Overexpression of EIF5A2 is associated with poor survival and aggressive tumor biology in gallbladder cancer.

Gallbladder cancer (GBC) is a malignant tumor of the biliary tract. The main problem affecting the treatment of gallbladder cancer is late diagnosis and poor prognosis. EIF5A2 is one of two isoforms of the EIF5A family and is reported to be a new oncogenic protein in many human cancers. In this study, our results showed for the first time that EIF5A2 was overexpressed in GBC samples compared with non-tumor tissue. Overexpression of EIF5A2 was associated with lymph node metastasis, tumor differentiation, UICC (Union for International Cancer Control) staging, histological type, metastasis, and tumor size. Overexpression of EIF5A2 in gallbladder carcinoma tissues is also associated with poor prognosis in patients. The interference of EIF5A2 significantly inhibited the proliferation, cell cycle, migration and colony formation of GBC-SD cells in vitro. Our results suggest that EIF5A2 is a target oncogene and may be an important prognostic biomarker in the pathogenesis of gallbladder cancer.

Surface expression of anti-CD3scfv stimulates locoregional immunotherapy against hepatocellular carcinoma depending on the E1A-engineered human umbilical cord mesenchymal stem cells.

Tumor antigens is at the core of cancer immunotherapy, however, the ideal antigen selection is difficult especially in poorly immunogenic tumors. In this study, we designed a strategy to modify hepatocellular carcinoma (HCC) cells by surface expressing anti-CD3scfv within the tumor site strictly, which depended on the E1A-engineered human umbilical cord mesenchymal stem cells (HUMSC.E1A) delivery system. Subsequently, membrane-bound anti-CD3scfv actived the lymphocytes which lysed HCC cells bypassing the expression of antigens or MHC restriction. First, we constructed the anti-CD3scfv gene driven by human α-fetoprotein (AFP) promoter into an adenoviral vector and the E1A gene into the lentiviral vector. Our results showed that anti-CD3scfv could specifically express on the surface of HCC cells and activate the lymphocytes to kill target cells effectively in vitro. HUMSC infected by AdCD3scfv followed by LentiR.E1A could support the adenoviral replication and packaging in vitro 36 h after LentiR.E1A infection. Using a subcutaneous HepG2 xenograft model, we confirmed that AdCD3scfv and LentiR.E1A co-transfected HUMSC could migrate selectively to the tumor site and produce considerable adenoviruses. The new generated AdCD3scfv infected and modified tumor cells successfully. Mice injected with the MSC.E1A.AdCD3scfv and lymphocytes significantly inhibited the tumor growth compared with control groups. Furthermore, 5-fluorouracil (5-FU) could sensitize adenovirus infection at low MOI resulting in improved lymphocytes cytotoxicity in vitro and in vivo. In summary, this study provides a promising strategy for solid tumor immunotherapy.

Gene therapy in B-NHL cell line using adenovirus-mediated transfer of secretable trimeric TRAIL gene expression driven by CD20 promoter.

CD20 is a crucial target to B-non-Hodgkin lymphoma (NHL), in fact, a humanized anti-CD20 monoclonal antibody, rituximab, is widely applied in clinical practice. However, resistance to rituximab often occurs in B-NHL patients, which has encouraged us to find new medications to treat B-NHL. In this study, we designed a gene therapy strategy targeting CD20 at a transcriptional level mediated by adenovirus, in which the stTRAIL gene was driven by a specific CD20 promoter fragment. We cloned the CD20 promoter from genome DNA of BJAB cell, a CD20-positive cell line, and identified its specific transcriptional activity with a dual-luciferase reporter assay system. Meanwhile, we constructed the stTRAIL gene sequence, which contained secretion signal, isoleucine zipper, and soluble TRAIL gene sequence, in which the isoleucine zipper facilitated the product of this gene sequence to form a functional homotrimer. The recombinant adenovirus was termed as AdP20-stTRAIL, which carried on the fused gene of the CD20 promoter fragment and the stTRAIL gene. Our studies confirmed that the stTRAIL could be expressed and secreted from BJAB cells infected with AdP20-stTRAIL specifically, and it inhibited the growth of these infected BJAB cells in vitro and in vivo. Our results indicate that the gene therapy using stTRAIL gene driven by a CD20 promoter may be an effective strategy in B-NHL treatment.

[The regulation of EKI-785 to the growth of U937 cell line].

OBJECTIVE: The aim of this study was to explore the expression of erbBs in U937, an acute monocyte leukemia cell line, and their impact on the growth of this cell line. METHODS: Expression of erbBs was detected by RT-PCR and expression of erbB2 at protein level by Western blot. After U937 cells was treated with EKI-785, an irreversible specific inhibitor of erbBs, the growth was assessed by MTT and growth curve, apoptosis was detected by Annexin V-FITC Apoptosis Detection Kit, and signal pathway was detected by Western blot. RESULTS: erbB2-4 were expressed in U937 cell line, but not erbB1. Especially, protein of erbB2 was expressed in this cell line. After treating with EKI-785, the growth of U937 cells was inhibited and early apoptosis was induced. Moreover, the Ras/MAPK and the PI3K/Akt signaling pathways were all blocked. CONCLUSION: erbBs may play key roles in the development of some leukemia. Therefore, erbBs may become new targets of treatment to leukemia, and EKI-785 has a potency of clinic use to leukemia.

An effective and simple expansion system for megakaryocyte progenitor cells using a combination of heparin with thrombopoietin and interleukin-11.

Transfusion of ex vivo expanded megakaryocyte (MK) progenitor cells has been suggested to shorten the time of platelet recovery in the thrombocytopenia induced by radiotherapy or chemotherapy. Here, we report an effective and simple expansion system of MK progenitor cells from cord blood (CB) CD34(+) cells using a combination of thrombopoietin (TPO), interleukin (IL)-11, and heparin. When the CB CD34(+) cells were cultured in a liquid expansion system in the presence of TPO + recombination human (rh)IL-11 + heparin for 7 days, the number of CB CD34(+)/CD41a(+) cells was significantly increased compared to control groups (p < 0.05). When the suspension cells collected from 7-day liquid culture were replated in semisolid cultures, increased large MK colonies were observed in the culture with combination of TPO + IL-11 + heparin compared to those of control groups. In vivo, transfusion of CD34(+) cells expanded with TPO + IL-11 + heparin into irradiated nonobese diabetic/severe combined immunodeficient mice significantly accelerated platelet recovery. These data indicate that heparin as effective cofactor for TPO and IL-11 promotes expansion of MK progenitor cells from CB CD34(+) cells. This expansion system is simple and effective and could be used for the treatment of thrombocytopenia after radiotherapy or chemotherapy.