AH-8 peptide(BCN Peptides SA)

The synthesis of new analogues of cyclolinopeptide A (CLA) and their linear precursors modified with (R)- and (S)-4-methylpseudoproline in the Pro³-Pro⁴ fragment are presented. The peptides were tested in comparison with cyclosporine A (CsA) in concanavalin A (Con A) and pokeweed mitogen (PWM)-induced mouse splenocyte proliferation and in secondary humoral immune response in vitro to sheep erythrocytes (SRBC). Their effects on expression of selected signaling molecules in the Jurkat T cell line were also determined. In addition, the structural features of the peptides, applying nuclear magnetic resonance and circular dichroism, were analyzed. The results showed that only peptides 7 and 8 modified with (R)-4-methylpseudoproline residue (c(Leu¹-Val²-(R)-(αMe)Ser(ΨPro)³-Pro⁴-Phe⁵-Phe⁶-Leu⁷-Ile⁸-Ile⁹) and c(Leu¹-Val²-Pro³-(R)-(αMe)Ser(ΨPro)⁴-Phe⁵-Phe⁶-Leu⁷-Ile⁸-Ile⁹), respectively) strongly suppressed mitogen-induced splenocyte proliferation and the humoral immune response, with peptide 8 being more potent. Likewise, peptide 8 more strongly elevated expression of Fas, a proapoptotic signaling molecule in Jurkat cells. We postulate that the increased biological activity of peptide 8, compared to the parent molecule and other studied peptides, resulted from its more flexible structure, found on the basis of both CD and NMR studies. CD and NMR spectra showed that replacement of Pro³ by (R)-(αMe)Ser(¬Pro) caused much greater conformational changes than the same replacement of the Pro⁴ residue. Such a modification could lead to increased conformational freedom of peptide 8, resulting in a greater ability to adopt a more compact structure, better suited to its putative receptor. In conclusion, peptide 8 is a potent immune suppressor which may find application in controlling immune disorders.

We aimed to develop inhibitory short peptides that can prevent protein interactions of SOS1/EPS8/ABI1 tri-complex, a key component essential for ovarian cancer metastasis.

Plasmids containing various regions of HA-tagged ABI1 were co-transfected into ovarian cancer cells with Flag-tagged SOS1 or Myc-tagged EPS8. Co-immunoprecipitation and GST-pulldown assay were used to identify the regions of ABI1 responsible for SOS1 and EPS8 binding. Inhibitory short peptides of these binding regions were synthesized and modified with HIV-TAT sequence. The blocking effects of the peptides on ABI1-SOS1 or ABI1-EPS8 interactions in vitro and in vivo were determined by GST-pulldown assay. The capability of these short peptides in inhibiting invasion and metastasis of ovarian cancer cell was tested by Matrigel invasion assay and peritoneal metastatic colonization assay.

The formation of endogenous SOS1/EPS8/ABI1 tri-complex was detected in the event of LPA-induced ovarian cancer cell invasion. In the tri-complex, ABI1 acted as a scaffold protein holding together SOS1 and EPS8. The SH3 and poly-proline+PxxDY regions of ABI1 were responsible for SOS1 and EPS8 binding, respectively. Inhibitory short peptides p + p-8 (ppppppppvdyedee) and SH3-3 (ekvvaiydytkdkddelsfmegaii) could block ABI1-SOS1 and ABI1-EPS8 interaction in vitro. TAT-p + p-8 peptide could disrupt ABI1-EPS8 interaction and suppress the invasion and metastasis of ovarian cancer cells in vivo.

TAT-p + p-8 peptide could efficiently disrupt the ABI1-EPS8 interaction, tri-complex formation, and block the invasion and metastasis of ovarian cancer cells.