Reprogramming of B16-F10 melanoma-educated macrophages by STING agonists loaded in mannose-decorated reconstituted high density lipoprotein nanoparticles.
0000-0002-9844-8860 (Dossou, Akpedje)
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Purpose: Representing a large portion of tumor-infiltrating cells, tumor-associated macrophages (TAMs) have the potential to mediate an immune response against the tumor. Instead, they are educated by the tumor microenvironment (TME) to display an immunosuppressive (M2) phenotype that favors tumor progression. The utilization of stimulator of interferon genes (STING) agonists to re-program TAMs to an immunostimulatory (M1) phenotype leads to tumor regression. However, the whole-body distribution of macrophages, the complex TME architecture, and low bioavailability at the TME are challenges to this therapeutic approach. The selective delivery of STING agonists by suitable nanoparticles can help address these challenges. Reconstituted high-density lipoprotein nanoparticles (rHDL NPs) are biocompatible, can penetrate the TME, and interact readily with macrophages. The scavenger receptor class B type 1 (SR-B1) mediates the intracellular delivery of rHDL NPs' payload. Since TAMs highly express scavenger receptors and the mannose receptor CD206, we hypothesize that mannose-decorated rHDL NPs will efficiently deliver STING agonists to TAMs for their repolarization to an M1 phenotype. Thus, the purpose of this study is to assemble and characterize mannose-decorated rHDL NPs and assess the ability of the formulation to deliver STING agonists and polarize B16F10 melanoma-conditioned macrophages to an M1 phenotype. Methods: DSPE-PEG-Mannose (DPM) was used to introduce a mannose moiety onto the rHDL NPs. Two STING agonists (DMXAA, MSA-2) were loaded separately in the rHDL-DPM to form rHDL-DPM-DMXAA and rHDL-DPM-MSA-2. Dynamic light scattering, absorbance- and fluorescence-based measurements were used to evaluate the chemical composition and characterize the formulations. Murine macrophages incubated in B16-F10 melanoma-conditioned media served as an in vitro TAM model. Lipopolysaccharide + interferon-gamma- and interleukin-4 -treated macrophages respectively served as M1 and M2 macrophage references. Western blots and ELISA were used to assess the expression of M1 markers (CXCL10, HLA-DR) and M2 markers (CD36). Results: Similar characteristics, including size, were found for rHDL-DPM-DMXAA and rHDL-DPM-MSA-2. In addition to CD206 expression, B16F10-conditioned macrophages show expression of M2 markers. M2 macrophages and B16-F10-conditioned macrophages showed a higher SR-B1 expression and higher uptake of the payload from rHDL-DPM NPs than M1 macrophages. Treatment with STING agonist-loaded rHDL-DPM diminished CD36 expression and induced HLA-DR and CXCL10 expression in B16F10-conditioned macrophages. Conclusions: The above findings show that the rHDL-DPM NPs can serve as a delivery vehicle for both DMXAA and MSA-2 and potentially can be extended to other TAM-repolarizing drugs. The expression of SR-B1 and payload uptake by the B16-F10-conditioned macrophages validate the utility of rHDL-DPM NPs to efficiently target TAMs. In addition, the induced expression of CXCL10 could be beneficial for the recruitment of CD8+ T-cells when rHDL-DPM NPs are used in combination with T-cell-based immunotherapies to improve treatment outcomes for cancer patients.