Trehalose dehydrate

5-FU, Ch of low molecular weight, β-glycerophosphate (β-GP) disodium salt, and trehalose dehydrate were from Sigma-Aldrich (St Louis, MO, USA). ChHCl was prepared as previously described.13 (link) Fluorescein isothiocyanate (FITC) was from Sigma-Aldrich. The QA-Ch conjugates QA-Ch50 and QA-Ch60 and thiolated derivatives QA-Ch50-SH and QA-Ch60-SH were synthesized from Ch according to Zambito et al.10 (link) For the Ch derivatives, the figures 50 and 60 refer to the respective temperatures (°C) at which the synthesis was carried out. Hyaluronic acid (viscometric molecular weight 470 kDa) was prepared as described by Zambito et al.10 (link) Nylon 6.6 and cellulose membranes were soaked for at least 24 hours in water before use. All aqueous solutions/dispersions were prepared with deionized water. All other chemicals and solvents were of reagent grade. FITC labeling of Ch was carried out as previously described.14 (link)–17 (link)

The mIDH1-U87 (U87 cell line stably transfected with IDH1-R132H with a lentiviral vector, and was kindly provided by Marc Sanson [Hôpital de la Salpêtrière, Paris, France]) was maintained in Eagle’s minimal essential medium (EMEM) with 10% foetal calf serum, 2 mM L-glutamine, 100 U/mL penicillin, and 100 μg/mL streptomycin (Lonza, Verviers, Belgium). bevacizumab (Roche, Paris, France) was diluted with culture medium to working concentrations before use. As a control, a stock solution containing the corresponding excipient was prepared with 60 mg/mL of trehalose dehydrate, 5.8 mg/mL sodium dihydrogen phosphate monohydrate, and 1.5 mg di-sodium hydrogen phosphate dihydrate (all from Sigma Aldrich, Saint-Quentin Fallavier, France). VEGF secretion was assessed with the Quantikine ELISA kit for human VEGF (R&D Systems, Abingdon, UK), following the manufacturer’s instructions. The cells proliferation was assessed as follows: The mIDH1-U87 cells were seeded into and allowed to attach overnight. Cells (seeded in 24-well plates; 30,000 cells/well) were treated for 72 h with bevacizumab (from 0.1 mg/mL to 1 mg/mL) in triplicate wells. Duplicate experiments were performed. Cell viability was then assessed with the MTT assay following the procedure of Mosmann.13 (link) The percentage of surviving cells is expressed as the ratio of optical density of treated cells versus untreated cells.