Antibiotic resistance is one of the main problems concerning public health or clinical practice. Antimicrobial peptides appear as good candidates for the development of new therapeutic drugs. In this study we de novo designed a group of cationic antimicrobial peptides, analyzed its physicochemical properties, including its structure by circular dichroism and studied its antimicrobial properties against a panel of clinical isolates expressing different mechanisms of resistance. Three cationic alpha helical peptides exhibited antimicrobial activity comparable to, or even better than the comparator omiganan (MBI-226). De novo designed alpha helical cationic peptides with antimicrobial activity against multi-resistant clinical isolates. Display omitted •De novo designed cationic peptides.•Physicochemical properties of peptides.•Antimicrobial resistant bacteria.