Journal: Cell reports

Article Title: Targeting the GPI transamidase subunit GPAA1 abrogates the CD24 immune checkpoint in ovarian cancer.

doi: 10.1016/j.celrep.2024.114041

Figure Lengend Snippet: Figure 5. Bestatin treatment increases phagocytosis and inhibits the growth of human ovarian cancer xenografts in mice (A) In vitro phagocytosis assay. Representative images (left) and quantification (right) of macrophage-mediated phagocytosis of OVCAR8 cells treated with bestatin (20 mM) or, as a control, DMSO. The results were normalized to that obtained with DMSO. Scale bars, 100 mm. Data are represented as mean ± SD (n = 3 independent experiments). (B–D) Quantification of macrophage-mediated phagocytosis of OVCAR3 (B), SKOV3 (C), and A2780 (D) cells treated with bestatin (20 mM) or DMSO. The results were normalized to that obtained with DMSO. Data are represented as mean ± SD (n = 3 independent experiments). (E and F) Left: representative in vivo bioluminescence images of NSG mice 5 weeks following implantation with OVCAR8 (E) or SKOV3 (F) cells and treated with bestatin or vehicle. Right: tumor growth, as measured by total flux. Data are represented as mean ± SEM (n = 5 mice per group). (G) Left: representative image depicting peritoneal ascites accumulation in mice 5 weeks following implantation with OVCAR8 cells and treatment with either vehicle or bestatin. Right: quantification of ascites volume from vehicle or bestatin-treated mice. Data are represented as mean ± SEM (n = 5 mice per group). (H) Quantification of ascites volume in mice 5 weeks after implantation with SKOV3 cells and treatment with vehicle or bestatin. Data are represented as mean ± SEM (n = 5 mice per group).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER PMSF Millipore Sigma Cat# 329-98-6 EGF Millipore Sigma Cat#SRP3027 ITS-G Thermo Scientific Cat# 41400045 HaltTM Protease Inhibitor Cocktail (100X) Thermo Scientific Cat# 78430 Bestatin (Ubenimex, NK421) SelleckChem Cat# S1591 Tosedostat MedChemExpress Cat# HY-14807 ARM-1 MedChemExpress Cat# HY-W027340 HFI-142 MedChemExpress Cat# HY-110259 Actinonin MedChemExpress Cat# HY-113952 Amastatin hydrochloride MedChemExpress Cat# HY-115194 SC-57461 MedChemExpress Cat# HY-103226 DG051 MedChemExpress Cat# HY-10825 Acebilustat MedChemExpress Cat# HY-17625 Firibastat MedChemExpress Cat# HY-109058 Docetaxel SelleckChem Cat# S1148 Carboplatin SelleckChem Cat# S1215 Doxorubicin SelleckChem Cat# E2516 DMSO Millipore Sigma Cat# D8418 Blasticidin Sigma Cat# 15205 Puromycin Sigma Cat# P8833 Sodium azide Millipore Sigma Cat# S2002 7-AAD BioLegend Cat# 420404 Trizol Reagent invitrogen Cat# 15596026 Paraformaldehyde Millipore Sigma Cat# 158127 Tween 80 Millipore Sigma Cat# P4780 Triton X-100 Millipore Sigma Cat# 9036-19-5 Goat serum Gibco Cat# 16210064 ER-Tracker Blue-White DPX Invitrogen Cat# E12353 ProLong Glass Antifade Mountant Invitrogen Cat# P36980 pHrodo Red, SE Invitrogen Cat# P36600 Matrigel Sigma Cat# CLS356237 Critical commercial assays PrestoBlue Cell Viability Reagent Thermo Scientifc Cat# A13261 ProtoScript II First Strand cDNA Synthesis Kit NEB Cat# E6560 PureYield Plasmid Miniprep System Promega Cat# A1222 PureLink HiPure Plasmid Midiprep Kit Invitrogen Cat# K210004 RNeasy Mini kit Qiagen Cat# 74104 True-Nuclear Transcription Factor Buffer Set BioLegend Cat# 424401 Human Tumor Dissociation Kit Miltenyi Biotec Cat#130-095-929 Phusion High-Fidelity PCR Kit Thermo Scientific Cat# F553L Deposited data CRISPRscreen Zenodo Zenodo: 10.5281/zenodo.10712119 scRNA-seq data from Zhang et al.75 GEO GEO: GSE165897 LOGpc GEO GEO: GSE63885 cBioPortal database Cerami et al.25, Li et al.36 http://www.cbioportal.org/ TIMER 2.0 Li et al.36 http://timer.cistrome.org/ (Continued on next page) Cell Reports 43, 114041, April 23, 2024 19

Techniques: In Vitro, Phagocytosis Assay, Control, In Vivo