Seyed H. Ghaffari

Phase II Study of Single-Agent Arsenic Trioxide for the Front-Line Therapy of Acute Promyelocytic Leukemia

PURPOSEnThe long-term follow-up results of patients with acute promyelocytic leukemia (APL) treated with all-trans retinoic acid and chemotherapy show high cure rates. Several studies have shown high efficacy of single-agent arsenic trioxide in newly diagnosed APL. However, long-term follow-up results are needed.nnnPATIENTS AND METHODSnOne hundred ninety-seven patients with newly diagnosed APL were treated with arsenic trioxide 0.15 mg/kg daily intravenous infusion until complete remission (CR). After achieving CR, the patients received one to four more courses of therapy with arsenic trioxide as consolidation and were observed with reverse-transcriptase polymerase chain reaction studies from peripheral blood (to detect of minimal residual disease) every 3 months or until relapse or death.nnnRESULTSnThe morphologic CR rate was 85.8%. The most common cause of remission failure was early death owing to APL differentiation syndrome (13.2%). The most important prognostic factor for early mortality was a high WBC count at presentation. The 5-year disease-free survival (DFS) rate was 66.7% ± 4% (SE). Relapse after 5 years in CR was rare. The 5-year overall survival (OS) rate by intention-to-treat analysis was 64.4% ± 4%. In patients who achieved CR, OS and DFS were identical.nnnCONCLUSIONnThe long-term follow-up of newly diagnosed patients with APL treated with single-agent arsenic trioxide shows high rates of DFS and OS.

Effects of silibinin on cell growth and invasive properties of a human hepatocellular carcinoma cell line, HepG-2, through inhibition of extracellular signal-regulated kinase 1/2 phosphorylation.

The purpose of the current study is to evaluate the effect of silibinin on human hepatocellular carcinoma HepG-2 cells. Microculture tetrazolium test (MTT assay), Lactate dehydrogenase (LDH) release, Gelatin zymography, Griess reaction, Cell-based the extracelluar signal-regulated kinase (ERK) 1/2 phosphorylation assay and quantitative real-time RT-PCR were employed to appraise the effect of silibinin on cell proliferation, cytotoxicity, metastatic potential, nitric oxide (NO) production, ERK 1/2 phosphorylation and activation in HepG-2 cells. Silibinin inhibited cell proliferation, matrix metalloproteinase 2 enzymatic activity, NO production and ERK 1/2 phosphorylation in a dose-dependent manner without exerting any cytotoxicity effect. In addition, an expressive increase in mRNA levels of Raf kinase inhibitor protein (RKIP), sprouty-related protein 1 with EVH-1 domain (Spred-1), sprouty-related protein with EVH-1 domain 2 (Spred-2) coupled with a significant reduction in transcriptional levels of highly expressed in cancer (Hec1) and MMP-2 were observed. Altogether, these issues show for the first time that silibinin treatment could inhibit cell proliferation and invasive potential of HepG-2 cells through inhibition of ERK 1/2 cascade both directly (through suppression of ERK 1/2 phosphorylation) and indirectly (through up-regulation of RKIP, Spred-1 and Spred-2). In addition, cell growth and proliferation may be inhibited by silibinin through down-regulation of Hec1.

Anti-apoptotic and anti-inflammatory effects of Silybum marianum in treatment of experimental steatohepatitis

In this study, we were aimed to evaluate the probable effect of the crud extract of Silybum marianum, with high polyphenolic content, on experimental nonalcoholic steatohepatitis (NASH). To induce NASH, a methionine and choline deficient (MCD) diet was given to N-Mary rats for 8 weeks. After NASH development, MCD-fed rats were divided into two groups: MCD groups received MCD diet and MCD+S group was fed MCD diet plus crude extract of S. marianum orally for 3 weeks. Control group was fed a normal diet for 11 weeks. Finally, all rats were sacrificed. Plasma alanine amino transferase (ALT) and aspartate amino transferase (AST) levels were evaluated. In addition, the following hepatic factors were also evaluated: liver histology, malondialdehyde (MDA) and reduced glutathione (GSH) contents, gene expressions of TNF-α and TGF-β and immunoblot evaluations of caspase-3, ERK/p-ERK, JNK/pJNK and p38/pp38. Histopathological evaluations of the liver samples revealed that treatment with the S. marianum extract has abated the severity of NASH among the MCD-fed rats. Also, a significant reduction was observed in the sera ALT and AST activities. In addition, the extract caused dramatic reduction in the elevated hepatic TNF-α and TGF-β mRNA and MDA levels along with an increase in the GSH content. Moreover, the plant extract treatments significantly lowered activation of procaspase-3 to active caspase-3 and also lowered the phosphorylated form of JNK among the same group of rats. These results suggest that the S. marianum crude extract beneficial effects on NASH are mainly due to its antioxidant and anti-inflammatory activities.

Silibinin inhibits invasive properties of human glioblastoma U87MG cells through suppression of cathepsin B and nuclear factor kappa B-mediated induction of matrix metalloproteinase 9

Glioblastoma multiforme remains one of the most devastating human malignancies because of its high infiltrative capacity. This study aimed to investigate the effects of silibinin on human glioblastoma U87MG cells. The microculture tetrazolium test, bromodeoxyuridine cell proliferation assay, cell-based nuclear factor kappa B (NF-κB) activation assessment, cathepsin B activity assay, gelatin zymography, and quantitative real-time reverse transcription-PCR were performed to appraise the effects of silibinin on the metabolic activity, DNA synthesis, NF-κB phosphorylation, cathepsin B activity, and gelatinolytic activity of U87 cells. Silibinin inhibited metabolic activity, cell proliferation, NF-κB activation, cathepsin B enzymatic levels, and gelatinase B activity in U87 cells. In addition, an expressive decrease in mRNA levels of matrix metalloproteinase-9, cathepsin B, urokinase plasminogen activator receptor, urokinase plasminogen activator, and intercellular adhesion molecule 1 coupled with a significant induction in transcriptional levels of stefin A was observed. Altogether, these issues show for the first time that silibinin treatment could trammel invasive features of a highly invasive human glioma cell line, U87, through suppression of NF-κB-mediated stimulation of matrix metalloproteinase-9. Furthermore, silibinin might cripple the activation of gelatinase B by cramping transcriptional and enzymatic activities of cathepsin B in U87 cells.

Monitoring of Cytomegalovirus Reactivation in Bone Marrow Transplant Recipients by Real-time PCR

Cytomegalovirus (CMV) has been recognized as the most important viral pathogen in persons undergoing bone marrow transplantation (BMT). The aim was to develop a quantitative PCR assay to quantify CMV DNA in peripheral blood leukocytes (PBLs) of bone marrow transplantation (BMT) patients. An in-house real-time PCR assay based on TaqMan technology was developed to monitor the quantity of CMV DNA in PBLs of the BMT recipients. Sequential blood samples (415 specimens) were collected from 43 patients as weekly intervals until day 100 after transplantation. The CMV DNA was quantified in parallel with the pp65 antigenemia assay in PBL samples. Viral reactivation occurred in 51% and 41.8% of the recipients as detected by RQ-PCR and antigenemia assays respectively. There was a significant correlation between both assays (Pu2009<u20090.0001); however, the RQ-PCR was more sensitive than the antigenemia. CMV DNA was detected by the RQ-PCR by a median of 14xa0days earlier than the antigenemia. Preemptive therapy was implemented in the antigenemia positive cases. The administration of ganciclovir led to a rapid decrease in the viral load. After preemptive therapy, the antigenemia achieved a negative result earlier than the RQ-PCR assay (a median of 17.5xa0days). An increase of viral load in both quantitative assays and of cyclosporine serum level were identified as the most significant risk factors for CMV reactivation. The quantitative CMV PCR might be a useful tool for monitoring the CMV reactivation and guiding the efficacy of the CMV preemptive therapy in BMT recipients.

Blockade of vascular endothelial growth factor receptors by tivozanib has potential anti-tumour effects on human glioblastoma cells

Glioblastoma (GBM) remains one of the most fatal human malignancies due to its high angiogenic and infiltrative capacities. Even with optimal therapy including surgery, radiotherapy and temozolomide, it is essentially incurable. GBM is among the most neovascularised neoplasms and its malignant progression associates with striking neovascularisation, evidenced by vasoproliferation and endothelial cell hyperplasia. Targeting the pro-angiogenic pathways is therefore a promising anti-glioma strategy. Here we show that tivozanib, a pan-inhibitor of vascular endothelial growth factor (VEGF) receptors, inhibited proliferation of GBM cells through a G2/M cell cycle arrest via inhibition of polo-like kinase 1 (PLK1) signalling pathway and down-modulation of Aurora kinases A and B, cyclin B1 and CDC25C. Moreover, tivozanib decreased adhesive potential of these cells through reduction of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Tivozanib diminished GBM cell invasion through impairing the proteolytic cascade of cathepsin B/urokinase-type plasminogen activator (uPA)/matrix metalloproteinase-2 (MMP-2). Combination of tivozanib with EGFR small molecule inhibitor gefitinib synergistically increased sensitivity to gefitinib. Altogether, these findings suggest that VEGFR blockade by tivozanib has potential anti-glioma effects in vitro. Further in vivo studies are warranted to explore the anti-tumour activity of tivozanib in combinatorial approaches in GBM.

BIBR 1532 Increases Arsenic Trioxide-mediated Apoptosis in Acute Promyelocytic Leukemia Cells: Therapeutic Potential for APL

The current treatment of acute promyelocytic leukemia with arsenic trioxide (ATO) has increased long-lasting complete remissions; however, a proportion of patients continues to die eventually as a result of disease recurrence. In an effort to enhance the effectiveness of the APL treatment, we designed experiments to evaluate the effects of ATO in combination with the lead compound of non-nucleoside inhibitor of telomerase, BIBR 1532. After combined treatments with BIBR 1532 and ATO, decreased cell viability index with a concomitant increase in apoptotic cell death was observed in NB4 leukemic cells. Apoptosis induced by the combined treatments was accompanied by elevated Bax/Bcl-2 molecular ratio and enhanced caspase 3 activation. Our study has also demonstrated that the combined treatment suppressed NB4 cell proliferative capacity and inhibited telomerase activity probably via transcriptional suppression of c-Myc and hTERT. In conclusion, this study may supply insight into the application of this new combination therapy to APL cells intrinsically less sensitive to routine therapies and suggested a novel combination therapy for patients with more aggressive disease; those who may not respond favorably to the arsenic mono-therapy.

Teucrium polium reversed the MCD diet-induced liver injury in rats:

In the present study, we evaluated the ability of Teucrium polium ethyl acetate fraction, with high antioxidant activity, in the treatment of nonalcoholic steatohepatitis (NASH) in rats and its possible effect on factors involved in pathogenesis of the disease. To induce NASH, a methionine and choline deficient (MCD) diet was given to N-Mary rats for 8 weeks. After NASH development, MCD-fed rats were divided into 2 groups: NASH group that received MCD diet and NASH + T group which was fed MCD diet plus ethyl acetate fraction of T. polium orally for 3 weeks. Histopathological evaluations revealed that treatment with the extract has abated the severity of NASH among the MCD-fed rats. In addition, the fraction reduced the elevated levels of hepatic tumor necrosis factor-alpha (TNF-α) and transforming growth factor-beta (TGF-β) gene expression and also the elevated level of malondialdehyde (MDA). In addition, the extract increased the activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and enhanced the level of hepatic glutathione (GSH). Moreover, the fraction treatments lowered caspase-3 level and the phosphorylated form of C-Jun N-terminal kinase (JNK) and augmented the phosphrylated level of extracellular regulated kinase1/2 (ERK1/2). These results indicate that the ethyl acetate fraction of T. poium effectively reversed NASH, mainly due to its strong antioxidant and anti-inflammatory properties.

Arsenic Trioxide Induces Apoptosis and Incapacitates Proliferation and Invasive Properties of U87MG Glioblastoma Cells through a Possible NF-κB-Mediated Mechanism

Identification of novel therapeutics in glioblastoma remains crucial due to the devastating and infiltrative capacity of this malignancy. The current study was aimed to appraise effect of arsenic trioxide (ATO) in U87MG cells. The results demonstrated that ATO induced apoptosis and impeded proliferation of U87MG cells in a dosedependent manner and also inhibited classical NF-κB signaling pathway. ATO further upregulated expression of Bax as an important proapoptotic target of NF-κB and also inhibited mRNA expression of survivin, c-Myc and hTERT and suppressed telomerase activity. Moreover, ATO significantly increased adhesion of U87MG cells and also diminished transcription of NF-κB down-stream targets involved in cell migration and invasion, including cathepsin B, uPA, MMP-2, MMP-9 and MMP-14 and suppressed proteolytic activity of cathepsin B, MMP-2 and MMP-9, demonstrating a possible mechanism of ATO effect on a well-known signaling in glioblastoma dissemination. Taken together, here we suggest that ATO inhibits survival and invasion of U87MG cells possibly through NF-κB-mediated inhibition of survivin and telomerase activity and NF-κB-dependent suppression of cathepsin B, MMP-2 and MMP-9.

Anti-tumour activity of tivozanib, a pan-inhibitor of VEGF receptors, in therapy-resistant ovarian carcinoma cells

Epithelial ovarian cancer (EOC) is the most fatal gynaecological malignancy. Despite initial therapeutic response, the majority of advanced-stage patients relapse and succumb to chemoresistant disease. Overcoming drug resistance is the key to successful treatment of EOC. Members of vascular endothelial growth factor (VEGF) family are overexpressed in EOC and play key roles in its malignant progression though their contribution in development of the chemoresistant disease remains elusive. Here we show that expression of the VEGF family is higher in therapy-resistant EOC cells compared to sensitive ones. Overexpression of VEGFR2 correlated with resistance to cisplatin and combination with VEGFR2-inhibitor apatinib synergistically increased cisplatin sensitivity. Tivozanib, a pan-inhibitor of VEGF receptors, reduced proliferation of the chemoresistant EOC cells through induction of G2/M cell cycle arrest and apoptotic cell death. Tivozanib decreased invasive potential of these cells, concomitant with reduction of intercellular adhesion molecule-1 (ICAM-1) and diminishing the enzymatic activity of urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP-2). Moreover, tivozanib synergistically enhanced anti-tumour effects of EGFR-directed therapies including erlotinib. These findings suggest that the VEGF pathway has potential as a therapeutic target in therapy-resistant EOC and VEGFR blockade by tivozanib may yield stronger anti-tumour efficacy and circumvent resistance to EGFR-directed therapies.