Knockdown of DEPTOR inhibits cell proliferation and increases chemosensitivity to melphalan in human multiple myeloma RPMI-8226 cells via inhibiting PI3K/AKT activity

J Int Med Res. 2013 Jun;41(3):584-95. doi: 10.1177/0300060513480920. Epub 2013 Apr 15.

Abstract

Objective: The present study determined the role of DEP domain containing mTOR-interacting protein (DEPTOR) in the proliferation, apoptosis and chemosensitivity of RPMI-8226 multiple myeloma cells, using small hairpin RNA (shRNA) to knock down DEPTOR gene expression in vitro.

Methods: DEPTOR mRNA and protein levels in RPMI-8226 cells treated with DEPTOR-specific shRNA were evaluated by reverse transcription-polymerase chain reaction and Western blotting. Expression of apoptosis-associated proteins (including cleaved caspase-3 and cleaved poly-ADP ribose polymerase [PARP]) and activation of the phosphatidylinositol 3-kinase (PI3K)/v-akt murine thymoma viral oncogene homologue 1 (AKT) signalling pathway were detected by Western blotting.

Results: Transfection of DEPTOR-specific shRNA successfully knocked down DEPTOR gene expression in transfected RPMI-8226 cells. These transfected cells, together with control RPMI-8226 cells, were treated with 20 ┬Ámol/l melphalan for 24 h. Knockdown of DEPTOR exacerbated melphalan-induced growth inhibition and apoptosis, increased levels of cleaved caspase-3 and cleaved PARP, and reduced levels of phosphor-AKT.

Conclusion: Downregulation of DEPTOR inhibited proliferation and increased chemosensitivity to melphalan in human multiple myeloma RPMI-8226 cells via inhibiting the PI3K/AKT pathway.

Keywords: DEP domain containing mTOR-interacting protein (DEPTOR); Multiple myeloma; apoptosis; melphalan; phosphatidylinositol 3-kinase/v-akt murine thymoma viral oncogene homologue 1 (PI3K/AKT); proliferation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antineoplastic Agents / pharmacology*
  • Apoptosis
  • Caspase 3 / genetics
  • Caspase 3 / metabolism
  • Cell Line, Tumor
  • Cell Proliferation
  • Drug Resistance, Neoplasm / drug effects
  • Drug Resistance, Neoplasm / genetics*
  • Gene Expression Regulation, Neoplastic / drug effects*
  • Gene Knockdown Techniques
  • Humans
  • Intracellular Signaling Peptides and Proteins
  • Melphalan / pharmacology*
  • Multiple Myeloma / drug therapy
  • Multiple Myeloma / genetics*
  • Multiple Myeloma / metabolism
  • Multiple Myeloma / pathology
  • Phosphatidylinositol 3-Kinases / genetics*
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphoinositide-3 Kinase Inhibitors
  • Poly(ADP-ribose) Polymerases / genetics
  • Poly(ADP-ribose) Polymerases / metabolism
  • Proto-Oncogene Proteins c-akt / antagonists & inhibitors
  • Proto-Oncogene Proteins c-akt / genetics*
  • Proto-Oncogene Proteins c-akt / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Signal Transduction
  • TOR Serine-Threonine Kinases / antagonists & inhibitors
  • TOR Serine-Threonine Kinases / genetics*
  • TOR Serine-Threonine Kinases / metabolism

Substances

  • Antineoplastic Agents
  • Intracellular Signaling Peptides and Proteins
  • Phosphoinositide-3 Kinase Inhibitors
  • RNA, Small Interfering
  • Poly(ADP-ribose) Polymerases
  • DEPTOR protein, human
  • TOR Serine-Threonine Kinases
  • Proto-Oncogene Proteins c-akt
  • Caspase 3
  • Melphalan