Mcf 7 human breast adenocarcinoma cells

MCF-7 human breast adenocarcinoma cells (Fig. 1A) were purchased from the American type culture collection (ATCC) (Manassas, VA, USA). Cells were cultivated in RPMI 1640 medium (Life Technologies, Naerum, Denmark) supplemented with 10% fetal calf serum (FCS) (Biochrom, Berlin, Germany) and 1% penicillin/streptomycin (Biochrom) at 37 °C and 5% CO₂. One day prior to the experimental run on the RPM, cells were seeded either in slide flasks (Thermo Fisher Scientific, Roskilde, Denmark) (3 × 10⁵ cells/cm²) for fluorescence staining or in T25 (1 × 10⁶ cells) vented cell culture flasks (Sarstedt, Nümbrecht, Germany) for RNA and protein extraction. Before starting the run, flasks were filled up with medium, taking care that no air bubbles remain. A detailed procedure has been published previously^{9 (link),23 (link)}. To test the viability of the cells in the multicellular spheroids (MCS), the MCS were collected after 24 h (Fig. 1B) and seeded in slide flasks. The adhesion and migration behavior of the cells of the MCS was examined by phase contrast microscopy after 2 h, 4 h and 24 h (Fig. 1C–E).

CA, PEI (average molecular weight (MW) approximately 1300 Da), PArg (average MW approximately 10,000 Da), N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC), and triethylamine were purchased from Sigma-Aldrich (St. Louis, MO, USA). DOX was purchased from Calbiochem (Merck KGaA, Darmstadt, Germany). Spectra/Por™ dialysis membranes with MW cut-offs of 1000 or 13,000 g/mol were purchased from Spectrum Labs (Rancho Dominguez, CA, USA). MCF-7 human breast adenocarcinoma cells and WRL-68 human hepatic cells were purchased from the American Type Culture Collection (Manassas, VA, USA). Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), trypsin-ethylenediaminetetraacetic acid (EDTA), and (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT) were purchased from Life Technologies (Carlsbad, CA, USA).

MCF7 human breast adenocarcinoma cells were obtained from the American Type Culture Collection (ATCC: Manassas, VA, USA). Dulbecco’s modified Eagle’s medium (DMEM; Lonza, NSW, Australia) with 4.5 g/L glucose, L-glutamine, and sodium pyruvate (Lonza Australia Pty Ltd., Mount Waverley, VIC, Australia), supplemented with 10% foetal bovine serum (FBS; Interpath, Heidelberg West, VIC, Australia) and 100 U/mL of penicillin and streptomycin (Gibco^TM BRL, Scoresby, VIC, Australia) was used to culture MCF7 cell line at 37 °C in the presence of 5% CO₂.

HT29 human colorectal adenocarcinoma cells, HeLa human cervix adenocarcinoma cells, MCF7 human breast adenocarcinoma cells, PC-3 human prostate adenocarcinoma cells, J6 Jurkat Clone E6-1 acute T cell leukemia, the healthy I407 human intestine cells, and mouse embryonic fibroblast 3T3 fibroblasts as controls cell lines were purchased from American Type Culture Collection (ATCC, Manassas, VA). The human ovarian cancer cell line IGROV1 has been kindly provided by Prof. Colnaghi (Istituto Nazionale Tumori (IRCCS) (Milano, Italy). Cells were cultured in RPMI 1640 medium (Labtek Eurobio, Milan, Italy), supplemented with 10% FCS (Euroclone, Milano, Italy) and 2 mM L-glutamine (Sigma-Aldrich, Milano, Italy), at 37 °C, and a 5% CO₂ atmosphere. The compounds were dissolved in DMSO in a 30–40 mM stock solution. In cell treatments, the final DMSO concentration never exceeded 0.1%.