Fig. 1 |. Nanocomplex-decorated Microbubbles (ncMBs) targeting CD11b on antigen-presenting cells (APCs).

a, ncMBs are obtained by conjugating MBs with anti-CD11b antibodies (aCD11b) and spermine-conjugated dextran (SpeDex) and loading negatively charged cGAMP. b, Upon binding of ncMBs to APCs and under US exposure, cGAMP is delivered directly into the cytosol of the APCs by sonoporation to activate STING and downstream antitumour immunity, a process termed Microbubble-assisted UltraSound-guided Immunotherapy of Cancer (MUSIC). c, The lipid shell of the MB is partly composed of DSPE-PEG-maleimide, which was conjugated with thiolated SpeDex through a thiol-maleimide coupling reaction. d, Coulter counter measurements of SpeDex-aCD11b MBs (cMBs) show a size distribution of 1–10 μm, with a mean size of 2.6 μm. e,f, A fluorescent analog of cGAMP (DY547-c-diGMP) was used to verify binding to cMBs (forming ncMBs). Flow cytometry (e) and fluorescence microscopy (f) confirmed binding of the fluorescent analog to all ncMBs. Scale bar=50 μm. g, DiD-labeled cMBs were added to EO771 murine breast cancer cells and THP-1 human macrophages to confirm CD11b-specific targeting of cMBs. Confocal microscopy confirmed the binding of cMBs to THP-1 cells. h, Fluorescence microscopy of mouse bone marrow-derived macrophages (BMDMs) after sonoporation with DY547-c-diGMP loaded ncMBs indicate cytosolic delivery of the cyclic dinucleotide into all cells. g,h, Scale bar=100 μm. i, Intensity quantification of the DY547-c-diGMP uptake in BMDMs. The data represent mean ± s.d. with n=3 biologically independent samples (i). All data are shown as representative from at least three independently experiments (d-i), and were analysed by one-way analysis of variance (ANOVA) with Tukey’s multiple comparisons test (i).