Despite the intrinsic properties of AMPs that make them highly attractive for a potential use, relatively few of them have been successfully translated into the clinical use or as food preservatives (Mishra et al., 2017; Costa et al., 2019; Adaro et al.; Koniuchovaitė et al.). One of the key constraints is the mismatch between their in vivo and in vitro activities. Particularly frustrating is the fact that highly anticipated peptides such as pexiganan, iseganan, neuprex and omiganan have failed in phase III clinical trials due to low in vivo efficacy (http://dramp.cpu-bioinfor.org/). Many factors may contribute to the low bioavailability in vivo. However, poor stability of these molecules in complex microenvironments has been identified as the most significant factor (Jiang et al., 2021; Fu et al.; Guevara-Lora et al.; Skłodowski et al.).

One of the important prerequisites for clinical use is the drug safety, and this is the second major obstacle on the way of AMPs toward clinical translation (Payne et al., 2015). Toxicity of AMPs includes cytotoxicity and systemic toxicity (Li et al., 2017). Cytotoxicity is usually an inherent property of membrane-active AMPs, the cationic and hydrophobic components of which can directly interact with the membrane of host cells (Agrillo et al.), This interaction is exhibited in a concentration-dependent toxicity. Typical examples are melittin, CZS-1 and alamethicin, which exhibit potent cytotoxicity, including hemolysis (Askari et al., 2021; Farid et al., 2023; Bermúdez-Puga et al.; Brakel et al.). Considering the potential cytotoxic mechanisms of AMPs relative to their successful application, it can be generally concluded that narrow-spectrum peptides are relatively safer for clinical translation due to their lower cytotoxicity and the lack of off-target effects against the beneficial microbiota (Xu et al., 2020; Zong et al., 2020). Conversely, broad-spectrum AMPs tend to display the increased cytotoxicity toward the host and adverse effects on the microbiota, thereby limiting their potential for clinical use (Hao et al., 2023). Systemic toxicity may result from off-target effects, accumulation of drug in kidneys, undesirable immune responses or chronic inflammation (e.g., atopic dermatitis or hidradenitis suppurativa) due to the increased drug concentrations (Takahashi et al., 2018). Therefore, preclinical safety evaluation of AMPs should not be limited to basic hemolysis and cytotoxicity but also requires the evaluation of systemic toxicity. In fact, the antimicrobial and immunomodulatory properties and toxicity of AMPs are often compounded. Thus, a careful attention has to be paid to the delicate balance of antimicrobial properties, immunomodulation, and toxicity.

Although several papers in this Research Topic have discussed the pharmacokinetic (PK) properties of AMPs, it has to be emphasized here that PK is still a bottleneck for AMP translation. It is known that the physicochemical properties of AMPs are quite different from the traditional small-molecule chemical drugs. Hence, the PK of traditional small-molecule drugs should be further modified, improved and optimized for AMPs so that the quantitative PK methodology can be successfully applied for this class of antimicrobials (Wang et al., 2012). Therefore, the development of suitable quantification methods for PK of AMPs, which are different from small-molecule chemical drugs, is the 3rd key challenge for their entry into clinical applications (Ewles and Goodwin, 2011; Mercer and O'Neil, 2013). Usually, linear cationic AMPs are rapidly metabolized in vivo and degraded into smaller fragments or amino acids and absorbed as nutrients. This process interferes with the determination of the main four PK parameters such as absorption, distribution, metabolism, and excretion. Although the safety of AMP degradation products, especially amino acids, in vivo is not of a major concern from the nutrient metabolism point of view, it is difficult to determine the concentration of these products with the use of regular analytical tools. Therefore, there is an urgent need for updating PK principles so that they suit to AMPs, especially protocols for their clinical evaluation (Giguère et al., 2017). In brief, we believe that the use of the latest material analysis methods for exploratory pharmacokinetic detection combined with the calculation of PK parameters based on non-compartmental model is an important prerequisite for AMPs to resolve the bottleneck of drug development and transition to clinical practice (Zheng et al., 2022, 2024).

The likelihood of resistance development toward AMPs is generally much lower than that against conventional antibiotics. Numerous parameters influence resistance development, including the dose used, period of application, temperature, exposure/contact with inhibitory substances, and others. Metabolic pathways and genes within bacterial cells can be replaced or compensated over time, as has been shown for defensins derived from plants and polymyxin from microbes (Ouyang et al.); On the contrary, molecules that have multiple targets in bacteria are less likely to select for bacterial resistance. AMPs with the low probability of resistance development include melittin, bombesin, venoms and cecropins (Chen et al., 2022a). Additional attention has to be paid to AMP-induced cross-resistance. Chen et al. (2022b) found that Staphylococcus aureus acquired limited resistance to PIS-3, with a concomitant resistance toward polymyxin B, vancomycin, and tetracycline, but with no resistance development toward PIS-1. Thus, it is important to gain a better understanding of pharmacology, evolutionary effects and potential resistance acquisition during the development and application of AMPs, the above steps have been largely ignored in the past with traditional antibiotics (Lazzaro et al., 2020).

The molecular stability of AMPs is another important parameter to take into account. The stability of these agents needs to be sufficient to exert their function, ideally without causing off-target effects. At the same time, when assessing peptide stability, it is necessary to focus on the route of administration as this may substantially affect stability.

After exploration toward clinical translation for over 50 years, there is a huge number of publications and patents with innovative results on AMPs, but also there is still room for improvement, and it is expected that the original intentions could be realized as soon as possible (Zasloff, 2015; Czaplewski et al., 2016; Arciola et al., 2018).

Two main AMPs production routes include chemical synthesis and recombinant expression. Chemical synthesis can be executed via solid and liquid phase synthesis methods or their combination. The representative examples of peptide-based drug production at multi-ton scale are HIV fusion inhibitory peptide T-20 (Fuzeon, Roche), semaglutide and insulin (Walsh, 2005; Thayer, 2011; Aggarwal et al., 2021). With technological advances, more and more AMPs including those longer than 30 amino acids, with complex structures and modification processes, will be industrialized utilizing chemical synthesis or transgenic expressions at acceptable costs. Multiple studies have been published recently along this line. For example, optimisation of culture conditions of recombinant Pichia pastoris and induction process of cathelicidin BF expression allowed to reach the product concentration of 0.5 g/L after 240 h (Dong et al.). Li and Chen engineered synthetase to create a synthetic pathway for the production of a novel fusaricidin and constructed the recombinant M6 yielding a 55 mg/L of fusaricidin LI-F07a. In addition, the codon use optimisation in heterologous expression of AFP in E. coli allowed to reach its production at 780 μg/ml (Chen Y.-P. et al.). Different strategies have been used to optimize AMPs production and develop large-scale facilities, some of which were successfully accomplished. For example, after nearly two decades of efforts, Wang's team has successfully established a 20 and 30 cubic meter-scale production system for high-yield preparation of plectasin analogs with an affordable cost comparable to traditional antibiotics, a milestone for the translation of AMPs (Zhang et al., 2014; Yang et al., 2022; Hao et al., 2023; Jin et al., 2023; Li et al.).