Elevated erythrocyte adenosine deaminase activity in a patient with primary acquired sideroblastic anemia

We report a case of primary acquired sideroblastic anemia (PASA) associated with elevated erythrocyte adenosine deaminase (ADA) activity. The patient was an 85-year-old Japanese male. Analysis of the peripheral blood revealed pancytopenia, and the bone marrow findings showed marked ringed sideroblasts and chromosomal deletion (46XY, 11q-). The erythrocyte ADA activity was 17 times higher than that of normal control, the leukocyte ADA activity was within the normal range, and the plasma ADA activity was 2 times higher than the normal mean. The adenine nucleotides in the patient's erythrocytes were within normal range. According to starch gel electrophoresis, ADA isozyme of the patient was ADA 1. Western blotting showed an increased amount of ADA protein in the patient's erythrocytes. Southern blotting revealed no gene amplification or large structural change. Dot blot analysis of the reticulocyte mRNA showed no increase in the amount of ADA mRNA in the patient's reticulocytes compared with those of reticulocyte-rich controls. We considered that the mechanism of elevated ADA activity in this acquired defect was similar to that found in hereditary hemolytic anemia associated with ADA overproduction.     

A natural peptide, dolastatin 15, induces G2/M cell cycle arrest and apoptosis of human multiple myeloma cells

Several anti-cancer agents are derivative from natural products and microorganisms. The dolastatins are natural peptides derived from the marine mollusc Dolabella auricularia, which have recently been reported as an anti-cancer agent. Dolastatin 10 and 15 are small peptides; most preclinical studies have used dolastatin 10. It has been reported that dolastatins have cytotoxic activity by inhibiting microtubule assembly, and several clinical studies have already begun for solid tumors. However, the effects of dolastatin 15 against hematological malignancies such as myeloma cells have never been reported. We demonstrate here for the first time that dolastatin 15 induces cell cycle arrest at the G2/M phase followed by apoptosis in various human myeloma cell lines (RPMI8226, U266, and IM9), suggesting that it has effects on mitotic spindles. In addition, we showed that dolastatin 15 induces apoptosis of myeloma cells via activation of both mitochondrial- and Fas (CD95)/Fas-L (CD95-L)-mediated pathways. Our investigations have identified a novel inhibitor of microtubule assembly that induces mitotic arrest and apoptosis of myeloma cells. Therefore, it is possible that dolastatin 15 might be a novel and safe therapeutic agent for patients with multiple myeloma.     

Targeting NF-kappaB and induction of apoptosis by novel NF-kappaB inhibitor dehydroxymethylepoxyquinomicin (DHMEQ) in Burkitt lymphoma cells

A new NF-kappaB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), inhibited proliferation and induced apoptosis in human Burkitt lymphoma, HS-Sultan and Daudi cell lines. The activation of caspase-3 and the cleavage of caspase substrate PARP were observed after treatment with DHMEQ. The induction of apoptosis by DHMEQ was prevented by the pretreatment of Burkitt lymphoma cells with pan-caspase inhibitor, z-VAD-FMK. The expression of anti-apoptotic factors such as IAP-1 and XIAP was suppressed by DHMEQ. Phosphorylation of ERK and JNK was induced by DHMEQ. In conclusion, these results demonstrate that NF-kappaB might be an ideal target to develop for new anti-cancer drugs for Burkitt lymphoma.     

STI571-resistant KT-1 cells are sensitive to interferon-alpha accompanied by the loss of T-cell protein tyrosine phosphatase and prolonged phosphorylation of Stat1

OBJECTIVE: The high incidence of acquired drug resistance to STI571 during treatment of chronic myelogenous leukemia (CML) patients in blast crisis has become a problem. We studied the effects of interferon-alpha (IFN-alpha) on a novel STI571-resistant CML cell line and its molecular mechanisms in vitro. MATERIALS AND METHODS: KT-1 is a unique CML cell line that remains sensitive to the therapeutic IFN-alpha concentration. We developed novel STI571-resistant KT-1 cells (designated KTR cells) by gradually increasing the concentration of STI571. RESULTS: All seven KTR clones became more sensitive to IFN-alpha than KT-1 cells. IFN-alpha induced more prolonged phosphorylation of Stat1 for 24 hours in all seven KTR clones than in KT-1cells. Tyrosine phosphorylation of Jak1 in KTR cells was not prolonged compared to KT-1cells. T-cell protein tyrosine phosphatase (TC-PTP) was down-regulated in all KTR clones, and SH-PTP1 phosphatase also was down-regulated in some KTR clones. The transient transduction of TC-PTP cDNA into the KTR subline prevented the IFN-alpha-induced prolonged phosphorylation of Stat1 and recovered the sensitivity against IFN-alpha. These results indicated that the loss of TC-PTP is involved in the IFN-alpha-induced prolonged phosphorylation of Stat1 and in the higher sensitivity to IFN-alpha in KTR cells. CONCLUSION: We demonstrated that STI571-resistance does not confer cross-resistance to IFN-alphain KT-1 cells. The loss of TC-PTP contributed to the IFN-alpha-induced prolonged phosphorylation of Stat1 and the higher sensitivity to IFN-alpha in KTR cells.     

Erythroid crisis caused by parvovirus B19 transmitted via red blood cell transfusion

A 41-year-old man with hairy cell leukemia developed erythroid crisis after the transfusion of red cell concentrate. He was diagnosed with Parvovirus B19 infection based upon the presence of B19-specific IgM antibody and viral DNA in the sera. The repository sample from the corresponding red cell concentrate was negative for B19 antigen by red cell hemo-agglutination method, but was found to contain B19 virus DNA. Furthermore, a genomic PCR direct sequencing showed that B19 in both patient's sera and repository sample were identical. This is the first report directly demonstrating the transmission of B19 through B19 antigen-negative red cell concentrate transfusion. Further accumulation of the cases is warranted to estimate its incidence and to reconsider the screening methods of blood products.     

Characteristics and prognosis of synchronous multiple early gastric cancer

AIM:To assess the clinicopathologic characteristics,risk factors,and prognosis for synchronous multiple early gastric cancer(SMGC).METHODS:A total of 146 patients with SMGC and1194 patients with single gastric cancer who had undergone gastrectomy between 1989 and 2008 were retrospectively analyzed to determine their clinicopathologic characteristics and postoperative survival.Tumors were classified into groups on the basis of location and histology.Smoking habits were evaluated using the Brinkman index.Clinical and pathological factors were compared using either Fisher’s exact test or Pearson’sχ2test.Logistic regression analysis was performed to identify independent risk factors.Survival rate was calculated using the Kaplan-Meier method.RESULTS:SMGCs accounted for 10.9%of gastric cancer cases and occurred predominantly in elderly male patients with a family history of gastric cancer who were both smokers and drinkers.These tumors were typically macroscopically elevated and histologically differentiated.There were no significant differences between SMGC and single gastric cancer patients with respect to tumor location,tumor size,lymph node metastasis,the number of metastatic lymph nodes,venous invasion,or tumor stage(P=0.052,P=0.347,P=0.595,P=0.805,P=0.559,and P=0.408,respectively).Further,there was no significant difference in postoperative survival between the patient groups(P=0.200).Of the 146SMGC patients,a single patient had remnant cancer.CONCLUSION:A careful preoperative endoscopy is necessary for patients who are at high risk of SMGC,and minimally invasive treatment may be indicated in some cases.     

Regulation of manganese superoxide dismutase and other antioxidant genes in normal and leukemic hematopoietic cells and their relationship to cytotoxicity by tumor necrosis factor

Myeloid cells are a major source of superoxide and other oxygen metabolites. As a protective mechanism, cells express antioxidant enzymes including manganese superoxide dismutase (Mn-SOD), copper-zinc SOD (Cu/Zn-SOD), and glutathione peroxidase (GSX-PX). Even though hematopoietic cells are a major source of oxidants, little is known of their expression of antioxidants. We found that seven myeloid leukemic cell lines blocked at different stages of differentiation constitutively expressed Mn-SOD, Cu/Zn-SOD, and GSX-PX RNAs. Level of Mn-SOD activities paralleled levels of Mn-SOD RNA. Terminal differentiation of native HL-60 cells to either granulocytes or macrophages did not alter levels of Mn-SOD RNA but markedly decreased cell division. Myeloid leukemic lines sensitive to cytotoxic effects of tumor necrosis factor (TNF) as well as normal peripheral blood lymphocytes and monocytes, dramatically increased their levels of Mn- SOD RNA in the presence of TNF. In contrast, Cu/Zn-SOD and GSX-PX RNA levels did not increase in these same cells. TNF-resistant leukemic lines had higher constitutive levels of Mn-SOD RNA and activity; and these levels did not change in the presence of TNF. Antisense but not random oligonucleotides to Mn-SOD markedly increased the sensitivity to the inhibitory effects of TNF for both the native HL-60 (TNF-sensitive) and K562 (TNF-resistant) cell lines. Further studies showed that the antisense oligonucleotides entered the cells and resulted in decreased levels of Mn-SOD RNA. The data suggest that Mn-SOD may provide protection against cytotoxicity of TNF in hematopoietic cells.     

Quality assurance of MLC leaf position accuracy and relative dose effect at the MLC abutment region using an electronic portal imaging device

We investigated an electronic portal image device (EPID)-based method to see whether it provides effective and accurate relative dose measurement at abutment leaves in terms of positional errors of the multi-leaf collimator (MLC) leaf position. A Siemens ONCOR machine was used. For the garden fence test, a rectangular field (0.2 × 20 cm) was sequentially irradiated 11 times at 2-cm intervals. Deviations from planned leaf positions were calculated. For the nongap test, relative doses at the MLC abutment region were evaluated by sequential irradiation of a rectangular field (2 × 20 cm) 10 times with a MLC separation of 2 cm without a leaf gap. The integral signal in a region of interest was set to position A (between leaves) and B (neighbor of A). A pixel value at position B was used as background and the pixel ratio (A/B × 100) was calculated. Both tests were performed at four gantry angles (0, 90, 180 and 270°) four times over 1 month. For the nongap test the difference in pixel ratio between the first and last period was calculated. Regarding results, average deviations from planned positions with the garden fence test were within 0.5 mm at all gantry angles, and at gantry angles of 90 and 270° tended to decrease gradually over the month. For the nongap test, pixel ratio tended to increase gradually in all leaves, leading to a decrease in relative doses at abutment regions. This phenomenon was affected by both gravity arising from the gantry angle, and the hardware-associated contraction of field size with this type of machine.