Masuzu Ueda

Bortezomib overcomes cell adhesion-mediated drug resistance through downregulation of VLA-4 expression in multiple myeloma

Multiple myeloma (MM) is incurable, mainly because of cell adhesion-mediated drug resistance (CAM-DR). In this study, we performed functional screening using short hairpin RNA (shRNA) to define the molecule(s) responsible for CAM-DR of MM. Using four bona fide myeloma cell lines (KHM-1B, KMS12-BM, RPMI8226 and U266) and primary myeloma cells, we identified CD29 (β1-integrin), CD44, CD49d (α4-integrin, a subunit of VLA-4), CD54 (intercellular adhesion molecule-1 (ICAM-1)), CD138 (syndecan-1) and CD184 (CXC chemokine receptor-4 (CXCR4)) as major adhesion molecules expressed on MM. shRNA-mediated knockdown of CD49d but not CD44, CD54, CD138 and CD184 significantly reversed CAM-DR of myeloma cells to bortezomib, vincristine, doxorubicin and dexamethasone. Experiments using blocking antibodies yielded almost identical results. Bortezomib was relatively resistant to CAM-DR because of its ability to specifically downregulate CD49d expression. This property was unique to bortezomib and was not observed in other anti-myeloma drugs. Pretreatment with bortezomib was able to ameliorate CAM-DR of myeloma cells to vincristine and dexamethasone. These results suggest that VLA-4 plays a critical role in CAM-DR of MM cells. The combination of bortezomib with conventional anti-myeloma drugs may be effective in overcoming CAM-DR of MM.

Rituximab plus 70% cyclophosphamide, doxorubicin, vincristine and prednisone for Japanese patients with diffuse large B-cell lymphoma aged 70 years and older

Abstract In the rituximab era, several large studies have suggested that full-dose rituximab plus cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) might be the best treatment for patients with diffuse large B-cell lymphoma (DLBCL) aged 60 years and older. However, it remains unclear whether this is also the case for those aged 70 years and older. Previously untreated patients with DLBCL aged 70 years and older (elderly) were treated with R-70%CHOP, and patients younger than 70 years (younger) were treated with full-dose R-CHOP every 3 weeks, for a total of 6–8 cycles. Complete remission (CR) rates in elderly versus younger patients were 75 vs. 78% (p = 0.7), respectively. The 3-year overall survival, event-free survival and progression-free survival of elderly versus younger patients were 58 vs. 78% (p < 0.05), 45 vs. 70% (p < 0.05) and 64 vs. 72% (p = 0.43), respectively. Severe adverse events were more frequent in the elderly, even with the dose reduction in that age group. Three-year PFS with R-70%CHOP for patients aged 70 years and older was not significantly worse than that with full-dose R-CHOP for younger patients, suggesting that R-70% CHOP might be a reasonable choice for patients with DLBCL aged 70 years and older, especially for those with comorbidities.

The bone marrow hematopoietic microenvironment is impaired in iron-overloaded mice.

Increasing numbers of reports have described hematopoietic improvement after iron chelation therapy in iron‐overloaded patients. These observations indicate that excess iron could affect hematopoiesis unfavorably. To investigate how excess iron affects hematopoiesis in vivo, we generated iron‐overloaded mice and examined hematopoietic parameters in these mice.

Renal failure caused by plasma cell infiltration in multiple myeloma

We report on a case of severe renal failure in a 61-year-old female with multiple myeloma (MM). Two months prior to admission, the patient was diagnosed to have anemia and progressive renal failure associated with urinary Bence Jones protein and was referred to our hospital. A bone marrow biopsy revealed 40% plasma cells with κ light chain restriction. Thus, she was considered to have MM. A renal biopsy revealed neoplastic plasma cell infiltration within the kidney, moderate interstitial fibrosis, tubular atrophy, and punctate, electron-dense material along the peripheral capillary walls, tubular basement membrane, and in the interstitium of the kidney. This suggested that a combination of compression of the tubules and the microvasculature by the infiltrative process, and local light chain deposition-mediated tissue damage might be implicated in the development of renal failure in this patient. Despite a remission of bone marrow plasmacytosis with a bortezomib-based regimen, her renal function gradually deteriorated and a periodic hemodialysis program was finally required. Although the clinical impact of the direct kidney infiltration of neoplastic plasma cells on the longitudinal changes in renal function remains to be delineated, it is reasonable to consider that the infiltration of neoplastic plasma cells associated with local light chain depositions may result in irreversible renal injuries. Obviously, further studies and accumulation of additional experience with renal biopsy are required to better determine the precise and prognostic relationship between renal outcome and morphological alterations among MM patients with varying degrees of renal impairment.

Correlation between flow cytometric identification of CD33-positive cells and morphological evaluation of myeloblasts in bone marrow of patients with acute myeloblastic leukemia

Abstract Determination of the percentage of myeloblasts in bone marrow is important for the evaluation of acute myeloblastic leukemia (AML) and related disorders. Using flow cytometry with a CD45-blast gate (FCM/CD45), 226 bone marrow aspiration samples serially collected from 71 patients with de novo AML were analyzed. Bone marrow smears were evaluated independently by pathologists who did not know the corresponding flow cytometric data in advance. Patients received remission induction followed by consolidation. The CD33+ cell percentages evaluated by FCM/CD45 were strongly correlated to the myeloblast percentages determined by microscopic examination (r=0·8360, p<0·001). When only samples containing leukemic cells demonstrated by chromosomal or fluorescence in situ hybridization (FISH) analysis after induction were evaluated, positive correlations were found between CD33+ cell percentages determined by FCM/CD45 and myeloblast percentages determined by morphology (r=0·672, p<0·001). The identification of CD33+ cells by FCM/CD45 is useful for the evaluation of bone marrow myeloblasts in AML.

Prediction of response to imatinib in patients with chronic myelogenous leukemia by flow cytometric analysis of bone marrow blastic cell phenotypes.

Chronic myelogeneous leukemia (CML) is characterised by a specific translocation t(9; 22)(q34; q11), the Philadelphia (Ph) chromosome, giving rise to the novel BCR/ABL fusion gene [1]. The deregulated tyrosine kinase activity of the protein encoded by the fusion gene has been shown to be both necessary and sufficient for the initiation and maintenance of CML [1]. High response rates and favourable toxicities of imatinib in patients with newly diagnosed CML in chronic phase have been reported [2]. Imatinib induces a complete cytogenetic response (CCyR), i.e. disappearance of the Ph chromosome, in most patients with CML in chronic phase after one year of treatment with the drug [3–5]. However, some patients with CML in chronic phase who received treatment with imatinib show residual Ph chromosome after one year of treatment, and event-free survival in such patients was worse than that in patients showing a CCyR [3–5]. We analysed phenotypes of the cells in the blast region of bone marrow in CML by flow cytometry (FCM) and determined whether the phenotypes of the cells predict response to imatinib. During the period from January 2005 to April 2007, 32 patients in Jichi University Hospital were diagnosed as having CML in chronic phase. The diagnosis of CML in chronic phase was based on peripheral blood pictures, bone marrow pictures and existence of the Ph chromosome in bone marrow. Three-colour FCM combined with two-colour FCM was performed to evaluate phenotypes of the cells in the blast region of bone marrow before the beginning of imatinib therapy. Bone marrow mononuclear cells were stained with fluorescein isothiocyanateconjugated monoclonal antibody and/or phycoerythrin-conjugated monoclonal antibody and peridinin chlorophyll protein-conjugated CD45 [6,7]. Monoclonal antibodies used in this study were as follows: CD5, CD7, CD2, CD19, CD20, CD10, CD13, CD14, CD15, CD33, CD34, CD117, CD41, CD11b, CD11c, CD36, CD25, CD56, CD45 and HLA-DR. A gate was set for identifying immature cells characterised by intermediate CD45 expression and low side scatter properties [6,7]. Phenotypes of the cells in the blast region were analysed using a flow cytometer (FACSCalibur; BD Biosciences, San Jose, CA). Karyotypic analysis of bone marrow cells was performed using a standard Giemsa-banding method, and approximately 20 metaphases were analysed. BCR-ABL transcripts in bone marrow cells were measured using a real-time quantitative polymerase chain reaction assay (SRL Inc., Tokyo) [8]. All patients received treatment with imatinib only after the diagnosis of CML in chronic phase. To compare phenotypes of the cells in the blast region of bone marrow, bone marrow cells from patients with