Current Protein and Peptide Science - Online First

Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer with a high recurrence rate. A new therapeutic intervention is urgently needed to combat this lethal subtype. The identification of biomarkers is also crucial for improving outcomes in TNBC.

The cell cytotoxicity of ML364 (2-(4-Methylphenylsulfonamido)-N-(4-phenylthiazol-2-yl)-4-(trifluoromethyl)benzamide) was measured at different concentrations of ML364 in TNBC-treated and untreated cells. The 2DE and LC-MS/MS analysis were used for protein identification of differentially expressed proteins. Furthermore, the quantitation of gene expression was demonstrated using RT-qPCR. TIMER, HPA, and UALCAN databases were utilized for further analysis.

Differentially expressed proteins and genes after ML364 treatment in TNBC were found to be linked with the USP2 (ubiquitin specific peptidase 2)-mediated pathway. Our results demonstrate that differentially identified proteins, including PPA1, TRIM68, and FBXO46, could be a potential prognostic biomarker for TNBC. Further analysis through the UALCAN and HPA databasess shows the high expression of these proteins in primary breast tumors, which is in contrast to normal. The induction of ML364 significantly reduced the expression of PPA1, TRIM68, and FBXO46 proteins and induced cell cytotoxicity in TNBC cells.

This study provides an understanding of the USP2-mediated signaling pathway in TNBC, emphasizing the role of USP2 and its substrates with apoptotic genes. Our results offer insight into the USP2-mediated cellular mechanism after ML364 treatment in TNBC that could be a potential therapeutic candidate.

Clinic infections caused by various microorganisms are a public health concern. The rise of new strains resistant to traditional antibiotics has exacerbated the problem. Thus, the search for new antimicrobial molecules remains highly relevant.

The current study purified, characterized, and assessed the antimicrobial activity of a papain inhibitor from Terminalia catappa L. seeds.

The inhibitor was purified by heating the crude extract at 80°C for 30 min, followed by ion exchange chromatography on a DEAE cellulose column. The purification index was 9-fold, yielding 2.3%. SDS-PAGE and size exclusion chromatography revealed that the protease inhibitor (TcPI) is a 15.9 kDa monomeric protein. The inhibition kinetics showed that TcPI is a competitive inhibitor specific to papain (Ki = 1.02 x 10^-4 M). TcPI remained active even after heating at 100 ºC for 120 min and at pH conditions varying from 2.0 to 10.0. Even after 60 min, TcPI was resistant to papain proteolysis. TcPI exhibited antimicrobial activity against Candida parapsilosis and Staphylococcus aureus.

Here, we show that TcPI is a highly stable type-1 cystatin with the potential to combat infections caused by C. parapsilosis and S. aureus. Additional investigations into TcPI's structural aspects and mechanism of action, as well as safety assessments, are essential prerequisites for its potential application as a novel therapeutic intervention.

Crotalus Neutralizing Factor (CNF) is a γ-type Phospholipase A2 (PLA2) inhibitor present in the blood of Crotalus durissus terrificus snake. Particularly, CNF inhibits the toxic action of Crotoxin (CTX), which is a major neurotoxin found in C. d. terrificus venom. CTX induces also myotoxic action and demonstrates high selectivity for skeletal muscle fibers. Consequently, CTX can diffuse beyond the site of infection, which can potentially evoke rhabdomyolysis. The present study has evaluated the effects of CTX on myoblasts and myotubes of muscle cells C2C12 in vitro and the effect of CNF on CTX-induced damage.

Cytotoxicity assays were performed by measuring the mitochondrial enzyme dehydrogenase levels. Furthermore, creatine kinase and lactate dehydrogenase levels were used as indicators of muscle damage.

Crotoxin has been found to have cytotoxic effects on C2C12 myoblast cells, while CNF has not shown toxic effects on these cells. Furthermore, the findings have shown CNF (50 µg/mL) to abolish CTX toxicity in myoblasts. The myotubes, differentiated cells, showed no change in mitochondrial respiration when exposed to CNF or CTX, showing greater resistance to the toxic actions of crotoxin.

The data have confirmed the potential of CNF as an anti-myotoxic agent to prevent CTX-damaged myoblasts and increase resistance to the toxic effects of crotoxin on differentiated cells.