Supplementary information Stability, Permeability and Growth-Inhibitory Properties of Gonadotropin-Releasing Hormone Liposaccharides Daryn Goodwin1, Pegah Varamini1, Pavla Simerska1, Istvan Toth1,2* 1 School of Chemistry and Molecular Biosciences, The University of Queensland, St. Lucia 4072, Queensland, Australia; 2The School of Pharmacy, Pharmacy Australia Centre of Excellence, Wooloongabba 4102, Queensland, Australia *Corresponding author: School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, Queensland, Australia, 4072 Tel: +61 7 33469892 Fax: +61 7 33654273 email: i.toth@uq.edu.au Figure S1. Lipidation strategy using activated lipoamino acid-NHS ester on Tyr. Figure S2. Stability of C8 conjugated GnRH(glyco)lipids in Caco-2 cell homogenates (mean ± SEM). Addition of lipoamino acids and carbohydrates conferred increases in half-life during incubation. Peptides modified with dimeric addition of lipoamino acid were detected in the highest concentrations at completion of the assay. Figure S3. The generation of native GnRH (10) in the receiver chamber demonstrated indicated terminal cyclisation of [E]GnRH (1) during incubation with Caco-2 cell monolayers (mean ± SD). Figure S4. The trans-epithelial electrical resistance (TEER) of the cell monolayer measured after incubation with (glyco)lipopeptides (mean±SD). TEER was measured at t0 (before addition of conjugates), at 3 h (after removal of conjugates), and 16 h (overnight). Figure S5. Viability of PC-3 cells as a percentage of cells treated with PBS (mean±SEM) after 48h incubation with C8- and C12-(glyco)lipopeptides (at 100 µg/mL) in 10% DMSO. Statistical analysis was performed using a one-way ANOVA followed by the Dunnett’s post hoc test and compared to DMSO group (*, p > 0.05; **, p < 0.01; ***, p < 0.001). a b Figure S6. Viability of SKOV-3 cells as a percentage of cells treated with PBS (mean±SEM) after 48h incubation with C8- (a) and C12-(glyco)lipopeptides (b) in 10% DMSO. Statistical analysis was performed using a one-way ANOVA followed by the Dunnett’s post hoc test and compared to DMSO group (*, p > 0.05;***, p<0.001).