Fig. 9: si < (EG18L)2 uptake is driven by hydrophobicity and does not depend on active endocytic processes, while being partially susceptible to FcRn-mediated extracellular recycling when albumin bound. | Nature Communications

Fig. 9: si < (EG18L)2 uptake is driven by hydrophobicity and does not depend on active endocytic processes, while being partially susceptible to FcRn-mediated extracellular recycling when albumin bound.

From: Structural optimization of siRNA conjugates for albumin binding achieves effective MCL1-directed cancer therapy

Fig. 9

a Cy5 fluorescence measured by flow cytometry of MDA-MB-231 cells incubated with 100 nM Cy5-labeled si < (EG18L)2 or si < (EG18Ldiacid)2 in serum-free media, with or without albumin, for 4 h at 37 °C. b Cy5 fluorescence measured by flow cytometry of MDA-MB-231 cells incubated with 100 nM Cy5-labeled si < (EG18L)2 or si < (EG18Ldiacid)2 in OptiMEM for 2 h at 37 °C vs. 4 °C. c HEK293 cells were transfected to express NanoLuc-tagged human FcRn and treated with media only, Alexa Fluor 647-BSA (positive control), or Cy5-labeled siRNA si < (EG18L)2 with or without BSA. BRET signal (ratio of 670 nm vs. 460 nm signal) was measured. d Western blot of cell lysates collected from WT MDA-MB-231 or cells treated with shFcRn. Time course of siRNA < (EG18L)2 recycling in MDA-MB-231 parental vs. shFCGRT cells in the presence (e) or absence (f) of albumin, measured as siRNA concentration in media following 24 h incubation with 1 µM siRNA < (EG18L)2 with or without 1 µM unlabeled human serum albumin. Concentrations were adjusted for cell quantity using CellTiter Glo. For (a–c), significance was assigned by one way ANOVA with Sidak’s multiple comparisons test (n  = 2–3 experimental replicates). For (d, e) significance was assigned using an unpaired t-test on the AUC of the curves (n = 3 experimental replicates per timepoint).

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