Rational fusion design inspired by cell-penetrating peptide:SS31/S-14 G Humanin hybrid peptide with amplified multimodal efficacy and bio-permeability for the treatment of Alzheimer’s disease

Alzheimer’s disease is a neurodegenerative disease induced by multiple interconnected mechanisms.Peptide drug candidates with multi-modal efficacy generated from fusion strategy are suitable for addressing multi-facet pathology.However,clinical translation of peptide drugs is greatly hampered by their low permeability into brain.Herein,a hybrid peptide HNSS is generated by merging two therapeutic peptides(SS31 and S-14 G Humanin(HNG)),using a different approach from the classical shuttle-therapeutic peptide conjugate design.HNSS demonstrated increased bio-permeability,with a 2-fold improvement in brain distribution over HNG,thanks to its structure mimicking the design of signal peptide-derived cell-penetrating peptides.HNSS efficiently alleviated mitochondrial dysfunction through the combined effects of mitochondrial targeting,ROS scavenging and p-STAT3 activation.Meanwhile,HNSS with increased Aβaffinity greatly inhibited Aβoligomerization/fibrillation,and interrupted Aβinteraction with neuron/microglia by reducing neuronal mitochondrial Aβdeposition and promoting microglial phagocytosis of Aβ.In3×Tg-AD transgenic mice,HNSS treatment efficiently inhibited brain neuron loss and improved the cognitive performance.This work validates the rational fusion design-based strategy for bio-permeability improvement and efficacy amplification,providing a paradigm for developing therapeutic peptide candidates against neurodegenerative disease.

Hybrid Antimicrobial Peptide Buforin Ⅱ-Cecropin B Exhibits Antimicrobial Activity

[ Objective ] This study aimed to investigate the antimicrobial activity of hybrid antimicrobial peptide buforin II-cecropin B. [ Method ] Gene fragment BC encoding the hybrid antimicrobial peptide Buforin II-Cecropin B was synthesized by SOE-PCR with six primers designed according to the published amino acid sequences. Then, the BC gene fragment was ligated into pET32a vector and expressed in BI21. Antimierobial ability of the crude hybrid antimicrobial peptide was detected with the Oxford cup method. [ Result] The BC gene fragment was correctly amplified by SOE-PCR and ligated into pET32a vector, to construct the expression vector pET32-BC. The hybrid antimicrobial peptide buforin II-cecropin B was expressed in BI21 after induction by IPTG. The optimal induction time was 2 h. Antimicrobial activity assay suggested that the hybrid antimicrobial peptide possessed antimicrobial activity to Escherichia coli and Bacillus subtilis. [ Conclusion] The hybrid antimicrobial peptide buforin II-cecropin B exhibited antimicrobial activity to Gram-negative E. coli and Gram-positive B. subtilis strains.

In situ aneuploidy assessment in human sperm:the use of primed in situ and peptide nucleic acid-fluorescence in situ hybridization techniques

Both the primed in situ(PRINS)and the peptide nucleic acid-fluorescence in situ hybridization(PNA-FISH) techniques constitute alternatives to the conventional(fluorescence in situ hybridization,FISH)procedure for chro- mosomal investigations.The PRINS reaction is based on the use of a DNA polymerase and labeled nucleotide in an in situ primer extension reaction.Peptide nucleic acid probes are synthetic DNA analogs with uncharged polyamide backbones.The two procedures present several advantages(specificity,rapidity and discriminating ability)that make them very attractive for cytogenetic purposes.Their adaptation to human spermatozoa has allowed the development of new and fast procedures for the chromosomal screening of male gametes and has provided efficient complements to FISH for in situ assessment of aneuploidy in male gametes.(Asian J Androl 2006 Jul;8:387-392)

Design and characterization of a bifunctional bybrid antibacterial peptide LLH for bactericidal/endotoxin neutralization effects

Objective:To design a bi-functional antimicrobial peptide with bactericidal and endotoxin neutralization activity,and explore its bactericidal properties.Methods:The LBP(86-99)peptides and HLF(1-11)peptides were connected by a GGGS flexible 4-peptide linker to obtain the bi-functional antimicrobial peptide,which was named LLH.The secondary structure characteristics of LLH were analyzed by Emboss software.The minimum inhibitory concentration(MIC)and minimum bactericidal concentration(MBC)of LLH against Escherichia coli ATCC25922 and DH5αwere determined by the microtiter broth dilution method.The bactericidal kinetics of LLH was characterized and its effect on endotoxin neutralization was determined.The hemolysis of LLH was evaluated.Results:LLH carried a positive charge of+9,exhibitedβ-folding andβ-corner structure,and had strong hydrophobicity.The MIC of LLH against Escherichia coli ATCC25922 and DH5αwas 4μM,and the MBC of LLH against Escherichia coli ATCC25922 and DH5αwas 8 and 4μM,respectively.LLH showed rapid bactericidal effects and significantly neutralized the endotoxin released in the sterilization process as well as reducing release of endotoxin.LLH showed no significant hemolysis at concentrations up to 400μg/mL.Conclusion:LLH produces dual effects of rapid sterilization and endotoxin neutralization,and does not induce significant hemolysis.

Design and characterization of a bi-functional bybrid antibacterial peptide LLM against Pseudomonas aeruginosa

Objective:To design a bifunctional antimicrobial peptide with high antibacterial activity and endotoxin neutralization against P.aeruginosa and explore its bactericidal properties.Methods:The neutralizing endotoxin peptides and antimicrobial peptide against P.aeruginosa were connected by a GGGS-linker to get a bi-functional bybrid peptide.Its structural parameter was tested by EMBOSS software.The minimum inhibitory concentration(MIC),minimum bactericidal concentration(MBC)and bactericidal kinetics against P.aeruginosa were determined.Its effect on endotoxin neutralization and hemolysis were also evaluated.Results:We designed and obtained a bybrid peptide LLM,which carried+8 positive charge with high activity against P.aeruginosa and neutralizing endotoxin.The MIC of LLM against P.aeruginosa CMCC10104 and P.aeruginosa ATCC 9027 were 2 and 4μmol/L,and MBC/MIC equal 1.LLM showed rapid anti-P.aeruginosa effects and significantly neutralized the endotoxin released,and not exhibited hemolysis as high as 115μmol/L(400μg/mL).Conclusion:The MICs of LLM against P.aeruginosa were 2~4μmol/L,it showed significant activity of anti-P.aeruginosa and neutralizing endotoxin,and could kill bacteria quickly,and did not show significant hemolytic under 115μmol/L.