A series of 6-polyaminosteroid analogues of squalamine were synthesized with moderate to good yields and evaluated for theirin vitroantimicrobial properties against both susceptible and resistant Gram-positive (vancomycin-resistantEnterococcus faeciumand methicillin-resistantStaphylococcus aureus) and Gram-negative (carbapenem-resistantAcinetobacter baumanniiandPseudomonas aeruginosa)bacterial strains. Minimum inhibitory concentrations against Gram-positive bacteria ranged from 4 to 16 µg/mL for the most effective compounds,4kand4n, and showed an additive or synergistic effect with vancomycin or oxacillin. On the other hand, the derivative4f, which carries a spermine moiety like that of the natural trodusquemine molecule, was found to be the most active derivative against all the resistant Gram-negative bacteria tested, with an MIC value of 16 µg/mL. Our results suggest that 6-polyaminosteroid analogues of squalamine are interesting candidates for Gram-positive bacterial infection treatments, as well as potent adjuvants to fight Gram-negative bacterial resistance.