Mammalian cathelicidins are a class of innate antimicrobial peptides isolated from leukocytes and epithelial cells that aid host defense against bacterial infections. Synthetic analogs of cathelicidins offer the promise of potent broad-spectrum antimicrobial efficacy. We developed a combined lung infection and ex vivo whole-blood assay model to characterize the toxicity and efficacy of synthetic cathelicidin-derived peptides. Male C57BL/6 mice were administered saline or Klebsiella pneumoniae by intratracheal instillation. Five hours later, the Klebsiella-infected mice were instilled with saline, tobramycin (1 mg/kg of body weight or 10 mg/kg), novispirin G10 (0.4 mg/kg), or a combination of tobramycin (1 mg/kg) and G10 (0.4 mg/kg). At 24 h, bronchoalveolar lavage fluid (BAL) was collected for analysis of culturable bacteria and for markers of inflammation and lung toxicity. Blood samples were analyzed for circulating cytokines. Recovery of Klebsiella from the lung, recruitment of neutrophils, and production of interleukin-6 (IL-6) in BAL samples were highly correlated (r ؍ 0.68 and 0.84, respectively; P < 0.01). Animals treated with G10 or G10 plus tobramycin had increased hemoglobin (P < 0.001) and protein (P < 0.001) levels compared to those for Klebsiella-infected or tobramycin-alonetreated animals. The levels of circulating IL-6 in mice infected with Klebsiella were 1000-to 10,000-fold higher than in the noninfected controls. The highest levels of IL-6 were measured in mice given G10 alone or in combination with tobramycin. These studies demonstrated that G10 was relatively nontoxic in saline-treated mice but was highly toxic in mice infected with Klebsiella. This finding establishes the importance of investigating candidate antimicrobial agents in an in vivo infection model.The search for new and effective antimicrobial agents to treat bacterial or fungal infections has focused in recent years on components of the innate defense systems of plants and animals. Peptides isolated and purified from phagocytic cells have been especially promising as agents against multidrugresistant bacteria and special-niche bacteria associated with chronic conditions such as cystic fibrosis (6,7,10,14,17,27). A class of antimicrobial peptides that has been particularly well studied is the mammalian-derived cathelicidins. These peptides share a highly conserved N-terminal domain identical to a cathelin protein (hence the class name) but are structurally diverse at the C terminus, which also determines the antimicrobial activity of the peptide (14,27,28,29). Lipid A binding and endotoxin neutralization are also determined by the C terminus (2,5,11,12,15,18,29).The members of one class of promising antimicrobial peptides resemble the 18 amino acids of the N terminus of SMAP-29, derived from sheep neutrophils (16,19). While showing a broad spectrum of antimicrobial activity and low toxicity in ovine models, the parent compound, SMAP-29, exhibits unacceptably high hemolytic and cytotoxic activity in human cells (1,16). A series ...