Aim To determine the effect of VIR576, a dimeric 20mer peptide potently inhibits HIV1 entry, on antigenspecific T cell and nonantigenspecific T cell activation.Methods In vitro Tcell proliferation assays were estalished to investigate the potential effect of FP16, VIR576 on the proliferation of A2b cells in response to MOG3555.A fluorescencebased binding assay using Rhodamine (Rho)conjugated VIR576 was estalished to evaluate the potential interaction between VIR576 and TCRTMD.Fluorescence confocal microscopy was used to to study whether VIR576 could colocalize with CD4 molecule in the CD4 + T cell membrane to interact with TCR.Results The effects of VIR576 on the proliferation of MOGspecific A2b T cells in response to the stimulation of MOG 3555 peptide wasevaluated.VIR576 itself could directly inhibit antigenspecific Tcell activation.Further studies confirmed that VIR576 also inhibited the proliferation of splenocytes and primary CD4 + CD25T cells isolated from the spleens of DOll.10 OVA Tg mice in response to OVA stimulation in vitro.However, VIR576 had no effect on the proliferation of normal mouse splenocytes and T lymphocytes stimulated with Con A or antiCD3 antibody.The FRET assay confirmed that VIR576 effectively binds to the core peptide (CP), corresponding to the Nterminal 9residue region of TCRTMD.Confocal microscopy revealed that VIR576 colocalizes with CD4 on the activated CD4 + Tcell membrane, particularly within the activation cluster including reassembled CD4 and TCR molecules.Conclusion These results suggest that VIR576 is effective in suppressing antigenspecific Tcell activation, but it has no effect on nonspecific Tcell proliferation, and VIR576 has the ability to downregulate antigenspecific Tcell activation by interaction with TCR transmembrane domain.