Mine Harada

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.

Characterization of non-small-cell lung cancer cell lines established before and after chemotherapy

We established several in vitro drug-resistant cell lines after continuous, long-term exposure of each drug to elucidate mechanisms of drug resistance. Whether drug resistance in these in vitro resistant cell lines reflects clinical drug resistance still remains unanswered. In this study, a pair of lung cancer cell lines was established from one patient with squamous cell carcinoma of the lung, with one line being established before and one line after combination chemotherapy (cisplatin/ifosfamide/vindesine). Combination chemotherapy selected resistant EBC-2/R cells, which showed cross-resistance to 4-hydroxyifosfamide (3.2-fold), cisplatin (2.3-fold), and methotrexate (3.7-fold) and collateral sensitivity to vindesine (0.77-fold) compared with parent EBC-2 cells. EBC-2/R cells showed decrease in intracellular accumulation of cisplatin, increase in intracellular concentration of glutathione (GSH), and overexpression of multidrug resistance-associated protein (MRP) 3 when compared with EBC-2 cells. A single cycle of chemotherapy was not sufficient to select other mechanisms of drug resistance, such as multidrug resistance-1/P-glycoprotein, MRPs 1, 2, 4, and 5, lung resistance-related protein, metallothionein IIa, glutathione S-transferase pi, gamma-glutamylcysteine synthetase (light and heavy chain), and excision repair cross complementing 1. Sequentially we established two cell lines, which cell lines showed the differences of the cisplatin resistance, expression level of MRP3, intracellular GSH level and intracellular accumulation of cisplatin. A pair of cell lines will be useful to elucidate resistant mechanisms of cisplatin in heterogeneous lung cancer cells.

Simultaneous induction of matrix metalloproteinase-9 and interleukin 8 by all-trans retinoic acid in human PL-21 and NB4 myeloid leukaemia cells

Summary. All‐trans retinoic acid (ATRA) has been shown to induce differentiation of human acute promyelocytic leukaemia (APL) cells and eventual elimination of the malignant clone. Matrix metalloproteinase‐9 (MMP‐9) is produced by neutrophils and its expression appears to be linked with myeloid cell differentiation. We investigated effects of ATRA on MMP expression in two human myeloid leukaemia cell lines, PL‐21 and NB4. Both cells could differentiate into neutrophils after exposure to ATRA. Both the activity and antigen levels of MMP‐9 were much higher in NB4 cells than in PL‐21 cells. Stimulation with ATRA significantly increased MMP‐9 levels approximately three‐ to fivefold in both PL‐21 and NB4‐conditioned media. MMP‐9 mRNA levels increased in ATRA‐treated cells and was almost in parallel with the increase in MMP‐9 activity, suggesting that ATRA induced MMP‐9 by activating its gene expression. ATRA can induce interleukin 8 (IL‐8) in APL cells. IL‐8, chemokine for neutrophils and a potent inducer of MMP‐9, was also induced by ATRA in PL‐21 cells. However, recombinant IL‐8 did not induce MMP‐9 expression. In addition, a neutralizing antibody against IL‐8 did not inhibit ATRA‐induced MMP‐9 expression in either cell type. These observations suggest that ATRA can induce both MMP‐9 and IL‐8, but IL‐8 is not involved in ATRA‐induced MMP‐9 expression. As MMP‐9 can truncate and activate IL‐8, simultaneous induction of MMP‐9 and IL‐8 by ATRA could activate leucocytes excessively, causing the hyper‐inflammatory events in retinoic acid syndrome.

Predictive value of circulating immature cell counts in peripheral blood for timing of peripheral blood progenitor cell collection after G–CSF plus chemotherapy-induced mobilization

BACKGROUND: Enumeration of CD34+ cells in peripheral blood (PB) before apheresis predicts the number of CD34+ cells collected, although flow cytometric techniques used are complex and expensive. In an attempt to determine the optimal timing for peripheral blood progenitor cell (PBPC) collection, the usefulness of circulating immature cell (CIC) counts in PB was evaluated.

Expression of cutaneous lymphocyte antigen is associated with a poor outcome of nasal-type natural killer-cell lymphoma

Summary. Nasal and nasal‐type natural killer (NK) lymphoma is a distinct clinicopathological entity mostly associated with Epstein–Barr virus. Cases that have widespread lesions are resistant to ordinary anti‐cancer therapy and take a highly aggressive course. To date, there are no available data on the relationships between the localization, clinical outcome and expression of adhesion molecules in such cases. We examined the expression of cutaneous lymphocyte antigen (CLA) in 52 cases of NK‐cell lymphoma. CLA was highly expressed in cutaneous cases. Also, the CLA+ group (n=29) had a much worse prognosis than the CLA– group (n=23), regardless of the primary site or clinical staging. Univariate analysis identified some significant prognostic factors, and multivariate analysis of these factors showed that the expression of CLA was an independent prognostic indicator. In conclusion, the present findings established that CLA is an independent and important prognostic factor in patients with NK‐cell lymphomas.

Multilineage involvement in hypereosinophilic syndrome terminating in granulocytic sarcoma and leukaemic transformation with trisomy 8

Summary. We report a patient with hypereosinophilic syndrome (HES), which, 8 years later, transformed into granulocytic sarcoma in the brain and, subsequently, into acute myelocytic leukaemia. Repeated chromosome analyses showed a normal karyotype, until the time of leukaemic transformation when trisomy 8 was confirmed in cells from the bone marrow and cerebrospinal fluid. The combined techniques of May–Grunwald–Giemsa staining and fluorescence in situ hybridization identified trisomy 8 not only in blasts and eosinophils but also in neutrophils and erythroblasts. Our observation suggests that HES is a multilineage myeloproliferative disorder involving precursors of at least the eosinophil, neutrophil and erythroid lineages.

Therapy-related chronic myelogenous leukaemia following autologous stem cell transplantation for Ewing's sarcoma

Summary. A 17‐year‐old Japanese woman with Ewings sarcoma was initially treated with conventional chemotherapy and local irradiation, and then with high‐dose chemotherapy supported by autologous peripheral blood stem cell transplantation. Four years later she was diagnosed with chronic myelogenous leukaemia (CML). The BCR/ABL fusion gene was detected in both peripheral blood and bone marrow cells by reverse transcription–polymerase chain reaction, but not in the harvest product of peripheral blood stem cells which were infused at the time of transplantation. This case adds to the accumulating evidence of therapy‐related CML developing after high‐dose chemotherapy and autologous stem cell transplantation.

A Filter That Prevents the Spread of Mail-Attachment-Type Trojan Horse Computer Worms

The malicious code “W32/Sircam” is spread via e-mail and potentially through the file space shared by local area networks. Such Trojan-horse-type computer worms easily penetrate Internet firewalls and propagate via the “backdoor” to other computers. Once a malicious code, such as “W32/Sircam,” has been executed on a system, it may reveal or delete confidential data, such as clinical records. In order to protect against the leakage of clinical records, we devised a mail filter that successfully prevents the spread of mail containing this malicious code. It is significant that neither access control nor packet filtering is guaranteed to prevent the spread of this mail-attachment-type Trojan horse computer worm.