title: Silencing of p130cas in ovarian carcinoma: a novel mechanism for tumor cell death.
abstract: BACKGROUND
  We investigated the clinical and biological significance of p130cas, an important cell signaling molecule, in ovarian carcinoma.
  
  
  METHODS
  Expression of p130cas in ovarian tumors, as assessed by immunohistochemistry, was associated with tumor characteristics and patient survival. The effects of p130cas gene silencing with small interfering RNAs incorporated into neutral nanoliposomes (siRNA-DOPC), alone and in combination with docetaxel, on in vivo tumor growth and on tumor cell proliferation (proliferating cell nuclear antigen) and apoptosis (terminal deoxynucleotidyl transferase dUTP nick-end labeling) were examined in mice bearing orthotopic taxane-sensitive (HeyA8 and SKOV3ip1) or taxane-resistant (HeyA8-MDR) ovarian tumors (n = 10 per group). To determine the specific mechanisms by which p130cas gene silencing abrogates tumor growth, we measured cell viability (MTT assay), apoptosis (fluorescence-activated cell sorting), autophagy (immunoblotting, fluorescence, and transmission electron microscopy), and cell signaling (immunoblotting) in vitro. All statistical tests were two-sided.
  
  
  RESULTS
  Of 91 ovarian cancer specimens, 70 (76%) had high p130cas expression; and 21 (24%) had low p130cas expression. High p130cas expression was associated with advanced tumor stage (P < .001) and higher residual disease (>1 cm) following primary cytoreduction surgery (P = .007) and inversely associated with overall survival and progression-free survival (median overall survival: high p130cas expression vs low expression, 2.14 vs 9.1 years, difference = 6.96 years, 95% confidence interval = 1.69 to 9.48 years, P < .001; median progression-free survival: high p130cas expression vs low expression, 1.04 vs 2.13 years, difference = 1.09 years, 95% confidence interval = 0.47 to 2.60 years, P = .01). In mice bearing orthotopically implanted HeyA8 or SKOV3ip1 ovarian tumors, treatment with p130cas siRNA-DOPC in combination with docetaxel chemotherapy resulted in the greatest reduction in tumor growth compared with control siRNA therapy (92%-95% reduction in tumor growth; P < .001 for all). Compared with control siRNA therapy, p130cas siRNA-DOPC reduced SKOV3ip1 cell proliferation (31% reduction, P < .001) and increased apoptosis (143% increase, P < .001) in vivo. Increased tumor cell apoptosis may have persisted despite pan-caspase inhibition by the induction of autophagy and related signaling pathways.
  
  
  CONCLUSIONS
  Increased p130cas expression is associated with poor clinical outcome in human ovarian carcinoma, and p130cas gene silencing decreases tumor growth through stimulation of apoptotic and autophagic cell death.
authors.affiliation: Department of Gynecologic Oncology, University of Texas M. D. Anderson Cancer Center, Houston, TX 77230-1439, USA.
authors.forename: Alpa M
authors.initials: AM
authors.lastname: Nick
authors.forename: Rebecca L
authors.initials: RL
authors.lastname: Stone
authors.forename: Guillermo
authors.initials: G
authors.lastname: Armaiz-Pena
authors.forename: Bulent
authors.initials: B
authors.lastname: Ozpolat
authors.forename: Ibrahim
authors.initials: I
authors.lastname: Tekedereli
authors.forename: Whitney S
authors.initials: WS
authors.lastname: Graybill
authors.forename: Charles N
authors.initials: CN
authors.lastname: Landen
authors.forename: Gabriel
authors.initials: G
authors.lastname: Villares
authors.forename: Pablo
authors.initials: P
authors.lastname: Vivas-Mejia
authors.forename: Justin
authors.initials: J
authors.lastname: Bottsford-Miller
authors.forename: Hye Sun
authors.initials: HS
authors.lastname: Kim
authors.forename: Ju-Seog
authors.initials: JS
authors.lastname: Lee
authors.forename: Soo Mi
authors.initials: SM
authors.lastname: Kim
authors.forename: Keith A
authors.initials: KA
authors.lastname: Baggerly
authors.forename: Prahlad T
authors.initials: PT
authors.lastname: Ram
authors.forename: Michael T
authors.initials: MT
authors.lastname: Deavers
authors.forename: Robert L
authors.initials: RL
authors.lastname: Coleman
authors.forename: Gabriel
authors.initials: G
authors.lastname: Lopez-Berestein
authors.forename: Anil K
authors.initials: AK
authors.lastname: Sood
mesh_terms: D000328:Adult; D000368:Aged; D000818:Animals; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins / Q000032:analysis; D001343:Autophagy / Q000235:genetics*; D000071186:Beclin-1; D002277:Carcinoma / Q000737:chemistry / Q000235:genetics*; D002470:Cell Survival; D050719:Crk-Associated Substrate Protein / Q000235:genetics*; D018572:Disease-Free Survival; D000077143:Docetaxel; D015536:Down-Regulation; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D049452:Green Fluorescent Proteins; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D007150:Immunohistochemistry; D053208:Kaplan-Meier Estimate; D008565:Membrane Proteins / Q000032:analysis; D051379:Mice; D046529:Microscopy, Electron, Transmission; D008869:Microtubule-Associated Proteins / Q000032:analysis; D008875:Middle Aged; D015999:Multivariate Analysis; D010051:Ovarian Neoplasms / Q000737:chemistry / Q000235:genetics*; D010713:Phosphatidylcholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000032:analysis; D051057:Proto-Oncogene Proteins c-akt / Q000032:analysis; D034741:RNA, Small Interfering / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D043823:Taxoids / Q000494:pharmacology; D014162:Transfection; D014183:Transplantation, Heterologous; D015854:Up-Regulation
chemical_list: D051017:Apoptosis Regulatory Proteins; D000071186:Beclin-1; D050719:Crk-Associated Substrate Protein; D000077143:Docetaxel; D049452:Green Fluorescent Proteins; D008565:Membrane Proteins; D008869:Microtubule-Associated Proteins; D010713:Phosphatidylcholines; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D034741:RNA, Small Interfering; D043823:Taxoids; C493841:BCAR1 protein, human; C491997:BECN1 protein, human; C477012:MAP1LC3A protein, human; C017251:1,2-oleoylphosphatidylcholine