Chronic respiratory infections are the main cause of morbidity and mortality in cystic fibrosis (CF) patients, and are characterized by the development of multidrug resistance (MDR) phenotype and biofilm formation, generally recalcitrant to treatment with conventional antibiotics. Hence, novel effective strategies are urgently needed. Antimicrobial peptides represent new promising therapeutic agents. Here, we analyze for the first time the efficacy of three versions of a cryptide identified in human apolipoprotein B (ApoB, residues 887-922) towards bacterial strains clinically isolated from CF patients. Antimicrobial and anti-biofilm properties of ApoB-derived cryptides have been analyzed by broth microdilution assays, crystal violet assays, confocal laser scanning microscopy and scanning electron microscopy. Cell proliferation assays have been performed to test cryptide effects on human host cells. ApoB-derived cryptides have been found to be endowed with significant antimicrobial and anti-biofilm properties towards Pseudomonas and Burkholderia strains clinically isolated from CF patients. Peptides have been also found to be able to act in combination with the antibiotic ciprofloxacin, and they are harmless when tested on human bronchial epithelial mesothelial cells. These findings open interesting perspectives to cryptide applicability in the treatment of chronic lung infections associated with CF disease.