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Background Multi-parametric flow cytometry (MFC) is a helpful tool for detecting neoplastic cells in malignant lymphoma; however, lymphoma cells can be difficult to detect when characteristic immunophenotypic abnormalities are not evident. We evaluated the stainability of VS38, which is used for multiple myeloma, in normal and abnormal B cells using MFC to develop a new strategy for detecting lymphoma cells. Methods We compared the median fluorescence intensity of VS38 staining in lymphocytes from patients without hematopoietic neoplasms and in B cells from 26 patients with B cell lymphoma (BCL). To evaluate the performance of VS38 gating, we compared VS38-positive B cells with the percentages of BCL cells, and with the mutation ratios of MYD88 L265P measured by droplet digital PCR in patients with lymphoplasmacytic lymphoma (LPL)/Waldenstrom macroglobulinemia (WM). Results CD27-positive memory B cells were stained with VS38, whereas normal lymphocytes were faintly stained. Lymphoma cells were stained with VS38 in 11 of 12 patients with LPL/WM, 3 of 3 with chronic lymphocytic leukemia, 3 of 5 with mantle cell lymphoma, 2 of 4 with follicular lymphoma, and 1 of 1 with splenic marginal zone lymphoma. The percentages of VS38-positive B cells in VS38-positive BCL were equivalent to those of lymphoma cells and the mutation ratios of MYD88 L265P in LPL/WM. Conclusions VS38 identified neoplastic cells in plasma cell disorders and BCL. This might improve the accuracy of BCL diagnosis, especially in patients with LPL/WM.

Background SF3B1 (splicing factor 3B subunit-1) somatic mutation is specifically detected in myelodysplastic syndrome (MDS) with ring sideroblasts (MDS-RS). We investigated the sensitivity and utility of SF3B1 mutation analysis as a clinical laboratory test. Method Detection limit for SF3B1 mutations by high-resolution melting (HRM) analysis was investigated by plasmid mixture. In 67 MDS patients, we examined the association between SF3B1 mutation and prognostic evaluation using the Revised International Prognostic Scoring System and revalidated MDS classifications based on the revised 4th edition of the WHO classification. Results HRM analysis enabled mutation detection in the 12.5% SF3B1 mutant alleles. SF3B1 mutation was detected in 9 cases, mostly in the low-risk group. Cases of MDS with ring sideroblasts unrelated to SF3B1 mutation were detected in the high-risk group. Two cases were reclassified as MDS-RS after detecting SF3B1 mutation. Conclusions SF3B1 mutation analysis as an initial screening at diagnosis increases the accuracy of prognostic prediction and disease classification.

Purpose: The aim of this study was to investigate the efficacy of combination therapy of ionizing radiation (IR) and adenoviral p53 gene therapy and to evaluate its molecular mechanisms. Methods and Materials: Two human prostate cancer cell lines, DU145 and PC-3 cells, containing different types of p53 gene mutations, were investigated. The recombinant adenovirus vector containing the wild-type p53 gene (Ad5CMV- p53 ) was used for this study. Cells were irradiated (in 0, 2, 4, and 6 Gy, 300 cGy/min) and after 12 h of irradiation, the cells were infected with various doses of Ad5CMV- p53 (0–40 multiplicity of infection [MOI]). Cytotoxicity was determined by clonogenic assay. The molecular mechanisms were evaluated by quantitative reverse transcriptase–polymerase chain reaction (RT-PCR), apoptotic cell detection, and cell cycle analysis. Results: The cell growth inhibition in DU145 ( p53 -mutated) cells by IR was strongly enhanced by additional Ad5CMV- p53 infection in a viral dose–dependent manner. In DU145 cells, IR alone induced minimal p53 mRNA expression. However, IR combined with Ad5CMV- p53 infection stimulated significant increase in p53 mRNA expression supplemented with Bax and p21 mRNA expressions. In PC-3 ( p53 -null), IR induced Bax and p21 mRNA expression, while the combination effects were observed in p53, Bax , and p21 mRNA expression. Apoptotic cell deaths were rarely observed after IR alone (DU145: 3%, PC-3: 5%). However, after combination therapy, the proportion of apoptotic cells greatly increased (sevenfold in DU145 cells, and twice in PC-3 cells). G1 cell cycle arrest was observed after Ad5CMV- p53 infection and the combination in both cell lines. Conclusion: In this study, we demonstrated that the combination of IR and Ad5CMV- p53 gene therapy resulted in remarkable synergistic effects in human prostate cancer cells. This combination therapy could be one of the optimal treatment strategies for radioresistant prostate cancer.

Abstract 3648 Background: Radioimmunotherapy (RIT) with 90Y-ibritumomab tiuxetan has been used for the treatment of relapsed or refractory indolent B-cell lymphoma. Early prediction of response to the therapy may offer the potential to identify patients who will benefit this therapy. We evaluated the efficacy of fluorine 18 fluorodeoxyglucose (FDG) combined positron emission tomographic-computed tomographic (FDG-PET/CT) imaging for assessment of therapy and predicting outcome in 90Y-ibritumomab tiuxetan radioimmunotherapy. Methods: Radioimmunotherapy with 90Y-ibritumomab tiuxetan was preformed in 52 patients with relapsed or refractory indolent B cell lymphoma (follicular lymphoma, 45; mucosa-associated lymphoid tissue lymphoma, 5; lymphoplasmacytic lymphoma, 2) at Hyogo College of Medicine Hospital from June 2009 through August 2012. FDG-PET/CT scanning was performed at two weeks and the later after 90Y-ibritumomab tiuxetan therapy. Results: In FDG-PET at 2 weeks after 90Y-ibritumomab therapy, 26 (50%) showed complete response (defined as early CR), 20 (38%) showed partial response (PR-2W) and 6 (12%) showed non response (NR-2W). Further follow-up revealed 10 later CR (8 CR out of 20 PR-2W and 2 CR out of 6 NR-2W), showing 69 %(36) of overall CR rate. Relapse was occurred in 4(17%) of 26 early CR and in 6 (60%) of 10 later CR, indicating significantly low relapse late in early CR (P= 0.0074, by Chi-square test). Patients with early CR (CR in 2 weeks after the RIT) showed significantly better progression free survival than those with later CR (P=0.0046, by Logrank test). In contrast, analysis at six weeks after the RIT showed 36 CR patients (10 of which eventually relapse), but failed to discriminate patients who had high risk of relapse. Conclusion: Our results suggest that CR assessed by FDG-PET/CT at 2 weeks after 90Y-ibritumomab tiuxetan therapy discriminates good responder to the RIT and predicts better progression free survival in relapsed or refractory indolent B cell lymphoma. Disclosures: No relevant conflicts of interest to declare.

A regimen which incorporates cyclophosphamide, doxorubicin, vincristine and prednisolone (CHOP) is the standard treatment for patients with non-Hodgkin's lymphoma (NHL), but it has not been effective in patients with aggressive NHL who are at high risk. The aim of the present trial was to investigate the feasibility of high-dose chemotherapy (HDC) without stem cell support as a first-line treatment. The primary endpoint was a complete remission rate. The second endpoint was survival. Fourteen patients with aggressive NHL entered the study and were treated according to the K93 protocol (3 cycles of CHOP, high-dose etoposide and ifosfamide, and high-dose methotrexate) Eleven patients (79%) achieved complete remission (CR) and two (14%) achieved partial remission (PR). Overall survival (OS) after five years was 79%. The actuarial five year disease free survival (DFS) for the eleven cases of CR was 75%. During chemotherapy, grade IV hematologic toxicity was observed in all patients and grade IV non-hematologic toxicity in only one patient, who experienced oral ulcers. Peripheral blood stem cell (PBSC) apheresis was performed in eight cases. One harvesting was enough to provide an adequate number of CD34+ cells for the subsequent PBSC transplantation (PBSCT). In conclusion our study confirmed the efficacy of the K93 protocol in obtaining a good response (CR + PR) rate and a very good DFS rate in most cases of aggressive NHL, with acceptable toxicity. This regimen may improve the outcome in cases of aggressive NHL without stem cell support. It seems worthwhile to conduct a randomized controlled study comparing the K93 protocol with the standard CHOP regimen.