Caov3

Human ovarian cancer cell lines were purchased from the American Type Culture Collection (ATCC; Manassas, VA) and cultured in Dulbecco's modified Eagle's medium (SKOV3, CaOV3) or RPMI-1640 (OVCAR8, HeyA8) supplemented with 5 % fetal bovine serum (FBS; Wisent). Early-passage cell lines (designated “iOvCa”) are derived from ascites cultures (designated “EOC”) of the corresponding number (e.g., iOvCa201 is a line derived from EOC201). Early-passage lines were maintained in DMEM/F12 medium (Gibco/Invitrogen, Carlsbad, CA) supplemented with 10 % FBS. The iOvCa147-E2 line is a clone of iOvCa147. To establish stable expression of eGFP-LC3B, sub-confluent (~60-70 %) OVCAR8 cells were transfected with the pBMN-ires-puro-eGFP-LC3B construct (gift of Dr. C. McCormick, Dalhousie University) and transferred to Puromycin-containing selection medium (1 μg/mL) where clones with robust eGFP-LC3B expression were isolated. All cells were maintained in a 37 °C humidified atmosphere of 95 % air and 5 % CO₂. Adherent cells were maintained on tissue culture-treated polystyrene (Sarstedt, Newton, NC) and non-adherent cells were maintained on Ultra-low Attachment (ULA) cultureware (Corning, Corning, NY).

The human OC cell lines CaOV3 and OVCAR3 were obtained from the American Type Culture Collection (Manassas, VA) and maintained in a humidified 5 % CO₂ incubator at 37 °C. Cells were passaged twice weekly. OVCAR3 cells were maintained in RPMI-1640 (Wisent, St-Bruno, QC, Canada) supplemented with 20 % FBS, insulin (10 mg/L), glutamine (2 mM) and antibiotics. CaOV3 cells were cultured in DMEM/F12 (Wisent) supplemented with 10 % FBS, 2 mM glutamine and antibiotics. Acellular ascites fractions were obtained at the time of initial cytoreductive surgery from women with advanced serous ovarian carcinomas. Samples were supplied by the Banque de tissus et de données cliniques et biologiques sur les cancers gynécologiques et du sein de Sherbrooke as part of the Banque de tissus et de données du Réseau de Recherche en Cancer des Fonds de Recherche du Québec en Santé (FRQS) affiliated to the Canadian Tumor Repository Network (CTRNet). HRP-conjugated anti-mouse and rabbit antibodies and anti-FAK antibody were purchased from Cell Signaling Technology (Danvers, MA). Anti-phospho FAK and anti-phospho Pyl2 were from Thermo Fisher (Waltham, MA). Anti-Pyk2 and anti-Tubulin were purchased from Sigma-Aldrich (Oakville, ON). CCL18 and CCL18 neutralizing antibody were from RnD Systems (Minneapolis, MN). Plasmid pCMV6-ENTRY-PTK2B was obtained from Origene (Rockville, MD).