Abstract 2198: Cancer cell membrane coated biomimetic nanoparticles: Synthesis, characterization, and functionality

Abstract

Abstract Introduction: Biomimetic nanoparticles (NPs) combining synthetic and biological materials have flexibility and functionality for drug delivery and immunomodulation.1 Red blood cell (RBC) membranes coated poly(lactic-co-glycolic acid) (PLGA) NPs that mimic RBCs and act as nanosponges for toxins were recently described.2-3 Here, we coated cancer cell membranes onto PLGA NPs in a “right-side” out manner, to translocate membrane anchored proteins onto NPs. Cancer cell membrane coated PLGA NPs hold promise for disrupting cancer cell-stromal cell interactions, and for priming the immune system in cancer immunotherapy. Method: Plasma membrane fractions (MFs) of U87 (low CXCR4) and U87-CXCR4 (high CXCR4) cells were isolated upon homogenization, and sucrose density gradient centrifugation. MFs and PLGA NPs were mixed and physically extruded through a porous membrane to obtain MF-coated PLGA NPs. MFs were probed with antibodies against cell fraction markers, and CXCR4. MFs-coated NPs, MFs and PLGA NPs were characterized on size, morphology, and zeta-potential. The orientation of MFs and MF-coated NPs was investigated by confocal microscopy and flow cytometry. Transwell migration assays were performed to investigate the migration of cancer cells towards human mammary fibroblasts (HMFs). Immune-competent Balb/c mice were immunized with IR700-labeled MFs or MF-coated NPs via subcutaneous injection through hock and imaged in vivo and ex vivo by near-infrared (NIR) fluorescence. Immune responses to MFs or MF-coated NPs were examined on activating CD4+ and CD8+ T lymphocytes in lymph nodes and spleens by flow cytometry. Results: Plasma membrane purity was confirmed from western blot analysis that showed the significant enrichment of Na+/K+-ATPase, negligible amount of GPR78 or GAPDH in MFs. PLGA NPs, U87-CXCR4 MFs and U87-CXCR4 MFs-coated NPs had average diameters of 50 nm, 200 nm, and 70 nm, respectively. Z-average diameters and zeta-potential of PLGA NPs, U87-CXCR4 MFs, and U87-CXCR4 MFs-coated NPs were 79.8 nm, 336 nm and 168 nm, and -34.3 mV, -24.9 mV and -25.0 mV, respectively. Confocal fluorescence microscopy and flow cytometry detected intense PE fluorescence and higher CXCR4 in U87-CXCR4 MFs and U87-CXCR4 MFs-coated NPs than U87 counterparts, confirming a “right-side” out orientation. When U87 or U87-CXCR4 MFs were added to HMFs in the transwell assay, fewer cancer cells migrated towards HMFs, identifying the unique ability of MFs in disrupting HMF-cancer cell interactions. U87-CXCR4 MFs and MF-coated NPs were observed in the popliteal lymph nodes, and triggered the induction of CD8+ T lymphocytes, identifying a role for MF-coated NPs in providing cancer cell membranes to antigen presenting cells to induce a tumor-specific immune response.