Guanghua Chen

Combination of DNA methylation inhibitor 5‐azacytidine and arsenic trioxide has synergistic activity in myeloma

XAF1 is a newly identified candidate tumour‐suppressor gene that can antagonise XIAP and sensitise cells to cell death triggers. This study was undertaken to study the effect of 5‐azacytidine (AZA) on XAF1 expression in myeloma cells and efficacy of 5‐AZA and arsenic trioxide (ATO) combination treatment in myeloma in vivo and in vitro. XAF1 expression was analysed by semi‐quantitative PCR and western blotting. Methylation specific PCR was used to detect methylation status of XAF1 promoter CpG islands. RPMI 8226 and XG‐7 cells were treated with various concentrations of 5‐AZA and ATO. Expression of XAF1 mRNA variants were confirmed by gel electrophoresis and sequencing. Untreated RPMI 8226 cell expresses two variants of XAF1 mRNA. Untreated XG‐7 cell has no expression of XAF1. Hypermethylation of XAF1 promoter CpG islands was detected in both cell lines. Both cell lines express full‐length XAF1 transcript after treated with 2.5 μmol/L 5‐AZA for 72 h. Our studies demonstrated that 5‐AZA exhibits anti‐myeloma synergy with ATO. In addition, ATO alone, 5‐AZA alone, or combination of 5‐AZA and ATO was effective in slowing myeloma growth and prolonging survival of myeloma‐loaded nude mice. The findings suggested that 5‐AZA and ATO may be an effective combination in the therapy of myeloma patients.

Expression and function of toll‐like receptors in multiple myeloma patients: toll‐like receptor ligands promote multiple myeloma cell growth and survival via activation of nuclear factor‐κB

Toll like receptors (TLRs) are the major agents for innate immunity that recognize invading microbial products and regulate the growth of normal and malignant human B lymphocytes. Multiple myeloma (MM) is a clonal plasma cell malignancy, though the regulatory role of TLRs in MM plasma cells has been reported, the molecular mechanism remains unclear. We first compared the transcripts of TLR1 to TLR10 in MM patients and healthy donors and found that TLR2, ‐4 and ‐9 transcripts were higher in bone marrow mononuclear cells (BMMCs) from patients than those from donors; in addition the expression of TLR4 and TLR9 were higher in MM cells than normal cells as demonstrated by flow cytometric analyses. The ligands of these two TLRs were capable to promote the growth of MM cells and protect them from serum‐deprivation‐induced apoptosis but not normal plasma cells, which could be attenuated with anti‐IL6 neutralizing antibodies or blockage of NF‐κB activities. Further investigation demonstrated that these TLR ligands could trigger the nuclear translocation of NF‐κB p65 and the activated NF‐κB was sufficient to increase the expression of IL6 transcript in MM cells. These data suggested that activated NF‐κB signalling probably plays a crucial role for the ligands of TLR4 and TLR9 to promote the growth and survival of MM cells partially through IL6 autocrine.

The Expression of Th17-Associated Cytokines in Human Acute Graft-versus-Host Disease

The role of Th17 cells and Th17-associated cytokines in the development of acute graft-versus-host disease (aGVHD) in clinical allogeneic hematopoietic stem cell transplantation (allo-HSCT) recipients is not well established. In the current study, a cohort of 69 allo-HSCT patients was examined for the percentages of Th17 and FoxP3(+) Treg cells and the expressions of RORγt and FoxP3 in peripheral blood mononuclear cells (PBMCs). The Th17 percentage and RORγt expression were significantly higher, whereas Treg percentage and FoxP3 expression were significantly lower in severe aGVHD (grade 3 to 4) and mild aGVHD (grade 1 to 2) patients than in patients without aGVHD (grade 0) and healthy donors. We then investigated the expressions of Th17-associated cytokines, including TGF-β, IL-6, IL-1β, IL-17, IL-21, IL-22, IL-23, as well as IL-23R in the PBMCs of patients after allo-HSCT. The expressions of IL-17 and IL-22 in CD4(+) T cells were also examined. The results showed that the expressions of IL-6, IL-1β, IL-17, IL-21, IL-23, and IL-23R were all increased, whereas IL-22 expression was decreased in aGVHD patients. The changes were also correlated with the severity of aGVHD. We also investigated the dynamic changes of Th17/Treg cells and Th17-associated cytokines in patients during the onset and resolution of aGVHD. The results demonstrated a reciprocal relationship between Treg and Th17 cells. Th17-associated cytokine expressions, namely IL-17 and IL-23, were closely related to the occurrence and resolution of aGVHD. We conclude that the dynamic balance between the Th17 and FoxP3(+) Treg cells and the changes of Th17-associated cytokines could be the indicators of the disease progression and promising candidates of prognostic biomarkers of aGVHD.

Multiple myeloma cells undergo differentiation upon exposure to rosiglitazone and all-trans retinoic acid

Activation of PPARγ by its ligands has shown differentiating effects in solid tumors. However, few reports addressed its role in myeloma cells. Our study demonstrated that exposure to PPARγ ligand (rosiglitazone, RGZ) induced proliferation inhibition and cell cycle arrest in myeloma cells. A combination of RGZ with all-trans retinoic acid (ATRA) can enhance the growth inhibition effects of RGZ. Further study shows that RGZ-treated myeloma cells displayed morphological characteristics of cell differentiation, and more evident signs of differentiation were observed when RGZ was combined with ATRA. These changes were confirmed by the detection of CD49e expression and light chain protein secretion. Similar results were also observed when primary CD138+ cells were treated with RGZ and ATRA. Collectively, our study revealed that RGZ can induce cell differentiation in myeloma cells and concomitant treatment with ATRA can enhanced the effects of RGZ.

Haploidentical allogeneic hematopoietic stem cell transplantation compared to matched unrelated transplantation for Philadelphia chromosome-positive acute lymphoblastic leukemia

To investigate the effect of haploidentical allogeneic hematopoietic stem cell transplantation (Haplo-HCT) in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL), the outcome of 58 patients with Ph+ ALL who received Haplo-HCT (n=42) or matched unrelated donor transplantation (MUD-HCT) (n=16) during the same period were analyzed retrospectively. All patients received a tyrosine kinase inhibitor (TKI)-based regimen before transplantation, and TKI was resumed primarily after transplantation. At the 3-year follow-up, the overall survival (OS), leukemia-free survival (LFS), the cumulative incidence of relapse (CIR), and non-relapse mortality (NRM) rates in Haplo-HCT group were 69.1, 64.3, 19.0, and 14.3%, respectively, without significant differences from that of MUD-HCT. Haplo-HCT was not related to higher incidences of severe acute graft-versus-host disease (GvHD) (17.6±5.2% vs. 20.0±10.0%, P=0.603) or chronic GvHD (19.5±7.1% vs. 13.3±8.6%, P=0.637) as compared to MUD-HCT. Multivariate analysis showed that chronic GvHD was associated with lower relapse rate in Haplo-HCT group. Haplo-HCT is a promising choice for improving the long-term survival in Ph+ ALL patients.