Short peptide-based inhibition of fusion remains an attractive goal in anti-human immunodeficiency virus (HIV) research based on its potential for the development of technically and economically desirable antiviral agents. Herein, we report use of the dithiol bis-alkylation reaction to generate a series of m-xylene thioether-stapled 22-residue α-helical peptides that have been identified as fusion inhibitors targeting HIV-1 glycoprotein 41 (gp41). The peptide sequence is based on the helix-zone binding domain (HBD) of the gp41 C-terminal heptad repeat (CHR) region. We found that one of these stapled peptides, named hCS6ERE, showed promising inhibitory potency against HIV-1 Env-mediated cell-cell fusion and viral replication at a level comparable to the clinically used 36-mer peptide T20. Furthermore, combining hCS6ERE with a fusion inhibitor having a different target site, such as HP23, produced synergistic anti-HIV-1 activity. Collectively, our study offers new insight into the design of anti-HIV peptides with short sequences.