Regulatory Effect and Mechanism of Erythroblastic Island Macrophages on Anemia in Patients with Newly Diagnosed Multiple Myeloma.

Objective: To examine the clinical characteristics and anemia-related factors in patients with newly diagnosed multiple myeloma (NDMM), as well as the effect and mechanism of erythroblastic islands (EBIs) and EBI macrophages in NDMM patients with anemia. Methods: We collected and analyzed clinical data to find anemia-related factors. Using flow cytometry, the numbers and ratios of erythroblasts and EBI macrophages were determined. RNA sequencing (RNA-seq) was used to determine the differences of EBI macrophages in NDMM patients with or without anemia. Results: Based on the clinical characteristics of NDMM patients with anemia, MCV, abnormal levels of albumin, osteolytic lesions, and Durie-Salmon (DS) stage are risk factors for anemia. Patients with anemia have fewer erythroblasts, erythroblastic islands (EBIs), and EBI macrophages in their bone marrow than patients without anemia. RNA-seq analysis of EBI macrophages from the bone marrow of patients with and without anemia revealed that macrophages from patients with anemia are impaired and tend to promote the production of interleukin-6, which has been demonstrated to be an essential survival factor of myeloma cells and protects them from apoptosis. Conclusion: In NDMM patients with anemia, EBI macrophages are impaired, which causes anemia in those patients. Our finding highlights the significance of EBI macrophages in anemia in NDMM patients and provides a new strategy for recovery from anemia in these patients.

Efficacy and safety of pomalidomide and low-dose dexamethasone in Chinese patients with relapsed or refractory multiple myeloma: a multicenter, prospective, single-arm, phase 2 trial.

BACKGROUND: Pomalidomide in combination with dexamethasone has demonstrated positive results in patients with relapsed or refractory multiple myeloma (RRMM), but no data are available in China. We conducted a multicenter, single-arm trial to examine the efficacy and safety of bioequivalent generic pomalidomide plus low-dose dexamethasone in Chinese RRMM patients. METHODS: Adult (≥ 18 years of age) RRMM patients who progressed after at least two previous treatments, including bortezomib and lenalidomide, were eligible. Pomalidomide was given orally at 4 mg/day on days 1 to 21 of a 28-day cycle. Dexamethasone was given at 40 mg/day (either orally or intravenously; 20 mg/day at 75 years or older) on days 1, 8, 15, and 22 of each cycle. Treatment continued until disease progression or intolerable adverse events (AEs). The primary end point was objective response rate (ORR). RESULTS: Seventy-four patients were enrolled between February 2017 and February 2019. All patients had progressed within 60 days of their last therapy. 74.3% of the patients were resistant to lenalidomide, 31.1% had renal insufficiency and 33.8% had high-risk cytogenetic RRMM. The median follow-up duration was 33.0 months (range 31.1-34.8 months). The ORR was 37.8% in the overall analysis, 32.7% in lenalidomide-refractory patients, 36.0% in patients with high-risk cytogenetics and 34.8% in RRMM patients with renal impairment. The median progression-free survival was 5.7 months (95% CI 3.7-8.8 months). The median overall survival was 24.3 months (95% CI 14.4-41.1 months). The most common grade 3 and 4 treatment-emergent adverse events (TEAEs) were neutropenia (63.5%), leukopenia (37.8%), thrombocytopenia (28.4%), and anemia (31.1%). Pulmonary infection (27.0%) was the most frequent grade 3 and 4 nonhematologic TEAE. No previously unreported AEs were observed. No venous thromboembolism was reported. CONCLUSIONS: Pomalidomide in combination with low-dose dexamethasone is effective and safe in Chinese RRMM patients. TRIAL REGISTRATION: The study is registered at Chinese Clinical Trial Registry (ChiCTR) ( ChiCTR-OIC-17013234 , first registered on 03/11/2017).

[Research Progress of Clonal Hematopoiesis of Indeterminate Potential in Multiple Myeloma --Review].

With the progress of medical technology, cloning hematopoietic was found to be widely exist in normal people. Because of its clinical significance and prognosis is unclear, it is named clonal hematopoiesis of indeterminate potential(CHIP), which has been detected in blood diseases such as myelodysplastic syndrome and lymphoma, and proven to be related to poor prognosis. Recently, CHIP has been also detected in patients with multiple myeloma (MM). In this article, the definition and influencing factors of CHIP, clinical significance, prognosis and treatment in MM were reviewed.

[PX-12 Promoting Apoptosis of Multiple Myeloma Cell Line H929 Induced by Bortezomib].

OBJECTIVE: To study the effect of PX-12 on apoptosis of multiple myeloma (MM) cell line induced by bortezomib. METHODS: MM cell line H929 cells were divided into PX-12 group, bortezomib group, combination group, and control group. 5.0 µmol/L PX-12, 20 nmol/L bortezomib, combination of the two drugs, and DMSO were given to the above mentioned group, respectively. After culture for 24, 48, and 72 hours, the changes of cell viability were observed, the MM cell activity was detected by MTT method, and the cell cycle distribution and apoptosis of each group was detected by flow cytometry. The intracellular ROS level was measured by H2DCFDA probe labeling. RESULTS: MTT assay showed that after culture for 72 hours, the activity of H929 cells in PX-12 group (P<0.05) and bortezomib group (P<0.01) was significantly lower than that in the control group, while that in the combination group was decreased most significantly (P<0.01). After culture for 48 hours, cells in G1 phase in PX-12 group was decreased to 40%, while cells in S phase and G2/M phase was increased to 28% and 40%, respectively. The cells in bortezomib group also showed a similar distribution after being treated. After treated with PX-12 and bortezomib, the cells in G1 phase were decreased significantly to 19% and 12% in S phase, but increased significantly to 68% in G2/M phase, which was significantly different from PX-12 group and bortezomib group (P<0.01). After culture for 72 hours, the apoptosis rate was 71.3% in the combination group, which was significantly higher than that in PX-12 group, bortezomib group, and control group (20.6%, 33.3%, 10.6%)(P<0.01). After culture for 24 hours, the intracellular ROS level in the combination group was 12015±430.2, which was higher than that in the PX-12 group, bortezomib group, and control group (6729±352.8, 2651±228.3, 1098±164.6, respectively) (P<0.01). CONCLUSION: PX-12 can increase the apoptosis of MM cell line H929 induced by bortezomib, which may be caused by increasing of ROS level.

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.

Analysis of the efficacy and safety of bortezomib for treating newly diagnosed multiple myeloma through different administration methods.

BACKGROUND: This study aims to compare the efficacy and adverse reactions of bortezomib for treating newly diagnosed multiple myeloma (MM) through two different administration methods: intravenous (IV) injection and subcutaneous (SC) injection. METHODS: A retrospective analysis was performed in 205 patients with newly diagnosed MM, who were treated by the Department of Hematopathology, Henan Cancer Hospital, from June 2009 to December 2017. These patients were divided into two groups according to the treatment methods: IV injection group, IV injection of bortezomib; SC injection group, SC injection of bortezomib. RESULTS: After the first course of treatment, the effect of very good partial remission (VGPR) or above (≥VGPR) in the IV injection group (IV group) and SC injection group (SC group) was 31.0% and 14.3%, respectively (P=0.004), while the overall response rate (ORR) was 72.0% and 49.5%, respectively (P=0.001). From the 2nd course to the 6th course of treatment, the ORR was not statistically different between these two groups. No significant difference was found in median progression-free survival (37 vs 45 months) and overall survival (63 vs 59 months). A lower frequency of adverse events, especially Grade 3 peripheral neuropathy, was observed in SC group compared with the IV group. CONCLUSION: Compared with IV administration, SC bortezomib can provide a better balance between efficacy and toxicity.

The role of long non-coding RNAs in multiple myeloma.

Multiple myeloma (MM) is still an incurable disease, and its pathogenesis involves cytogenetics and epigenetics. In recent years, the roles of long non-coding RNAs (lncRNAs) in MM have been deeply studied by scholars. LncRNAs are defined as a class of non-protein-coding transcripts greater than 200 nucleotides in length, which are involved in a large spectrum of biological processes, including proliferation, differentiation, apoptosis, invasion, and chromatin remodeling. However, little is known about the specific mechanisms of these lncRNAs. They can act as oncogenic and/or tumor-suppressive factors in the development and progression of MM. But that how do they work remains unclear. In this review, the recent progress in the study of functional lncRNAs associated with MM was summarized and the present knowledge about their expression and roles was discussed, to provide guidance for the in-depth functional study of lncRNAs.

Retrospective analysis of the efficacy and influencing factors of autologous hematopoietic stem cell transplantation for multiple myeloma.

This study aims to review the clinical efficacy and factors affecting the treatment of multiple myeloma (MM) by autologous hematopoietic stem cell transplantation (ASCT). The clinical data of 47 patients with MM from the Department of Hematology of Henan Cancer Hospital from September 2010 to July 2018 were retrospectively analyzed. At pre-transplantation of autologous cells, 25.5% were in complete remission (CR), 14.9% were in very good partial remission (VGPR) and 59.6% were in partial remission (PR). Among these cases, one case had PR after three recurrences. At post-transplantation, 51% were in CR, including two cases who received double transplantations, 27.7% were in VGPR, and 21.3% were in PR. The median follow-up time was 27.6 months (4-96 months). The 3-year progression free survival (PFS) and overall survival (OS) were 47.9% and 79.6%, respectively. The Analysis of variance (ANOVA) results revealed that factors that affected OS were international staging system (ISS) stage (P = 0.002), CR and VGPR post-transplantation (P = 0.002), while factors that affected PFS were ISS stage (P = 0.005), pre-transplant induction therapy (P = 0.032), and disease risk stratification (P = 0.017). The curative effects for PFS were CR and VGPR pre-transplantation (P = 0.013) and post-transplantation (P = 0.011). The Cox multivariate regression analysis revealed that ISS stage and CR and VGPR post-transplantation were independent prognostic factors of OS. At post-transplantation, CR and VGPR, ISS stage, and pre-transplant induction therapy were independent prognostic factors for PFS. In conclusion, ASCT can improve the clinical efficacy and survival rate of MM patients. ISS stage, CR and VGPR post-transplantation are independent prognostic factors of OS and PFS, while pre-transplant induction therapy is an independent prognostic factor for PFS.

[Efficacy and Influencing Factors of Autologous Hematopoietic Stem Cell Transplantation for Multiple Myeloma: Retrospective Analysis].

OBJECTIVE: To analyze the clinical efficacy and possible influencing factors of autologous hematopoietic Stem cell transplantation (auto-HSCT) in the treatment of patients with multiple myeloma (MM). METHODS: Clinical data of 40 MM patients received auto-HSCT in the Department of Hematology of Henan Cancer Hospital from September 2010 to November 2017 were retrospectively analyzed, the clinical curative efficiency was summarized and the related factors were analyzed. RESULTS: The curative efficiency of the patients before transplantation was 9(22.5%) with complete remission(CR), 5(12.5%) with very good partial remission(VGPR), 26(65%) with partial remission(PR), respectively, one of them was PR after 3 recurrences. The curative efficiency after transplantation was 22(55%) with complete remission(CR), 12(30%) with very good partial remission(VGPR), 6(15%) with partial remission(PR), respectively. And 2 cases were CR after double transplantation. Median follow-up time was 28.4 (3.1 to 88) months,15 cases presented disease progression, 7 cases were dead, 3-year estimated progression-free survival(PFS) and overall survival(OS) rate were 45.1% and 82% respectively. Unvariate analysis showed that the OS was affected by ISS stage (P<0.05), CR and VGPR (P<0.05) after transplantation; PFS was affected by ISS stage (P<0.01), before transplantation induction therapy (27 cases with bortezomizomi or thalidomide) (P<0.05), disease risk stratification (6 cases in high risk group) (P<0.05) , CR and VGPR (P<0.05) before transplantation, CR and VGPR (P<0.01) after transplantation. Cox multivariate regression analysis showed that the independent prognostic factors for OS were ISS stage, CR and VGPR after transplantation; the independent prognostic factors for PFS were the CR, VGPR, ISS stage after transplantation and induction therapy before transplant. CONCLUSION: Auto-HSCT can improve the clinical efficacy and survival rate of MM patients; ISS stage, CR and VGPR after transplantation are independent prognostic factors for OS and PFS, and induction therapy before transplantation is also an independent prognostic factor for PFS.

Clinical observation of recombinant human endostatin in treating relapsed refractory multiple myeloma.

Recombinant human endostatin (rhES) can inhibit multiple myeloma, while its clinical efficacy in treating relapsed refractory multiple myeloma (RRMM) has not been assessed. One hundred and eleven RRMM patients were treated with four different regimens: combination of VD (velcade+dexamethasone) and rhES (n = 25), Thalidomide (Tha) and VD (VTD, n = 22) combination, rhES and conventional chemotherapy combination (n = 32), and combination of conventional chemotherapy and Tha (n = 32). Significant differences were found in progression-free survival (PFS) between rhES combination groups and conventional chemotherapy combination groups. No statistical difference was found in overall response rate, overall survival or incidences of adverse effects. The combination of rhES with VD or conventional chemotherapy is active in patients with RRMM and prolongs the PFS to improve the quality of life.

[As2O3 Up-Regulates the Proportion of CD4+CD25+CD127low Tregs in Peripheral Blood of Patients with Severe Aplastic Anemia].

OBJECTIVE: To investigate the effect of ATO on the proportion of Treg in the peripheral blood of patients with severe aplastic anemia (SAA) in vitro. METHODS: The peripheral blood of 20 newlydiagnosed patients were collected, and the peripheral blood monomuclear cells (PBMNC) were extracted. After the PBMNC were treated with ATO of different concentrotions (0, 1, 2.5 and 5 µmol/L) for 96 hours, the proportion of CD44+ CD25+CD127low regulatatory T cells (Treg) were detected by flow cytometry. The expression levels of Foxp3 mRNA were detected by RT-PCR, and the levels of IFN-γ,IL-4,IL-17 and TGF-ß1 were detected by ELTSA to verify the results of flow cytomery. RESULTS: ATO significantly increased the proportion of Treg (P<0.01) at the concentration of 2.5 and 5 µmol/L, and the rising degree of Treg proportion improved with the increasing ATO concentration(r= 0.524). Treg proportion increased at a concentration of 1 µmol/L, but without statistical significance (P>0.05). At 1(P<0.05), 2.5(P<0.01) and 5 µmol/L(P<0.01), ATO significantly up-regulated the expression of Foxp3 mRNA, and the increase of Foxp3 mRNA positively and linearly correlated with the increase of Treg cell-frequency(r=0.523). ATO significantly reduced the levels of IFN-γ (at ATO 1,2.5 and 5 µmol/L, P<0.01), IL-4 (at ATO 2.5 µmol/L, P<0.01; at ATO 5 µmol/L, P<0.01) and IL-17(at ATO 2.5 µmol/L, P<0.05; at ATO 5 µmol/L, P<0.01). ATO had no significant effect on TGF-ß1 at 1(P>0.05) and 2.5 µmol/L (P>0.05), but significantly reduced TGF-ß1 level at 5 µmol/L (P<0.05). CONCLUSION: ATO can mediate the immune regulation through up-regulating the proportion of Treg in peripheral blood of patients with SAA and reducing the levels of IFN-γ, IL-4 and IL-17.

[Rituximab and Dexamethasone Combined with Cyclophosphamide for Treatment of Relapsed and Refractory Immune Thrombocytopenia].

OBJECTIVE: To evaluate the efficiency and safety of rituximab and dexamethasone combined with cyclophosphamide for treating patients with relapsed and refractory immune thrombocytopenia (ITP). METHODS: Twelve patients with relapsed and refractory immune thrombocytopenia were prospectively enrolled in this study, and received rituximab 375 mg/m(2) once a week for 4 weeks, dexamethasone 40 mg once a day for consecutive 4 days, and cyclophosphamide 500 mg/m(2) biweekly for 2 weeks. The levels of IFN-r and IL-4 in peripheral blood of patients were measured by enzyme-linked immunosorbent assay (ELISA), and the percentages of Breg, Treg and Th17 cells were detected by flow cytometry before and after treatment. Efficiency was evaluated according to platelet counts, and side effects were observed. RESULTS: Six out of 12 patients reached to complete remission and 4 patients reached to partial remission, with the total response rate 83.33%. The platelet counts [(115.42 ± 76.60) × 10(9)/L] after treatment were significantly higher than that before treatment [(115.42 ± 76.60) × 10(9)/L] (P < 0.001). The ratio of IFN-r/ IL4 after treatment (5.89 ± 2.30) was very significantly lower than that before treatment (7.00 ± 2.73) (P = 0.002). The percentage of Breg cells after treatment [(21.27 ± 4.28)%] were much significantly higher than that before treatment [(15.48 ± 1.67)%] (P < 0.001). The ratio of Treg/Th17 after treatment (3.07 ± 1.50) was significantly higher than that before treatment (0.98 ± 0.45) (P < 0.001). Infusion reaction was observed in 1 patient, secondary hypertension and hyperglycemia were in 1 patient, and pneumonia in 2 patients. CONCLUSION: Rituximab and dexamethasone combined with cyclophosphamide can improve the outcomes of patients with relapsed and refractory immune thrombocytopenia patients and they were well tolerated, its mechanism may be related with the balance between T cell sunsets and Treg cells.

[Regulatory effect of As2O3on imbalance between adipogenic and osteogenic differentiation of BM-MSC from patients with aplastic anemia].

This study was aimed to explore the regulation of arsenic trioxide (As2O3) on imbalance between adipogenic and osteogenic differentiation of BM-MSC from patients with aplastic anemia(AA). The BM-MSC from AA patients were separated and purified, placed into the adipogenic and osteogenic differentiation culture system, then added the As2O3, CsA, As2O3combined with CsA were added to corresponding differentiation culture system, the concentration of As2O3and CsA were set at 0.001 µmol/ml and 2.5 mmol/ml respectively, the cells were divided into As2O3group, the CsA group, combined group and control group (no drug). The cell morphological observation, oil red 'O' staining, Von-Kossa staining, and RT-PCR were used to detect corresponding differentiation marks. The results indicated that in respect to adipogenic differentiation, cellular morphology observing and oil red 'O' staining showed that the rate of adipocyte differentiation in As2O3group was (18.3 ± 1.9)%, which was lower than the (91.8 ± 2.7)% in the CsA group and (92.1 ± 1.2)% in control group (P < 0.05), there was no significant difference in comparison with (8.3 ± 1.9)% in the combined group (P > 0.05), but the rate of differentiation in CsA group was higher than that in combined group (P < 0.05), and there was no significant difference in comparison wtih control group. RT-PCR showed that the LPL-mRNA expression level in As2O3group were significantly lower than that in the CsA group and the control group (P < 0.05), no difference was observed while compared with the combined group (P > 0.05), but the LPL-mRNA expression level in CsA group was significantly higher than that in the combined group (P < 0.05). In terms of osteogenetic differentiation, the calcium deposition in As2O3group and combined group was obviously observed while rarely in the CsA group and the control group when detected by the Von-Kossa staining. OST-mRNA expression level in As2O3group were higher than that in CsA group and the control group (P < 0.05), while compared with the combined group, there was no significant difference (P > 0.05), but the OST-mRNA expression level in the CsA group was lower than that in the combined group (P < 0.05). It is concluded that As2O3can significantly enhance the ability of BM-MSC from AA patients to differentiate into osteoblast, also can inhibit the adipogenic differentiation, in contrast, the CsA can not promote the osteoblast differentiation of BM-MSC from AA patients.

[Hyperthermia increases the sensitivity of RPMI 8226 cells to adriamycin].

This study was purposed to explore the effect of hyperthermia on sensitivity of multiple myeloma cells RPMI 8226 to adriamycin (ADM) and its mechanism. The working concentration of ADM against RPMI 8226 cells was defined by MTT assay. RPMI 8226 cells were divided into 4 groups: control group, hyperthermia (42°C) group, chemotherapy (ADM) group and combination group (42°C + ADM), the survival rate of RPMI 8226 cells in 4 groups was detected by trypan blue exclusion, the inhibitory effect of hyperthermia on proliferation of RPMI 8226 cells was detected by MTT assay, the cell cycle distribution, apoptosis rate of cells, intracellular ADM concentration and P-gp expression level were measured by flow cytometry. The 1/4 IC(50) of ADM was defined as the working concentration in the experiment. The results indicated that the hyperthermia promoted the entering the cells from in G(0)/G(1) phase into S and G(2)/M phases, the expression of P-gp protein on cells in hyperthermia and combination groups was down-regulated, the intracellular ADM concentration in combination group obviously increased. It is concluded that the hyperthermia combined with ADM obviously enhance the inhibitory effect on proliferation of RPMI 8226 cells. The hyperthermia increases the sensitivity of RPMI 8226 cells to chemotherapy through down-regulating the expression of P-gp protein on cells and increasing the intracellular ADM concentration.