Antimicrobial peptides, in particular α-defensins expressed by Paneth cells, control microbiota composition and play a key role in intestinal barrier function and homeostasis. Dynamic conditions in the local microenvironment, such as pH and redox potential, significantly affect the antimicrobial spectrum. In contrast to oxidized peptides, some reduced defensins exhibit increased vulnerability to proteolytic degradation. In this report, we investigated the susceptibility of Paneth-cell–specific human α-defensin 5 (HD-5) and -6 (HD-6) to intestinal proteases using natural human duodenal fluid. We systematically assessed proteolytic degradation using liquid chromatography–mass spectrometry and identified several active defensin fragments capable of impacting bacterial growth of both commensal and pathogenic origins. Of note, incubation of mucus with HD-5 resulted in 255–8,000 new antimicrobial combinations. In contrast, HD-6 remained stable with consistent preserved nanonet formation. In vivo studies demonstrated proof of concept that a HD-5 fragment shifted microbiota composition (e.g., increases of Akkermansia sp.) without decreasing diversity. Our data support the concept that secretion of host peptides results in an environmentally dependent increase of antimicrobial defense by clustering in active peptide fragments. This complex clustering mechanism dramatically increases the host’s ability to control pathogens and commensals. These findings broaden our understanding of host modulation of the microbiome as well as the complexity of human mucosal defense mechanisms, thus providing promising avenues to explore for drug development.

Background:To evaluate the benefits of teduglutide in a real-life setting, we analyzed the data of 14 patients with short bowel syndrome treated with teduglutide. Additionally, we studied glucagon-like peptide 2 (GLP-2) receptor expression in samples of small intestinal and colonic tissue to provide explanations for clinical observations.Methods:Stool frequency and consistency, sensation of thirst, parental calorie or fluid uptake and the number of days on parenteral support per week were collected for up to 2 years. Quantitative real-time polymerase chain reaction of the GLP-2 receptor in healthy controls was performed to better understand clinical response in different patient subgroups.Results:There was a significant reduction in parenteral support after 24 and 48 weeks (by 11.0 and 36.6%, respectively; p < 0.05). Further major improvements were made in several patients after over 1 year (reduction by 79.3%, p < 0.05). The proportion of patients who reduced parenteral support by at least 20% was 33.3%, 54.5% and 71.3% after 24 weeks, 48 weeks and beyond 1 year, respectively. Patients on daily parenteral support showed late but strong amelioration. The reduction of thirst was the earliest marker for response. While stool consistency increased (p < 0.01), stool frequency decreased (p < 0.05) significantly after 12 weeks. This reduction was even more pronounced in patients with colon in continuity. Supporting these clinical observations, we found a stronger physiological expression of the GLP-2 receptor in the colon than in the small intestine.Conclusions:Patients benefit from teduglutide in a real-life setting, but in contrast to randomized, controlled studies reduction of parenteral support took longer. We identified early clinical markers of response, such as stool consistency and frequency as well as sensation of thirst. Clinical and molecular observations support the role of the colon as an important target organ of teduglutide.

Background/Aims Ustekinumab is effective in active Crohn’s disease. In a retrospective study, we assessed the clinical outcome in nonresponders to anti-tumor necrosis factor therapy, and/or conventional therapy and/or the α4β7-integrin inhibitor vedolizumab. As approval study populations do not always reflect the average “real world” patient cohort, we assessed weather patients who would not have qualified for approval studies show similar outcomes. Methods Forty-one patients with mild to severe active Crohn’s disease were treated with ustekinumab (intravenous 6 mg per kg/body weight) followed by subcutaneous ustekinumab (90 mg) at week 8. Depending on the clinical response maintenance therapy was chosen every 8 or 12 weeks. Clinical response was defined by Crohn’s Disease Activity Index (CDAI) decline, decline of stool frequency or clinical improvement. Inclusion criteria for approval studies were assessed. Results The 58.5% (24/41) showed clinical response to ustekinumab. The 58.3% of this group (14/24) achieved clinical remission. Clinical response correlated significantly with drop of stool frequency and improvement of CDAI score. The 39 out of 41 patients had no side effects and we observed no serious infections. About a third of our patients would not have met ustekinumab approval study criteria. However, patients who did not meet study criteria showed clinical improvement numerically in the same range compared to patients who would have qualified for approval studies. Conclusions Ustekinumab is effective, safe and well tolerated in a highly therapy refractory patient cohort. Even though a reasonable number of patients did not meet ustekinumab approval study criteria, approval study results seem to be representative to the overall patient cohort.

Microbial resistance against clinical used antibiotics is on the rise. Accordingly, there is a high demand for new innovative antimicrobial strategies. The host-defense peptide human beta-defensin 1 (hBD-1) is produced continuously by epithelial cells and exhibits compelling antimicrobial activity after reduction of its disulphide bridges. Here we report that proteolysis of reduced hBD-1 by gastrointestinal proteases as well as human duodenal secretions produces an eight-amino acid carboxy-terminal fragment. The generated octapeptide retains antibiotic activity, yet with distinct characteristics differing from the full-length peptide. We modified the octapeptide by stabilizing its termini and by using non-natural D-amino acids. The native and modified peptide variants showed antibiotic activity against pathogenic as well as antibiotic-resistant microorganisms, including E . coli , P . aeruginosa and C . albicans . Moreover, in an in vitro C . albicans infection model the tested peptides demonstrated effective amelioration of C . albicans infection without showing cytotoxity on human cells. In summary, protease degradation of hBD-1 provides a yet unknown mechanism to broaden antimicrobial host defense, which could be used to develop defensin-derived therapeutic applications.

The occurrence and spread of multidrug-resistant bacteria is a prominent health concern. To curb this urgent threat, new innovative strategies pursuing novel antimicrobial agents are of the utmost importance. Here, we unleashed the antimicrobial activity of human neutrophil peptide-4 (HNP-4) by tryptic digestion. We identified a single 11 amino acid long fragment (HNP-4 1−11 ) with remarkable antimicrobial potential, exceeding that of the full length peptide on both mass and molar levels. Importantly, HNP-4 1−11 was equally bactericidal against multidrug-resistant and non-resistant strains; a potency that was further enhanced by N-and C-terminus modifications (acetylation and amidation, respectively). These observations, combined with negligible cytotoxicity not exceeding that of the full length peptide, presents proteolytic digestion of innate host-defense-peptides as a novel strategy to overcome the current health crisis related to antibiotic-resistant bacteria.