Jian-Pei Fang

FAK silencing inhibits leukemogenesis in BCR/ABL-transformed hematopoietic cells†

Focal adhesion kinase (FAK) is constitutively activated and tyrosine phosphorylated in BCR/ABL‐transformed hematopoietic cells, but the role it plays during leukemogenesis remains unclear. Here, we examined the effects of RNA interference‐mediated FAK silencing on leukemogenesis induced by a BCR/ABL‐transformed cell line. Transduction of BCR/ABL‐BaF3 cells with FAK shRNA inhibited FAK expression and reduced STAT5 phosphorylation, but induced caspase‐3 activation. In vitro studies showed that treatment with FAK shRNA resulted in impaired cell proliferation and colony formation, while increasing cell apoptosis. Mice that received transplants of BCR/ABL‐BaF3 cells with FAK shRNA displayed significantly prolonged survival time and diminished leukemia progression. In addition, FAK silencing enhanced in vitro and in vivo efficacy of ABL tyrosine kinase inhibitor imatinib in BCR/ABL‐BaF3 cells. Our results suggest that FAK is critical for leukemogenesis and might be a potential target for leukemia therapy. Am. J. Hematol. 2009.

Autoimmune Hemolytic Anemia in Patients with β-Thalassemia Major

Hemolysis is a common feature in patients with β-thalassemia major. As a result, autoimmune hemolytic anemia complicating β-thalassemia is easily overlooked. Here, the authors described the clinical features and management of 7 patients with β-thalassemia major and autoimmune hemolytic anemia. These patients had fever, cough, and tea-colored urine on admission. The laboratory investigations showed a significant drop in hemoglobin and increased serum bilirubin. Coombs’ tests revealed that anti-immunoglobulin G (IgG) and anti-C3 was positive in 7 and 5 cases, respectively, whereas anti-Rh E alloantibody was positive in 3 cases. All the patients received corticosteroids treatments and blood transfusions. Patients with anti-Rh E alloantibodies also received immunoglobulin treatments. Six of the patients responded well to the management, but 1 patient developed recurrent autoimmune hemolytic anemia that required cyclosporin A treatment. All the patients remained well by following up for more than 6 months.

HMGB1 translocation is involved in the transformation of autophagy complexes and promotes chemoresistance in leukaemia.

Acute lymphoblastic leukaemia (ALL) is a common paediatric cancer and is among the most curable cancers. However, the acquisition of drug resistance is a significant obstacle to the achievement of favourable outcomes, and autophagy is regarded as a mechanism that underlies chemoresistance. In this study, RT-qPCR was used to measure the expression of HMGB1 and Beclin1 in bone marrow mononuclear cells. A CCK-8 test was conducted to assess cell viability. Western blot, immunofluorescence and transmission electron microscopic analyses were performed to evaluate the autophagy levels. Immunoprecipitation analysis was performed to detect protein-protein interactions in the autophagy complexes. We found that HMGB1 expression correlated with the clinical status of ALL. In vitro, anticancer agent-induced cytotoxic effects were associated with autophagy-related drug resistance, and these effects were ameliorated by FIP200 depletion or the application of autophagy inhibitors. Moreover, the Ulk1‑Atg13-FIP200 complex, which promotes HMGB1 trafficking, acted upstream of the HMGB1-Beclin1 and PI3KC3-Beclin1 complexes and played a critical role in autophagy. Targeting the transformation of autophagic complexes or HMGB1 translocation may suppress autophagy and consequently overcome chemoresistance in leukaemia.

Pure red cell aplasia associated with cytomegalovirus and Epstein–Barr virus infection in seven cases of Chinese children

Abstract Pure red cell aplasia (PRCA) associated with cytomegalovirus (CMV) and Epstein–Barr virus (EBV) infection is uncommon. Here, we describe the clinical features and management of seven cases of Chinese children with PRCA associated with viral infections. The patients presented with pallor on admission. Blood cell counts and marrow smears showed anemia, reticulocytopenia, and aplasia of erythroblasts. Serological investigation and DNA polymerase chain reactions for CMV were positive in four patients and those tests for EBV were positive in other three patients. All patients received blood transfusion, corticosteroids treatment, and ganciclovir injection. Two patients had a complete response and one had a partial response after the treatments. The other three patients had a complete response to second-line therapies, including high-dose methylprednisolone, cyclosporin A, and intravenous immunoglobulin. Only one patient had no response to the therapies. Our results indicated that it might be important to combine immunosuppressive drugs with an antiviral drug in the management of PRCA associated with CMV and EBV infection.

Mitophagy is increased during erythroid differentiation in β-thalassemia.

Mitophagy is a selective degradation of mitochondria, which also plays a critical role in hematopoiesis. However, it is unclear what role, if any, this process plays in the pathogenesis of β-thalassemia. To determine the role of mitophagy in β-thalassemia, CD34+ hematopoietic progenitor cells (HPCs) were isolated from peripheral blood of β-thalassemia patients and healthy controls and differentiated into erythrocytes. We found that the ratio of mitochondrial membrane depolarization was significantly increased, and that mitochondria co-localize with lysosomes at a higher level in β-thalassemia compared with control. Furthermore, the expression of LC3-II and Nix, as well as degradation of p62, in β-thalassemia was higher than in the control. In sum, our data suggest that selective mitophagy is enhanced during erythrocyte differentiation in β-thalassemia.

Synergism between the mTOR inhibitor rapamycin and FAK down-regulation in the treatment of acute lymphoblastic leukemia

BackgroundAcute lymphoblastic leukemia (ALL) is an aggressive malignant disorder of lymphoid progenitor cells in both children and adults. Although improvements in contemporary therapy and development of new treatment strategies have led to dramatic increases in the cure rate in children with ALL, the relapse rate remains high and the prognosis of relapsed childhood ALL is poor. Molecularly targeted therapies have emerged as the leading treatments in cancer therapy. Multi-cytotoxic drug regimens have achieved success, yet many studies addressing targeted therapies have focused on only one single agent. In this study, we attempted to investigate whether the effect of the mammalian target of rapamycin (mTOR) inhibitor rapamycin is synergistic with the effect of focal adhesion kinase (FAK) down-regulation in the treatment of ALL.MethodsThe effect of rapamycin combined with FAK down-regulation on cell proliferation, the cell cycle, and apoptosis was investigated in the human precursor B acute lymphoblastic leukemia cells REH and on survival time and leukemia progression in a non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mouse model.ResultsWhen combined with FAK down-regulation, rapamycin-induced suppression of cell proliferation, G0/G1 cell cycle arrest, and apoptosis were significantly enhanced. In addition, REH cell-injected NOD/SCID mice treated with rapamycin and a short-hairpin RNA (shRNA) to down-regulate FAK had significantly longer survival times and slower leukemia progression compared with mice injected with REH-empty vector cells and treated with rapamycin. Moreover, the B-cell CLL/lymphoma-2 (BCL-2) gene family was shown to be involved in the enhancement, by combined treatment, of REH cell apoptosis.ConclusionsFAK down-regulation enhanced the in vitro and in vivo inhibitory effects of rapamycin on REH cell growth, indicating that the simultaneous targeting of mTOR- and FAK-related pathways might offer a novel and powerful strategy for treating ALL.

Homing and engraftment of bone marrow cells derived from different donors in a murine model of sensitization

Abstract Sensitized recipients are at a high risk of graft rejection in hematopoietic stem cell transplantation. To explore the trace of donor cells, we tried to explore homing and engraftment of bone marrow cells (BMCs) derived from different donors in a murine model of sensitization. Sensitized BALB/c mice were used as transplanted recipients, which received BMCs derived from C57BL/6 or BALB/c donors after lethal irradiation. The homing study showed that the donor cells decreased along with time in recipients of the C57BL/6 donor group, but the donor cells increased along with time in recipients of the BALB/c donor group. For the engraftment assay, all the sensitized recipients transplanted with BMCs derived from C57BL/6 donors died after lethal irradiation. In contrast, all the recipients transplanted with BMCs derived from BALB/c donors got long-term survival. Our results suggest that it is crucial to have human leukocyte antigen identical donors for sensitized recipients during hematopoietic stem cell transplantation.

Evaluation of hepatic iron overload in Chinese children with β-thalassemia major.

Patients with β-thalassemia major require long-term blood transfusions, resulting in hepatic iron overload. Thirty-five Chinese children with β-thalassemia major were recruited in the present studies. Hepatic iron overload was evaluated by histological grading. The relationships between hepatic iron overload and both serum biochemical markers and magnetic resonance imaging (MRI) examination were studied. The majority of the patients showed high degrees of hepatic iron overload by histological study. The degree of hepatic iron overload was correlated with serum ferritin (r = .70, P < .01), hyaluronic acid (r = .58, P = .011), and type III precollagen (r = .55, P = .035). Moreover, hepatic iron overload showed a negative correlation with liver to muscle signal intensity ratio (r = −.44, P = .012), and a positive correlation with red marrow area percentage (r = .52, P < .01). These results indicated that hepatic iron overload might be assessed by serum biochemical markers and MRI examination.

Modified conditioning regimen improves outcomes of unrelated donor peripheral blood stem cell transplantation for β-thalassaemia major patients

The objective of this study was to evaluate the feasibility of a modified conditioning regimen for the treatment of patients with β‐thalassaemia major (TM), using unrelated donor peripheral blood stem cell transplantation (UD‐PBSCT).

Arsenic trioxide induces cell cycle arrest and affects Trk receptor expression in human neuroblastoma SK-N-SH cells

AbstractBackgroundArsenic trioxide (As2O3), a drug that has been used in China for approximately two thousand years, induces cell death in a variety of cancer cell types, including neuroblastoma (NB). The tyrosine kinase receptor (Trk) family comprises three members, namely TrkA, TrkB and TrkC. Various studies have confirmed that TrkA and TrkC expression is associated with a good prognosis in NB, while TrkB overexpression can lead to tumor cell growth and invasive metastasis. Previous studies have shown that As2O3 can inhibit the growth and proliferation of a human NB cell line and can also affect the N-Myc mRNA expression. It remains unclear whether As2O3 regulates Trks for the purposes of treating NB.MethodsThe aim of the present study was to investigate the effect of As2O3 on Trk expression in NB cell lines and its potential therapeutic efficacy. SK-N-SH cells were grown with increasing doses of As2O3 at different time points. We cultured SK-N-SH cells, which were treated with increasing doses of As2O3 at different time points. Trk expression in the NB samples was quantified by immunohistochemistry, and the cell cycle was analyzed by flow cytometry. TrkA, TrkB and TrkC mRNA expression was evaluated by real-time PCR analysis.ResultsImmunohistochemical and real-time PCR analyses indicated that TrkA and TrkC were over-expressed in NB, and specifically during stages 1, 2 and 4S of the disease progression. TrkB expression was increased in stage 3 and 4 NB. As2O3 significantly arrested SK-N-SH cells in the G2/M phase. In addition, TrkA, TrkB and TrkC expression levels were significantly upregulated by higher concentrations of As2O3 treatment, notably in the 48-h treatment period. Our findings suggested that to achieve the maximum effect and appropriate regulation of Trk expression in NB stages 1, 2 and 4S, As2O3 treatment should be at relatively higher concentrations for longer delivery times;however, for NB stages 3 and 4, an appropriate concentration and infusion time for As2O3 must be carefully determined.ConclusionThe present findings suggested that As2O3 induced Trk expression in SK-N-SH cells to varying degrees and may be a promising adjuvant to current treatments for NB due to its apoptotic effects.