title: Down-regulation of Mcl-1 by small interference RNA induces apoptosis and sensitizes HL-60 leukemia cells to etoposide.
abstract: BACKGROUND
  Acute myeloid leukemia (AML) is a fatal hematological malignancy which is resistant to a variety of chemotherapy drugs. Myeloid cell leukemia-1 (Mcl-1), a death-inhibiting protein that regulates apoptosis, has been shown to be overexpressed in numerous malignancies. In addition, it has been demonstrated that the expression level of the Mcl-1 gene increases at the time of leukemic relapse following chemotherapy. The aim of this study was to target Mcl-1 by small interference RNA (siRNA) and analyze its effects on survival and chemosensitivity of acute myeloid leukemia cell line HL-60.
  
  
  METHODS
  siRNA transfection was performed with a liposome approach. The expression levels of mRNA and protein were measured by real-time quantitative PCR and Western blot analysis, respectively. Trypan blue assays were performed to evaluate tumor cell growth after siRNA transfection. The cytotoxic effects of Mcl-1 siRNA (siMcl-1) and etoposide were determined using MTT assay on their own and in combination. Apoptosis was quantified using a DNA-histone ELISA assay.
  
  
  RESULTS
  Transfection with siMcl-1 significantly suppressed the expression of Mcl-1 mRNA and protein in a time- dependent manner, resulting in strong growth inhibition and spontaneous apoptosis. Surprisingly, pretreatment with siMcl-1 synergistically enhanced the cytotoxic effect of etoposide. Furthermore, Mcl-1 down-regulation significantly increased apoptosis sensitivity to etoposide. No significant biological effects were observed with negative control siRNA treatment.
  
  
  CONCLUSIONS
  Our results suggest that specific suppression of Mcl-1 by siRNA can effectively induce apoptosis and overcome chemoresistance of leukemic cells. Therefore, siMcl-1 may be a potent adjuvant in leukemia chemotherapy.
authors.affiliation: Immunology Research Center, Department of Medical Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran E-mail : esakhinia@yahoo.co.uk, baradaranb@tbzmed.ac.ir.
authors.forename: Hadi
authors.initials: H
authors.lastname: Karami
authors.forename: Behzad
authors.initials: B
authors.lastname: Baradaran
authors.forename: Ali
authors.initials: A
authors.lastname: Esfehani
authors.forename: Masoud
authors.initials: M
authors.lastname: Sakhinia
authors.forename: Ebrahim
authors.initials: E
authors.lastname: Sakhinia
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects; D005047:Etoposide / Q000494:pharmacology*; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D007938:Leukemia / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology*; D064549:Myeloid Cell Leukemia Sequence 1 Protein / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
chemical_list: D000972:Antineoplastic Agents, Phytogenic; C579095:MCL1 protein, human; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D005047:Etoposide