Cytotoxic and immunomodulatory activity of CD151-LEL-based peptides in breast cancer and THP-1 cells

CD151, a cell surface oncoprotein belongs to the tetraspanin family, drives metastasis, drug resistance, and immune escape, and is recognized as a tumor-associated antigen (TAA). In the screening phase, a positive correlation between CD151 expression and amplification of CCND1, ERBB2, and MYC genes as well with breast cancer (BC) metastasis was observed using data from different GEO datasets. This study identified three immunogenic peptides within the large extracellular loop (LEL) of CD151 using in silico approaches: one each of CTL and HTL epitope, which bind strongly to class I and II MHC molecules, and one B-cell epitope, eliciting a robust IgG immune response. In the validation phase, a multiepitope was constructed by incorporating high immunogenic peptides and adjuvants, which showed significant antigenicity and broader population coverage. The results also showed a hierarchy in the binding of multiepitope with key immune receptor molecules, highlighting their potential in activating the innate and adaptive immune responses. Furthermore, molecular dynamic simulation and normal mode analysis studies uncovered a strong interaction between the multiepitope construct and TLR2. The simulation of immune system activation by the multiepitope construct using the C-ImmSim web server showed a strong B- and T-cell response with potential activation of natural killer and dendritic cells. The results also showed active antigen internalization by macrophages, their presentation to other immune cells, and IL-2 and IFN-γ secretion. In addition, AntiCP web server predicted both CTL and HTL epitopes as anticancer peptides. The in vitro study validated that synthetic peptides corresponding to CD151-LEL-based CTL and HTL peptides induced cytotoxicity in BC cell lines MDA-MB 231 and MCF-7. They reduced the CCND1mRNA levels in MDA-MB-231 and MCF-7 cells, while MYC mRNA levels in MCF-7 cells. Also, they significantly induced the secretion of IFN-γ and IL-2 while reducing the secretion of IL-10. Furthermore, conditioned media from the treated BC cells induced the proliferation of PMA treated monocyte like THP-1 cells and provoked the secretion of IL-6, while reducing IL-10. These findings suggest that the CD151-LEL-based peptides and their engineered multiepitope construct represent a prospective vaccine candidate for in vivo experimental validation, which can be used as a therapeutic vaccine for breast cancer.