Dual‐functional Nanosized Albumin Facilitates CRISPR‐Cas9‐PAR2 to Mitigate Cytokine Storm via TLR4 and Pyroptosis Signaling

Abstract Protease‐activated receptor 2 (PAR2) widely modulates various cytokine secretions and inflammatory responses, while excessive cytokine releases and immune hyperactivation may trigger cytokine storm. However, the potential participation of PAR2 in cytokine storm remains elusive. CRISPR‐Cas9 as an efficient gene editing tool can be used for investigating the possible function of PAR2. In this study, albumin‐based nanoparticles (hAlbumin/cgPAR2) to protect and deliver CRISPR‐Cas9 for precise gene editing of PAR2 is constructed. Nanosized albumin largely assisted CRISPR‐Cas9‐PAR2 (cgPAR2) to enter cells via scavenger receptor‐mediated endocytosis, and then triggered potent transfection efficiency, consequently stimulating PAR2 knockout in vitro and in vivo. Apart from delivering CRISPR‐Cas9, nanosized albumin could be colocalized with TLR4 on endosome and promote PAR2 deficiency to attenuate TLR4/NF‐κB signaling, exerting the strong inhibitory effect of albumin in inflammatory responses depending on TLR4. Moreover, PAR2 deficiency induced by hAlbumin/cgPAR2 alleviated ERK/STING/NLRP3 signaling and GSDMD‐mediated pyroptosis, consequently inhibiting cytokine storm. Therefore, this study uncovered dual functions of nanosized albumin in delivering CRISPR‐Cas9 and facilitating PAR2 deficiency to prevent cytokine storm. It also uncovered that the significant role of PAR2 in modulating cytokine storm and provided a potential gene therapy strategy for treating this disease.