Shikun Zhang

The asialoglycoprotein receptor 1 (ASGR1) represents a highly promising target for drug development, with its expression regulation closely linked to various diseases. Consequently, research concentrating on targeted therapies against ASGR1 holds significant importance in devising effective treatment strategies. In this study, we utilized the CRISPR-Knockin technology to insert a firely luciferase reporter gene downstream of exon 9 of the ASGR1 gene in HepG2 cell line. This modification enables the expression level of luciferase to be directly proportional to the activity intensity of the ASGR1 protein. We successfully established a drug screening and evaluation model for ASGR1 and employed it for high-throughput screening of potential inhibitors from a microbial molecular metabolite library. After our screening process, several promising candidates were identified as potential ASGR1 inhibitors. Western blotting experiment was conducted to validate the efficacy of our drug screening model, thereby providing a solid experimental foundation for the development of novel targeted therapeutics targeting ASGR1.

SAAP-148, a derivative of LL-37, exhibits a well-defined amphipathic structure and enhanced antimicrobial activity; however, it also displays significant cytotoxicity towards human cells. In this study, we employed Lys-scan to produce a series of amphiphilic SAAP-148 analogs derived from the SAAP-148 sequence to investigate the impact of the distribution of positively charged residues on the biological viability of the antimicrobial peptides (AMPs). The physical properties and biological activity of the designed peptides were subsequently compared. The substitution of lysine resulted in an increase in the overall charge of SAAP-148 and a decrease in its overall hydrophobicity and hyd. moment, except for SAAP-10 where an analogue substitution occurred at the 18th residue. The replacement of lysine led to a reduction in hemolytic activity compared to SAAP-148, with slightly higher haemolysis rates observed in SAAP-11 and SAAP-13. The cytotoxicity of peptides towards human normal lung epithelial cells (BEAS-2B) was closely linked to their haemolytic activity, indicating that substituting lysine may mitigate the cytotoxic effects of SAAP-148. Additionally, the arrangement of positively charged residues in the peptides significantly influenced its antimicrobial activity. Our findings suggest that the positioning of a positively charged residue has a significant impact on the biophysical properties of the peptide. Additionally, the substitution of lysine at different positions demonstrates an influence on the anti-lipopolysaccharide (anti-LPS) activity of SAAP-148. These discoveries provide valuable insights for the design and optimization of antimicrobial peptides, which will be advantageous for the future development of antimicrobial agents.