IgH translocation with undefined partners is associated with superior outcome in multiple myeloma patients.

BACKGROUND: In multiple myeloma (MM), impact of specific chromosomal translocations involving IgH (14q21 locus, including t(4;14), t(11;14), and t(14;16)) has been explored extensively. However, over 15% MM patients harboring IgH translocation with undefined partners have long been ignored. METHODS: A prospective non-randomized cohort study with a total of 715 newly-diagnosed MM cases was conducted, 13.6% of whom were t(14;undefined) positive. The whole cohort was divided into four groups: no IgH split (47.7%); t(14;undefined) (13.6%); t(11;14) (17.6%); and t(4;14) or t(14;16) group (21.1%). RESULTS: Median OS for the four groups was 84.2, not reached (NR), 58.7, and 44.2 months, respectively, with P values for t(14;undefined) vs no IgH split, t(11;14), and t(4;14)/t(14;16) groups of 0.197, 0.022, and 0.001, respectively. In bortezomib-based group, the survival advantage gained by t(14;undefined) group was much more significant compared to t(11;14) and t(4;14)/t(14;16) groups. Importantly, t(14;undefined) turned out to be an independent predictive factor for longer OS of MM patients in multivariate analysis, especially in the context of bortezomib treatment. Similar results were also observed in the PUMCH external validation cohort. CONCLUSION: Collectively, our data confirmed and externally validated the favorable prognosis of the t(14;undefined) groups, especially in the era of novel agents.

[Treatment and Prognosis of Adult T Cell Acute Lymphoblastic Leukemia].

To analyze the treatment and prognosis of T cell acute lymphoblastic leukemia(T-ALL)in adults. Method The clinicobiogical and survival data of 68 adult patients with newly diagnosis T-ALL were retrospectively analzyed. Results The median age of these 68 patients was 23 years(14-60 years).T-ALL was more common in men(81%).After the first cycle of treatment,complete remission was achieved in 50 patients(73%).The highest complete remission(CR) rate was in patients with cortex T-ALL(100%),followed by other T-ALL(73%)and early T-cell precursor lymphoblastic leukemia(54%),(χ 2=5.712,P=0.058).The CR rate for adults aged >35 years was significantly lower than that of patients aged ≤ 35 years(40% vs. 79%,χ 2=6.364,P=0.012).The overall CR rate after the second treatment course was 93%.For patients treated with chemotherapy,autograft hematopoietic stem cell transplantation(auto-SCT),and allogeneic SCT,the median relapse free survival was 10 months,24 months,and not reached,respectively(P=0.002).The 5-year overall survival rate was 25% for all patients;for patients treated with chemotherapy,auto-SCT and allogeneic SCT,the median overall survival was 24 months,34 months,and 30 months,respectively(P=0.007),and the 5-year overall survival rate was 9%,33%,and 38%(P=0.037).Multivariate analysis showed leukocyte count ≥100×10 9/L was a risk factor for decreased relapse free survival(risk ratio 2.540,95%CI=1.058-6.099,P=0.037). Conclusion Adult T-ALL patients have poor prognosis,which may be improved by SCT.

Efficacy of Rituximab for Patients with Chronic Lymphocytic Leukemia.

Objective To evaluate the efficacy of rituximab in treating chronic lymphocytic leukemia (CLL). Methods The clinical data of CLL patients receiving fludarabine,cyclophosphamide±rituximab (with or without rituximab) regimen or cyclophosphamide,vincristine,and prednisone±doxorubicin±rituximab regimen in our hospital from March 2000 to February 2015 were analyzed retrospectively. Therapeutic efficacies and survivals of patients treated with different regimens were evaluated and compared. Results The complete response (CR) rate and the overall response rate (ORR) in 72 patients (43.6%) treated with rituximab were significantly higher than those treated without rituximab (38.9% vs. 21.5%,P=0.015;83.3% vs. 60.2%,P=0.001). The median PFS and OS for patients treated with rituximab were 53.0 (27.0-79.0) months and 112.0 (81.1-142.9) months,and the median PFS and OS for patients treated without rituximab were 28.0 (18.3-37.7) months and 89.0(72.0-106.0),but the results were not statistically significant (P=0.094,P=0.109). According to the cytogenetic features,patients were further divided into high-risk subgroup (with chromosome 17p deletion or 11q deletion) and non-high-risk subgroup. And in the high-risk subgroup,the ORR of patients treated with rituximab was 86.4%,which was significantly higher than that in patients treated without rituximab (53.3%)(P=0.012);in the non-high-risk subgroup,the PFS was marginally prolonged in patients treated with rituximab,but the difference was not statistically significant(P=0.050). Conclusions Compared with traditional chemotherapy,the chemoimmunotherapies with rituximab result in higher CR rate and ORR in CLL patients. In patients without 17p deletion or 11q deletion,the use of rituximab can marginally prolong PFS.

[The Optimal Storage Condition and Storage Time of Umbilical Cord Blood from Collection to Preparation].

OBJECTIVE: To explore the optimal storage condition and time of umbilical cord blood from collection to preparation. METHODS: Collect cord blood samples from 30 healthy newborns, with each new born's umbilical cord blood was divided into two parts on average. One part was stored in cold storage (4 ℃) and the other was stored at room temperature (20-24 ℃). Samples were taken at 24, 36, 48, 60 and 72 h, respectively, total nucleated cells (TNC) count and TNC viability was analyzed. Flow cytometry was used to detect the ratio of viable CD34+ cells to viable CD45+ cells and viability of CD34+ cells, and colony-forming unit-granulocyte-macrophage (CFU-GM) count was performed by hematopoietic progenitor cell colony culture. The change trend of each index over time was observed, and the differences in each index was compared between cold storage and room temperature storage under the same storage time. RESULTS: The TNC count (r 4 ℃=-0.9588， r 20-24 ℃=-0.9790), TNC viability (r 4 ℃=-0.9941， r 20-24 ℃=-0.9970), CD34+ cells viability (r 4 ℃=-0.9932， r 20-24 ℃=-0.9828) of cord blood stored in cold storage (4 ℃) and room temperature storage (20-24 ℃) showed a consistent downward trend with the prolongation of storage time. The percentage of viable CD34+ cells (r 4 ℃=0.9169， r 20-24 ℃=0.7470) and CFU-GM count (r 4 ℃=-0.2537， r 20-24 ℃=-0.8098) did not show consistent trends. When the storage time was the same, the TNC count, TNC viability, CD34+ cells viability and CFU-GM count of cord blood stored in cold storage were higher than those stored at room temperature. Under the same storage time (24, 36, 48, 60 or 72 h), TNC viability in room temperature storage was significantly lower than that in cold storage (P <0.001), but TNC count, percentage of viable CD34+ cells and CFU-GM count were not significantly different between room temperature storage and cold storage. When stored at room temperature for 24 h and 36 h, the viability of CD34+ cells was significantly lower than that in cold storage (P <0.001, P <0.01), when the storage time for 48, 60 and 72 h, there was no significant difference in the CD34+ cells viability between room temperature storage and cold storage. CONCLUSION: It is recommended that cord blood be stored in cold storage (4 ℃) from collection to preparation, and processed as soon as possible.

[Clinicopathologic features and differential diagnosis of splenic B-cell marginal zone lymphoma involving bone marrow].

OBJECTIVE: To study the clinicopathologic features and differential diagnosis of splenic B-cell marginal zone lymphoma (SMZL) involving bone marrow. METHODS: The clinical and pathologic features of 22 patients with SMZL were retrospectively studied. Immunophenotypic analysis was carried out by flow cytometry and immunohistochemistry. Immunoglobulin heavy chain rearrangement study was performed using polymerase chain reaction-based method. RESULTS: Villous lymphocytes were found in peripheral blood smears of 11/18 of the patients. In bone marrow aspirates, lymphocytosis (> 20%) was demonstrated in 15 cases (15/18) and villous lymphocytes in 6 cases (6/18). Flow cytometry showed CD19(+) CD20(+) FMC7(+) CD22(+) CD10(-) CD2(-) CD3(-) CD7(-) in 18 cases. Bone marrow biopsies of all the 22 patients revealed various degrees and patterns of neoplastic infiltration, as follows: mild (4 cases, 18.2%), moderate (11 cases, 50.0%) or severe (7 cases, 31.8%); intrasinusoidal (16 cases, 72.7%), interstitial (14 cases, 63.6%), nodular (11 cases, 50.0%) or diffuse (1 case, 4.5%). Reactive germinal center formation (CD23(+) bcl-2(-)) was found in 2 cases (91.0%). Immunohistochemical study showed the following results: CD20(+) PAX5(+) CD3(-) CD5(-) CD10(-) cyclin D1(-) CD23(-) CD43(-) Annexin A1(-) CD11C(-) CD25(-) in all the 22 cases, CD38(+) in 2 cases (9.1%) and CD138(+) in 2 cases (9.1%). CONCLUSIONS: Different and overlapping patterns of bone marrow involvement are observed in SMZL. As the histologic and immunophenotypic features are not specific to SMZL, distinction from other types of mature B-cell lymphomas is necessary.

[Regulation of miRNA-15a/-16 expression on the drug resistance of myeloma cells].

OBJECTIVE: To explore the effects and mechanism of bone marrow stromal cells (BMSCs) on the drug resistance of multiple myeloma (MM). METHODS: Fresh untreated MM patient (5 samples) and health donor (5 samples) bone marrow samples were collected from May 2011 to July 2011 at our hospital. Their BMSCs were separated respectively. The supernatant expression of cytokines in BMSCs was detected by enzyme-linked immunosorbent assay (ELISA). Myeloma cells were co-cultured with BMSCs and treated with melphalan or bortezomib. Cell proliferation and miRNA-15a/-16 expression of myeloma cells were measured in different culture conditions with thiazoyl blue tetrazolium bromide (MTT) assay and quantitative real-time PCR (qRT-PCR). miRNA-15a was transfected into myeloma cells. And the miRNA-15a functions on cell cycle and cell apoptosis were detected by flow cytometry. RESULTS: ELISA assay showed that cytokine IL-6 and VEGF were at higher levels in MM-BMSCs than healthy BMSCs ((189 ± 9) vs (115 ± 15) pg/ml, (1497 ± 40) vs (1239 ± 21) pg/ml, both P < 0.05). The miRNA-15a/-16 expressions of myeloma cells were up-regulated after the treatment of melphalan and bortezomib (all P < 0.05). However, the miRNA-15a/-16 up-regulation by melphalan or bortezomib became inhibited when MM cells were co-cultured with MM-BMSCs (melphalan: 4.690 ± 0.050 vs 34.440 ± 4.100, 0.760 ± 0.070 vs 12.030 ± 1.020, bortezomib: 1.440 ± 0.230 vs 11.480 ± 1.488, 0.880 ± 0.040 vs 3.680 ± 0.420, all P < 0.05). Furthermore, IL-6 suppressed the expression of miRNA-15a/-16 in a dose and time-dependent pattern. The transfection of miRNA-15a led to the arrest of MM cell cycle in G(1)/S phase. CONCLUSIONS: BMSCs suppress the proliferation of myeloma cells and regulate the drug sensitivity of myeloma cells through the inhibited expression of miRNA-15a/-16. IL-6 plays a pivotal role in the occurrence of drug resistance.

[Analysis of clinical and laboratory characteristics and survival with hairy cell leukemia].

OBJECTIVE: To explore the clinical and laboratory characteristics and survival of Chinese patients with hairy cell leukemia (HCL). METHODS: A total of 30 HCL patients from August 1990 to March 2012 were retrospectively analyzed. RESULTS: There were 22 cases with classical HCL (HCL-C) and 8 with variant HCL (HCL-V). Splenomegaly was the most common physical finding. Leukocytosis was found in all cases of HCL-V. But pancytopenia only accounted for 36.4% (8/22) in HCL-C. And 3/5 of HCL-V had abnormal chromosome karyotypes. Ribosomal-lamellae complexes (RLC) were found only in about 3/12 of HCL cases. Chemotherapy regimens including purine nucleoside analogues achieved a better complete remission (CR) rate than other regimens (3/4 vs 1/18, P = 0.012) in HCL-C. The median follow-up period was 27 (1 - 142) months. There was no follow-up loss. Eleven cases progressed and 6 died. The median overall survival (OS) was not reached. And the 1, 3, 6-year OS rates were 84%, 78% and 58% respectively. The median progression-free survival (PFS) was (63 ± 24) months and the 1, 2, 5-year PFS rates were 86%, 72% and 44% respectively. The median PFS of HCL-V was significant shorter than HCL-C ((23 ± 3) vs (78 ± 12) months, P = 0.014). In HCL-C group, fever (P = 0.038) and anemia (P = 0.000) at diagnosis were poor prognostic factors. But purine nucleoside analogues made no significant difference in PFS. CONCLUSIONS: Pancytopenia is infrequent in Chinese HCL patients. And classical RLC is rare under electron microscope. Purine nucleoside analogues may achieve a better CR rate, but fail to improve PFS rate. As compared with HCL-C, HCL-V is common with genetic abnormalities and has a worse prognosis with a shorter PFS.

[Richter syndrome: clinical characteristics and treatment experiences].

OBJECTIVE: To summarize the clinical characteristics of Richter syndrome and explore the methods of successful treatment and timely diagnosis. METHODS: Five patients with Richter syndrome in the last three years (from January 2009 to December 2011) were analyzed retrospectively at our hospital, including their clinical features and therapy before and after transformation. RESULTS: There were 4 males and 1 female with a median age on a diagnosis of chronic lymphocytic leukemia (CLL) at 47 (44 - 68) years. The median duration from a diagnosis of CLL to transformation was 52 (5 - 90) months. As for cytogenetic abnormalities, 3/4 patients had 17p deletion by fluorescence in situ hybridization (FISH). The clinical manifestations on transformation included regional enlargement of lymph node (n = 2) and systemic enlargement of lymph nodes (n = 3). All diagnoses were confirmed by lymph node biopsy and all transformed into diffuse large B cell lymphoma (classical transformation). The subgroups were germinal center B-cell like (GCB) (n = 3) and non-GCB (n = 1). After transformation, one patient underwent sibling allo-stem cell transplantation and survived 24 months until April 2012. Another patient with auto-stem cell transplantation relapsed and died 12 months later. One patient lost the treatment opportunity due to worsening condition. Another 2 patients gained partial remission after therapy and survived 20 and 8 months respectively. CONCLUSIONS: Richter syndrome may occur during a late stage of CLL. Such a high-risk cytogenetic abnormality as del17p may be correlated with transformation. Early identification and optimal therapy may extend the survival of Richter syndrome. Allo-stem cell transplantation remains a curable option.

[A preliminary study on the culture of single hematopoietic stem cell].

OBJECTIVE: To investigate the biological behavior including survival and proliferation of CD34 + CD38--Lin--cells when they are cultured at single cell level. METHODS: Purified umbilical cord blood CD34 + CD38--Lin--cells were separated at single cell level in 96-well plates using flow cytometry for four groups: control group (CD34 + CD38--Lin--cell plus stem cell medium) , Shh group (CD34 + CD38--Lin--cell plus stem cell medium and Shh), BMP-4 group (CD34 + CD38--Lin--cell plus stem cell medium and BMP-4), Jagged-1 group (CD34 + CD38--Lin--cell plus stem cell medium and Jagged-1). Methylcellulose medium was used in the colony-forming experiment which was also in four groups as previously. The number of cells and colony-forming units in each well for the four groups was evaluated at different time points (day 1, 3, 7) with fluorescence microscopy counting method. RESULTS: Division of single cell was observed to be amplified in all of these groups from day 3. And meanwhile, after 1-week culture, the survival rates for the treated groups were all higher than the control group (Jagged-1 group > BMP-4 group > Shh group > control), while the cell number in each well was also highest in the Jagged-1 group (Jagged-1 group > BMP-4 group > control). The number of wells with a cell number of zero was significantly fewer in all treated groups (especially the Jagged-1 group) than in the control group; meanwhile, the number of wells with a cell number higher than 17 was evidently higher in all the treated groups (especially the BMP-4 group) more than controls. Colony-forming units for erythroid (BFU-E), granulocyte (CFU-G), macrophage (CFU-M), and granulocyte macrophage (CFU-GM) were observed for all of these experimental groups, and there was no significant difference between the four experimental groups. CONCLUSIONS: CD34 + CD38 - Lin - cell can achieve the survival, self-renewal and proliferation when cultured at single cell level, and the adding of Shh, BMP-4, and Jagged-1 can enhance such capabilities. However, CD34 + CD38 - Lin - cell can only maintain cell totipotency in its niche.

[The Clinical Significance of Oligoclonal Bands in Patient with Multiple Myeloma].

OBJECTIVE: To investigate the clinical value of oligoclonal bands (OB) in patients with multiple myeloma (MM). METHODS: The laboratory test and clinical data of 624 newly diagnosed MM patients admitted to Blood Diseases Hospital of Chinese Academy of Medical Sciences from January 2013 to December 2019 were retrospectively analyzed, including 30 patients with OB, and the clinical characteristics, treatment effects and survival of OB and non-OB patients were analyzed and compared. RESULTS: OB occurred in 11.8% (22/187) of patients who received autologous stem cell transplantation(ASCT) and only 1.8% (8/437) of patients who did not receive ASCT (P=0.000). The median time to the appearance of oligoclonal bands was 3.2(0.6-10.5) months after transplantation. The M protein types of oligoclonal bands mainly include IgG κ, IgG λ, IgM λ and λ light chains. In the presence of oligoclonal bands, 90% of patients were evaluated as complete remission (CR) and above. There were no statistically significant differences in disease stage, tumor burden, and genetic abnormalities between OB and non-OB patients. Among the all patients, the prognosis of OB patients was significantly better than that of non-OB patients, and OB patients showed deeper disease remission (significantly higher CR rate, MRD negative rate, and longer MRD negative duration). Among patients who underwent ASCT, OB patients showed earlier immune recovery, but the depth of treatment response and survival outcomes were similar between OB and non-OB patients, it was no statistically difference. Although OB patients showed earlier immune reconstitution, this did not translate into better survival, suggesting that the better prognosis of OB patients was mainly related to deeper and durable remission rather than early immune reconstitution. Further analysis in patients who received ASCT and obtained MRD negative indicated that there was no additional survival benefit in patients with OB. CONCLUSION: The better prognosis of OB patients may be related to the deeper treatment response, but not to the early immune reconstitution. The appearance of OB is only a sign of deep remission and early immune reconstitution in patients, it cannot be translated into survival benefit of MM patients.

[The efficacy and safety of bortezomib plus thalidomide in treatment of newly diagnosed multiple myeloma].

OBJECTIVE: The aim of this phase II study was to determine the efficacy and safety of combined bortezomib and thalidomide (VT) regime as initial treatment for newly diagnosed multiple myeloma (MM) in China. METHODS: Thirty-four patients were enrolled in this study and received VT regime up to 21-day cycles. Bortezomib (1.3 mg/m(2)) was administered intravenously on days 1, 4, 8, and 11, while oral thalidomide (100 mg/day) was given from days 1 to 21. The primary end point was clinical response. The secondary end point was safety. RESULTS: Among the 34 patients, 20 were male, 14 were female, with a median age of 59 years, and 15 in international stage system (ISS) III, 18 in ISS II, 1 in ISS I. Among them, 28 completed 2 cycles' treatment and achieved an overall response rate (ORR) of 92.9%; 26 were able to complete the planned 8 cycles of therapy. After 8 cycles, the ORR was 100% (complete response 30.8%, near-complete response 23.1%, partial response 42.3%, minimal response 3.8%). After followed up with a median time of 12 months, the estimated rate without progress of disease was 62%, and the estimated continuous remission rate of 12 months was 62%. The median survival time was not achieved. The most common adverse events were mild to moderate (grades 1, 2). The main toxicities were hematologic (53.3%), gastrointestinal (40.0%), peripheral neuropathy (38.0%), fatigue (36.6%) and fever (32.0%). CONCLUSIONS: VT regime provides a very high ORR and complete response rate in the treatment of newly diagnosed MM patients. No patients experienced deep venous thrombosis. In conclusion, bortezomib in combination with thalidomide is a very effective regimen for newly diagnosed MM patients and the toxicities are manageable.

[Expression and prognostic significance of TOSO in CD19+ B cells from Chinese CLL patients].

OBJECTIVE: To investigate the expression and its prognostic significance of TOSO in CD19(+) B cells from Chinese chronic lymphocytic leukemia (CLL) patients. METHODS: The expression of TOSO was detected by absolute quantitative reverse transcription-polymerase chain reaction (RT-PCR) in a cohort of 85 untreated CLL patients from March 2006 to September 2010. Their status of immunoglobulin heavy chain variable (IGVH) somatic mutation, ZAP70 and CD38 were analyzed. RESULTS: The expression of TOSO was significantly elevated in CLL patients versus the healthy population (8.30 ± 2.99 vs 6.63 ± 1.22, P = 0.036) and other B cell lymphoproliferative diseases (8.30 ± 2.99 vs 7.12 ± 1.13, P = 0.023). The expression of TOSO was elevated in the IGVH non-mutated group versus the mutated group (9.87 ± 1.08 vs 7.61 ± 3.03, P = 0.000). The expression of TOSO was significantly elevated in Binet stage C patients versus in Binet stage B and Binet stage A patients (9.91 ± 3.03 vs 8.73 ± 1.86 vs 7.27 ± 2.83, P = 0.000). The expression of TOSO rose to (9.37 ± 2.12) in the chemotherapy group. And it was significantly higher than that in the observation group (7.19 ± 3.23, P = 0.001). At the cut-off point of 55 years old, the younger patients had a higher expression of TOSO than the elders (9.10 ± 2.06 vs 7.95 ± 3.22, P = 0.049). The expression of TOSO in ZAP70 and CD38-positive groups had no difference with those in the negative groups. CONCLUSION: The over-expressed TOSO in CLL cells is associated with a progressive disease. It may be an important prognostic factor for CLL.

[Expression of stereotyped B-cell receptor and its clinical value in chronic lymphocytic leukemia].

OBJECTIVE: To investigate the expression pattern of stereotyped B-cell receptor (BCR) and its prognostic significance in Chinese chronic lymphocytic leukemia (CLL) patients and evaluate the relationship to other prognostic markers. METHODS: Multiplex polymerase chain reaction (PCR) was used to identify the immunoglobulin variable heavy-chain (IGHV) segment and its mutation status in 116 CLL patients from April 1992 to April 2010. For CDR3-driven clustering, all in-frame IGHV-D-J rearrangements were aligned by the multiple sequence alignment software ClustalW2. RESULTS: There were 102 of 116 samples from newly diagnosed CLL were successfully analyzed. IGHV CDR3 genes were identified in 73/102 cases. Fourteen patients (19.2%) carried stereotyped BCR. A high percentage of carried stereotyped BCR was observed in IGHV non-mutated group versus mutated group (40.9% vs 11.8%, P = 0.005). Patients with stereotyped BCR had a higher frequency of deletion (17q) (33.3% vs 10.7%, P = 0.045). The median progression-free survival (PFS) in patients with stereotyped BCR was much shorter than other patients (39 vs 84 months, P = 0.002). CONCLUSION: The patients with stereotyped BCR have a poor prognosis. It highlights the importance of immunoglobulin mediated stimulation in the development of CLL.

[Expression of microRNA-223 and its clinical value in B lymphoproliferative disorders].

OBJECTIVE: To detect the expression of microRNA-223 and analyze its clinical value in B lymphoproliferative disorders. METHODS: Peripheral blood samples (n = 78) and bone marrow samples (n = 9) were collected from patients with chronic lymphocytic leukemia (CLL, n = 53), mantle cell lymphoma (MCL, n = 13), splenic marginal zone lymphoma (SMZL, n = 9) and healthy donors (n = 12) at our hospital from 2003 to 2010. Mononuclear cells were isolated and B cells purified with a CD19(+) magnetic-bead system. Total RNA was extracted from purified CD19(+) cells and the expression of microRNA-223 measured by TaqMan microRNA quantitative polymerase chain reaction (PCR). The clinical data of these patients were collected and their outcomes analyzed with SPSS 16.0 software. RESULTS: (1) The levels of microRNA-223 in CLL, MCL and SMZL were 4.58 ± 0.62, 4.03 ± 0.54 and 4.63 ± 0.57 respectively. And they were significantly lower than that in normal B cells (5.69 ± 0.60, P < 0.01). The expression of microRNA-223 decreased significantly in MCL versus CLL and SMZL (P < 0.05). There was no statistical difference between CLL and SMZL (P > 0.05). (2) The down-regulation of microRNA-223 was associated with disease aggressiveness in CLL. Patients with unmutated immunoglobulin heavy chain variable region (IgVH) expressed significantly a lower level of microRNA-223 (4.05 ± 0.69 vs 4.67 ± 0.51, P = 0.003). In 13q-negative patients, the expression of microRNA-223 decreased more significantly than that in 13q-positive patients (4.25 ± 0.67 vs 4.76 ± 0.45, P = 0.044). (3) Using receiver operating characteristic (ROC) curve analysis, the microRNA-223 cutoffs were defined according to the IgVH mutational status. The patients were divided into the positive and negative subgroups. The median progression-free survival (PFS) of microRNA-223 positive patient subgroup was 48 months. It was significantly longer than the negative subgroup (P = 0.001). In the microRNA-223 positive subgroup, no patient died at the end of follow-up. CONCLUSIONS: MicroRNA-223 may play an important role in the pathogenesis of B lymphoproliferative disorders. The down-regulation of microRNA-223 is associated with disease aggressiveness and poor prognostic factors in CLL. It may become a new reliable prognostic predictor.

[Cdc37 Expression in Multiple Myeloma and Its Role in Cell Proliferation].

OBJECTIVE: To investigate the expression of cell division cycle protein 37 (Cdc37) in multiple myeloma (MM) and its effect on MM cell proliferation. METHODS: The expression of Cdc37 mRNA in CD138+ cells derived from 63 newly diagnosed MM patients and 8 healthy people were detected by real-time quantitative PCR (RT-qPCR). Cdc37 was down-regulated by lentivirus in MM cell line NCI-H929. CCK-8 assay and soft agar clone formation assay were conducted to explore the role of Cdc37 on MM proliferation in vitro. To further verify the effect of Cdc37 on MM cell proliferation in vivo, NOD/SCID mice subcutaneous tumorigenesis model was established. Flow cytometry was carried out to explore the role of Cdc37 on cell cycle. Cell cycle associated proteins and NF-κB pathway were detected by Western blot (WB). RESULTS: Cdc37 was highly expressed in newly diagnosed CD138+cells compared with healthy people. After Cdc37 suppression by shRNA lentivirus infection in NCI-H929 cells, the proliferation of MM cells were decreased in vitro and in vivo. Compared with the control group, the ratio of cells arrested in G0/G1 phase significantly increased in NCI-H929-Cdc37 shRNA cells, the expression of cyclin D1 decreased, while the expression of p21 and p53 was significantly up-regulated. Meanwhile, the activation of NF-κB signaling pathway was hampered in NCI-H929-Cdc37 shRNA cells. CONCLUSION: Cdc37 is highly expressed in newly diagnosed MM patients. Inhibition of Cdc37 results in decreased proliferation activity and G0/G1 arrest in NCI-H929 cells. The possible mechanism may be to inhibit the activation of NF-κB signaling pathway.

[Clinical Analysis of Patients with MGUS, Primary Light Chain Amyloidosis, Multiple Myeloma or Multiple Myeloma with Concurrent Amyloidosis].

OBJECTIVE: To summarize and compare the clinical baseline characteristics of patients with monoclonal gammopathy of undetermined significance (MGUS), primary light chain amyloidosis (pAL), multiple myeloma (MM), or MM with concurrent amyloidosis, especially the differences in cytogenetic abnormalities. METHODS: The clinical data of 15 cases of MGUS, 34 cases of pAL, 842 cases of MM and 23 cases of MM with concurrent amyloidosis were analyzed and compared retrospectively. RESULTS: Cytogenetic statistics showed that the incidence of t (11; 14) in the four groups (MGUS vs pAL vs MM vs MM with concurrent amyloidosis) was 0%, 33.3%, 16.4%, and 15.8%, respectively (P=0.037); that of 13q deletion was 20.0%, 14.7%, 45.8% and 56.5%, respectively (P<0.001); gain of 1q21 was 50.0%, 12.5%, 47.4% and 40.9%, respectively (P=0.001). Proportion of pAL patients with 0, 1 and≥2 cytogenetic abnormalities (including 13q deletion, 17p deletion, 1q21 amplification and IgH translocation) accounted for 41.9%, 41.9% and 16.1%, respectively; while the proportion of the same category in MM was 17.6%, 27.3%, and 55.2% respectively; this ratio of MM with concurrent amyloidosis was more similar to MM. Subgroup analysis showed that genetic abnormalities (including 13q deletion, 17p deletion and 1q21 amplification) were comparable within t (11; 14) negative and positive groups. Compared with positive cases, t(11; 14) negative patients with MM or MGUS were more likely to have 13q deletions and multiple genetic abnormalities. CONCLUSION: Clinical characteristics of pAL, especially cytogenetic abnormalities, are significantly different from MM with concurrent amyloidosis. It suggests that although the onset characteristics are similar, actually the two diseases belong to different disease subtypes which should be carefully predicted and identified.

Treatment and outcome patterns of patients with Waldenström's macroglobulinemia: a large, multicenter retrospective review in China.

In this study, we aimed to investigate treatment options and the prognosis of patients with WM in China. This retrospective study included 1141 patients diagnosed with symptomatic WM between January 2003 and December 2019 at 35 tertiary hospitals in 22 provinces of China. Fifty-four patients (7.3%) received monotherapy, 264 (36.0%) received chemoimmunotherapy, 395 (53.8%) received other combination regimens without rituximab, and 21 (2.9%) received ibrutinib. Using a multivariable Cox regression model, age > 65 years old, platelets <100 × 109/L, serum albumin <3.5 g/dl, ß2 microglobulin concentration ≥4 mg/L and LDH ≥250 IU/L predicted poor OS. In summary, our study showed that frontline treatment choices for WM are widely heterogeneous. We validated most of the established prognostic factors in the rIPSS (age >65 years, LDH ≥250 IU/L, ALB <3.5 g/dl and ß2 microglobulin ≥4 mg/L) together with PLT ≤ 100 × 109/L indicate a poor prognosis for patients with WM.

[Effect of human umbilical cord mesenchymal stem cells on the CD34+ cells transplantation in NOD/SCID mice].

OBJECTIVE: To study the effect of human umbilical blood (UB) mesenchymal stem cells (MSC) on the CD34(+) cells transplantation in NOD/SCID Mice. METHODS: Umbilical blood CD34(+) cells (3.5 x 10(5) cells) alone or combined with umbilical cord MSC cells were transplanted into NOD/SCID mice that had been irradiated with (137)Cs (3.0 Gy) before transplantation. Changes in peripheral blood cells within 6 post-transplantation weeks were detected. The mice were sacrificed 6 weeks after transplantation. The human hematopoietic cells (hCD45(+)) and multi-lineage engraftment cells (CD3/CD19, CD33, CD14, CD61, and CD235a) in NOD/SCID recipients bone marrow, spleen, and peripheral blood were analyzed by flow cytometry. RESULTS: In the 3rd post-transplantation week, white blood cells (WBC), platelets (PLT), and red blood cells (RBC) began to increase in both two groups. In the 6th post-transplantation week, WBC and PLT counts in CD34(+) + MSC group reached peak levels and were significantly higher than CD34(+) alone group (P < 0.05), while RBC level was not significantly different between these two groups P > 0.05). hCD45(+) cell levels in bone marrow and peripheral blood were (42.66 +/- 2.57) % and (4.74 +/- 1.02) % in CD34(+) + MSC group, which were significantly higher than those in CD34(+) alone group [(25.27 +/- 1.67) % and (1.19 +/- 0.54) %, respectively, P = 0.006]. Also in the 6th post-transplantation week, the proportions of CD19(+), CD33(+), CD14(+), CD61(+), and CD235a(+) in CD34(+) + MSC group were significantly higher than those in CD34(+) alone group (P < 0.05), while the proportion of CD3(+) T lymphocyte in CD34(+) + MSC group was significantly lower than that in CD34(+) alone group (P = 0.003). The amplification of CD19(+) B lymphocyte was significantly higher than other blood cell lineages (P < 0.05). CONCLUSION: The co-transplantation of MSC cells and CD34(+) cells can promote hematopoietic stem cell transplantation and hematopoietic recovery in vivo.

[Clinicopathologic features of lymphoplasmacytic lymphoma].

OBJECTIVE: To explore the clinicopathologic features of lymphoplasmacytic lymphomas (LPL). METHODS: Routine histological examination was performed on hematoxylin-eosin stained sections of 24 bone marrow biopsies and available 6 concurrent lymph node specimens. Immunohistochemistry study was performed using EliVision methods. RESULTS: Among 24 cases, the male-to-female ratio was 2.4:1 and the median age was 59.5 years (42 - 75). The most common symptom was weakness (83.3%, 20/24). Hyperviscosity and "B" symptoms occurred in 20.8% (5/24) and 8.3% (2/24) respectively. 41.7% (10/24) patients presented with lymphadenopathy. Anemia, leukocytosis and thrombocytopenia were seen in 79.2% (19/24), 8.3% (2/24) and 37.5% (9/24) respectively. Monoclonal Ig light chain expression was detected by serum immunofixation electrophoresis in 23 cases (95.8%), including IgM (20 cases), IgG (2 cases) and IgA (1 case). Basing on the histology and immunohistochemistry findings, the diagnosis was made in 22 bone marrow and 2 lymph node biopsies, respectively. Histologically, the bone marrow and lymph node specimens composed of small lymphocytes, plasmacytoid lymphocytes and plasma cells. The most frequent pattern of bone marrow involvement was diffuse in appearance (63.6%, 14/22), while nodular and interstitial patterns were less common (22.7%, 5/22 and 13.6%, 3/22, respectively). Lymph node involvement was also to be diffuse in pattern. The proliferative cells expressed Pax5, CD20, CD38 and CD138, but were negative for CD5, CD10, CD23, CyclinD1, CD3, CD7 and MPO. CONCLUSIONS: LPL has distinct clinicopathological features. Histological and immunohistochemistry findings are important for its differential diagnosis with chronic lymphocytic leukemia/small lymphocytic lymphoma, splenic marginal zone lymphoma and follicular lymphoma. Waldenström macroglobulinemia is LPL.

TOSO interacts with SYK and enhances BCR pathway activation in chronic lymphocytic leukemia.

BACKGROUND: TOSO, also named Fas inhibitory molecule 3 (FAIM3), has recently been identified as an immunoglobulin M (IgM) Fc receptor (FcµR). Previous studies have shown that TOSO is specifically over-expressed in chronic lymphocytic leukemia (CLL). However, the functions of TOSO in CLL remain unknown. The B-cell receptor (BCR) signaling pathway has been reported to be constitutively activated in CLL. Here, we aimed to investigate the functions of TOSO in the BCR signaling pathway and the pathogenesis of CLL. METHODS: We over-expressed TOSO in B-cell lymphoma cell lines (Granta-519 and Z138) by lentiviral transduction and knocked down TOSO by siRNA in primary CLL cells. The over-expression and knockdown of TOSO were confirmed at the RNA level by polymerase chain reaction and protein level by Western blotting. Co-immunoprecipitation with TOSO antibody followed by liquid chromatography coupled with tandem mass spectrometry (IP/LCMS) was used to identify TOSO interacting proteins. Western blotting was performed to detect the activation status of BCR signaling pathways as well as B-cell lymphoma 2 (BCL-2). Flow cytometry was used to examine the apoptosis of TOSO-over-expressing B lymphoma cell lines and TOSO-down-regulated CLL cells via the staining of Annexin V and 7-AAD. One-way analyses of variance were used for intergroup comparisons, while independent samples t tests were used for two-sample comparisons. RESULTS: From IP/LCMS, we identified spleen tyrosine kinase (SYK) as a crucial candidate of TOSO-interacting protein and confirmed it by co-immunoprecipitation. After stimulation with anti-IgM, TOSO over-expression increased the phosphorylation of SYK, and subsequently activated the BCR signaling pathway, which could be reversed by a SYK inhibitor. TOSO knockdown in primary CLL cells resulted in reduced SYK phosphorylation as well as attenuated BCR signaling pathway. The apoptosis rates of the Granta-519 and Z138 cells expressing TOSO were (8.46 ±â€Š2.90)% and (4.20 ±â€Š1.21)%, respectively, significantly lower than the rates of the control groups, which were (25.20 ±â€Š4.60)% and (19.72 ±â€Š1.10)%, respectively (P < 0.05 for both). The apoptosis rate was reduced after knocking down TOSO in the primary CLL cells. In addition, we also found that TOSO down-regulation in primary cells from CLL patients led to decreased expression of BCL-2 as well as lower apoptosis, and vice versa in the cell line. CONCLUSIONS: TOSO might be involved in the pathogenesis of CLL by interacting with SYK, enhancing the BCR signaling pathway, and inducing apoptosis resistance.