dose escalation of donor leukocyte infusion (DLI) in the treatment of relapsed leukemia after allogeneic bone marrow transplant (BMT), data from 100 patients were collected from 46 facilities in Japan and analyzed with respect to indications and infused cell dose. Complete remission (CR) was achieved in 11 of 12 (91%) patients with relapsed chronic myelogenous leukemia (CML) in the chronic phase, 3 of 11 (27%) with CML in the acute phase, 8 of 21 (38%) with acute myelogenous leukemia (AML), 6 of 23 (25%) with acute lymphoblastic leukemia (ALL), and 5 of 11 (45%) with myelodysplastic syndrome (MDS). The probability of remaining in CR at 3 years was 82% in CML patients in the chronic phase, but 0% in those with CML in the acute phase, 7% in those with AML, 0% with ALL, and 33% with MDS. Acute graft-versus-host disease (GVHD) (> or = 2) developed in 31 of 89 (34%) patients with human leukocyte antigen identical related donors and was fatal for 7 (7%). A leukocyte dose of 1 x 10(7)/kg of recipient body weight with CML in the chronic phase, 3 x 10(7)/kg of recipient body weight with MDS, and 1 x 10(8)/kg of recipient body weight with acute leukemia appeared to be optimal as an initial dose of DLI. However, the minimal dose of leukocyte developing fatal GVHD was 7 x 10(7)/kg of recipient body weight. These suggest that a relatively small dose of DLI ranging from 1 x 10(7)/kg to 5 x 10(7)/kg of recipient body weight should be administered initially then the infused escalating dose 2 or 3 months later in patients with CML in the chronic phase and MDS. However, a large number of leukocytes around 1 x 10(8)/kg are needed to induce graft versus leukemia effects in patients with acute leukemia despite a 7% fatality in GVHD.

New diagnostic approach to intracystic lesions of the breast by fiberoptic ductoscopy.

Intracystic tumors of the breast are uncommon and, at the time of ultrasonography and aspiration cytology, it is difficult to distinguish cancer from a benign tumor. The Fiberoptic Ductoscopy System (FDS) is an emerging technique allowing direct visual access to the ductal system of the breast. FDS was inserted through the cannulae into the cavity and we observed the intracystic tumors (3 intracystic papillomas and 2 intracytsic papillary carcinomas). The appearance of the malignant tumors was irregular, rough-shaped and they tended to bleed. On the contrary, benign tumors had smooth surfaces without bleeding. Cytological findings showed malignant cells in one out of two breast cancer patients. In addition, in the immunohistochemical study of resected tumor tissues from 5 patients, we observed positive reactions with anti-ErbB-2 antibody in 2 intracystic papillary carcinomas. In contrast, none of the histologically confirmed benign lesions (3 intracystic papillomas) gave positive results. In conclusion, the use of FDS as a non-invasive technique may provide valuable information.