An Amphiphilic Cell-Penetrating Macrocycle for Efficient Cytosolic Delivery of Proteins, DNA, and CRISPR Cas9.

The discovery of safe platforms that can circumvent the endocytic pathway is of great significance for biological therapeutics that are usually degraded during endocytosis. Here we show that a self-assembled and dynamic macrocycle can passively diffuse through the cell membrane and deliver a broad range of biologics, including proteins, CRISPR Cas9, and ssDNA, directly to the cytosol while retaining their bioactivity. Cell-penetrating macrocycle CPM can be easily prepared from the room temperature condensation of diketopyrrolopyrrole lactams with diamines. We attribute the high cellular permeability of CPM to its amphiphilic nature and chameleonic properties. It adopts conformations that partially bury polar groups and expose hydrophobic side chains, thus self-assembling into micellar-like structures. Its superior fluorescence makes CPM trackable inside cells where it follows the endomembrane system. CPM outperformed commercial reagents for biologics delivery and showed high RNA knockdown efficiency of CRISPR Cas9. We envisage that this macrocycle will be an ideal starting point to design and synthesize biomimetic macrocyclic tags that can readily facilitate the interaction and uptake of biomolecules and overcome endosomal digestion.

Cell-Type-Specific CRISPR/Cas9 Delivery by Biomimetic Metal Organic Frameworks.

Effective and cell-type-specific delivery of CRISPR/Cas9 gene editing elements remains a challenging open problem. Here we report the development of biomimetic cancer cell coated zeolitic imidazolate frameworks (ZIFs) for targeted and cell-specific delivery of this genome editing machinery. Coating ZIF-8 that is encapsulating CRISPR/Cas9 (CC-ZIF) with a cancer cell membrane resulted in the uniformly covered C3-ZIF(cell membrane type). Incubation of C3-ZIFMCF with MCF-7, HeLa, HDFn, and aTC cell lines showed the highest uptake by MCF-7 cells and negligible uptake by the healthy cells (i.e., HDFn and aTC). As to genome editing, a 3-fold repression in the EGFP expression was observed when MCF-7 were transfected with C3-ZIFMCF compared to 1-fold repression in the EGFP expression when MCF-7 were transfected with C3-ZIFHELA. In vivo testing confirmed the selectivity of C3-ZIFMCF to accumulate in MCF-7 tumor cells. This supports the ability of this biomimetic approach to match the needs of cell-specific targeting, which is unquestionably the most critical step in the future translation of genome editing technologies.

Sustained and targeted delivery of checkpoint inhibitors by metal-organic frameworks for cancer immunotherapy.

The major impediments to the implementation of cancer immunotherapies are the sustained immune effect and the targeted delivery of these therapeutics, as they have life-threatening adverse effects. In this work, biomimetic metal-organic frameworks [zeolitic imidazolate frameworks (ZIFs)] are used for the controlled delivery of nivolumab (NV), a monoclonal antibody checkpoint inhibitor that was U.S. Food and Drug Administration-approved back in 2014. The sustained release behavior of NV-ZIF has shown a higher efficacy than the naked NV to activate T cells in hematological malignancies. The system was further modified by coating NV-ZIF with cancer cell membrane to enable tumor-specific targeted delivery while treating solid tumors. We envisage that such a biocompatible and biodegradable immunotherapeutic delivery system may promote the development and the translation of hybrid superstructures into smart and personalized delivery platforms.