Supplementary results, Supplementary Scheme 1 to 2, Supplementary Tables 1, and Supplementary Figures 1 through 7 from Folate Receptor–Targeted Polymeric Micellar Nanocarriers for Delivery of Orlistat as a Repurposed Drug against Triple-Negative Breast Cancer
Authors: Paulmurugan Ramasamy , Bhethanabotla Rohith , Mishra Kaushik , Devulapally Rammohan , Foygel Kira , Sekar Thillai V. , Ananta Jeyarama S. , Massoud Tarik F. , Joy Abraham 
Published: April 2023

<p>1. Supplementary Results: Synthesis of S-(2-aminoethyl) methanesulfonothioate hydrobromide (III); Synthesis of HEA-b-EHA polymer; Synthesis and characterization of HEA-b-EHA micelles for orlistat loading, hydrodynamic size, polydispersity index and zeta potential; Cytotoxic effects of orlistat alone in SkBr3 and MDA-MB-231 cells 2. Supplementary Schemes: Supplementary Scheme 1. Synthesis of NIR-DyLight747-B1 and folate ligand conjugated HEA-b-EHA polymer; Supplementary Scheme 2: Schematic illustration of HEA-b-EHA polymer micelles preparation for Orlistat loading. 3. Supplementary Tables 1: Size, surface charge, and polydispersity index (PDI) of various micelles prepared from HEA-b-EHA polymers. 4. Supplementary Figures: Supplementary Figure 1. Optimization of orlistat drug loading in HEA-b-EHA micellar nanoparticles by altering the polymer to drug ratio; Supplementary Figure 2. Standard graph for estimating orlistat loading in HEA-b-EHA polymer micelles by UV-Vis spectrophotometer; Supplementary Figure 3. a, Schematic structure of orlistat binding to FAS protein. b, Hydrodynamic size of orlistat loaded folic acid and DyLight747-B1 functionalized HEA-b-EHA polymer micelles analyzed by DLS; Supplementary Figure 4. Graph showing PI staining based FACS analysis measures apoptotic cells induced in response to the treatment of different concentration of orlistat for 48 h in SkBr3 (a) and MDA-MB-231 (b) cells; Supplementary Figure 5. Graphs and phase contrast microscopic images showing SkBr3 and MDA-MB-231 cells treated with different combination of orlistat loaded HEA-b-EHA-polymer micelles. SkBr3 cells treated with free orlistat (a), orlistat loaded in non-targeted micelles (b) and folic acid targeted micelles (c). MDA-MB-231 cells treated with free orlistat (d), orlistat loaded in non-targeted micelles (e) and folic acid targeted micelles (f). Phase contrast microscopic images of MDA-MB-231 cells treated with different concentration of free orlistat (g) and orlistat loaded HEA-b-EHA-polymer micelles (h) for 48 h; Supplementary Figure 6. Graphs and phase contrast microscopic images showing MDA-MB-231 cells treated with different concentrations of free orlistat aggregates and Fol-NP-orlistat for 48 h. FACS analysis of MDA-MB-231cells treated with free orlistat aggregates (a), phase contrast images of MDA-MB-231cells treated with free orlistat aggregates (b), FACS analysis of MDA-MB-231cells treated with Fol-NP-orlistat (c), phase contrast images of MDA-MB-231cells treated with Fol-NP-orlistat (d); Supplementary Figure 7. Graphs and phase contrast microscopic images showing MCF10A cells treated with different concentration of free orlistat (0-20μM) or Fol-NP-orlistat for 48 h. FACS analysis of free orlistat treated cells (a), Phase contrast microscopic images of free orlistat treated cells. (b), FACS analysis of Fol-NP-orlistat treated cells. (c), Phase contrast microscopic images of Fol-NP-orlistat treated cells (d). 5. Supplementary references.</p>