Skip Navigation
Skip to contents

Journal of Microbiology : Journal of Microbiology

OPEN ACCESS
SEARCH
Search
Advanced Search
mobile menu button

Articles

Page Path
HOME > J. Microbiol > Volume 64(3); 2026 > Article
Minireview
 Recent trends in dual-acting hybrid antibiotics and combination therapies against Gram-negative pathogens
Ji Eun Son, Umji Choi, Gyubin Han, Jeongho Lee, Chang-Ro Lee*
Journal of Microbiology 2026;64(3):e2601004.
DOI: https://doi.org/10.71150/jm.2601004
Published online: March 31, 2026

Department of Biological Sciences, Myongji University, Yongin 17058, Republic of Korea

*Correspondence Chang-Ro Lee crlee@mju.ac.kr
• Received: January 6, 2026   • Revised: February 26, 2026   • Accepted: March 4, 2026

© The Microbiological Society of Korea

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

  • 10 Views
  • 2 Download
Full Article

Download PDF Download PDF

  • Antibiotic resistance poses a serious challenge to public health worldwide; however, the development of new antibiotic classes for combating bacterial infections, especially those caused by Gram-negative pathogens, has slowed in recent years. Dual-acting hybrid antibiotics with a metabolically non-cleavable covalent bond represent an emerging strategy for developing novel antibiotic classes to overcome antibiotic resistance. The covalent connection between two antibiotics results in a fixed pharmacokinetic profile of a single molecule and can impede bacterial efflux. However, as most antibiotics do not have membrane-destabilizing activity, the resulting increase in molecular weight by connection of two antibiotics could limit their activity against Gram-negative bacteria, whose outer membrane forms a strong barrier blocking the penetration of high-molecular weight antibiotics. Here, we review recent developments in dual-acting hybrid antibiotics targeting Gram-negative bacteria, with a focus on their antibacterial efficacy. We also discuss combination therapy strategies in which the underlying molecular mechanisms of synergy have been characterized. Finally, we outline future directions for the rational design of hybrid antibiotics against Gram-negative pathogens.
  • Keywords: hybrid antibiotic, combination therapy, synergy, Gram-negative pathogen, molecular mechanism
Antibiotics have been central to clinical medicine since the second half of the 20th century; however, the evolution and spread of antibiotic resistance in pathogenic bacteria pose a serious threat to global public health (Lee et al., 2013). The World Health Organization (WHO) has identified six pathogens as high-priority pathogens because of their serious multidrug resistance abilities. They are collectively known as ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.). In 2019, bacterial antimicrobial resistance was directly linked to 1.27 million deaths, and approximately 95% of these deaths were associated with the six pathogens, including A. baumannii, E. coli, K. pneumoniae, P. aeruginosa, and S. aureus (Antimicrobial Resistance Collaborators, 2022). Among these, all except S. aureus are Gram-negative bacteria.
Bacteria can easily decrease their susceptibility to antibiotics through several mechanisms, including horizontal gene transfer and mutations (Thomas and Nielsen, 2005). Consequently, new weapons for combating bacterial infections are continually required. Despite this, only six new antibiotic classes have been clinically approved in the past 20 years, mostly discovered between the 1930s and the 1960s (Butler et al., 2017). Importantly, none of these classes are particularly effective in the treatment of infections caused by Gram-negative pathogens, such as A. baumannii, E. coli, K. pneumoniae, and P. aeruginosa (Butler et al., 2017). To overcome this decline in antibiotic discovery, various alternate strategies are currently being explored.
The development of dual-acting hybrid antibiotics has emerged as a promising strategy. These hybrids are constructed by ligating two antibiotics together or antibiotics and non-antibiotic moieties such as siderophores, adjuvants, or membrane-active peptides, via a metabolically stable linker, thereby forming a single heterodimer. In the following, we focus exclusively on hybrid antibiotics constructed by ligating two antibiotics together. In principle, such a heterodimeric entity should retain the antibacterial activity of its two constituent pharmacophores. In contrast with combination therapy, in which the two drugs retain independently distinct pharmacokinetic and pharmacodynamic properties, hybrid antibiotics connected by a covalent linker demonstrate fixed pharmacokinetic and pharmacodynamic profiles as a single molecular entity (Fig. 1). Additionally, covalent linkage between two antibiotics can impede bacterial efflux and reduce the development of resistance and toxicity when administrated in vivo (Theuretzbacher, 2020).
Despite the various advantages of hybrid antibiotics with metabolically non-cleavable bonds, they have serious disadvantages when used to treat infections caused by Gram-negative pathogens. These pathogens have an additional outer membrane that blocks the penetration of antibiotics with high molecular weights (O'Shea and Moser, 2008). As most antibiotics do not have outer membrane-destabilizing activity exhibited by colistin, an increase in molecular weight due to the covalent linkage between two antibiotics is a serious limitation of hybrid antibiotics. Indeed, most hybrid antibiotics with dual mechanisms mainly target Gram-positive bacteria (Koh et al., 2023; Pokrovskaya and Baasov, 2010; Theuretzbacher, 2020). While it is true that molecular weight influences uptake of hybrid antibiotics, other physicochemical factors, including polarity, charge distribution, conformational flexibility, amphiphilicity, and susceptibility to efflux, are also important. Therefore, modification of physicochemical properties can enhance the penetration of hybrid antibiotics.
In this review, we summarize recent advances in dual-acting hybrid antibiotics designed to target Gram-negative bacteria and discuss their antibacterial activities. Additionally, we explore recent examples of combination therapy in which the molecular mechanisms underlying the synergistic effects have been elucidated. Finally, we discuss future research directions for the development of hybrid antibiotics targeting Gram-negative bacteria.
Quinolone-containing antibiotic hybrids are the most comprehensively represented hybrid antibiotics. The structure–activity relationships of quinolones has been widely studied and is well defined. The amine group in the C-7 piperidino moiety is a readily acceptable and tolerated position; therefore, it can be used for the attachment of bulky compounds via N-alkylation, carbamate formation, or quaternary salt formation. Additionally, quinolones are chemically stable under various synthetic conditions, facilitating their conjugation with other antibiotics. Ouinolones target two enzymes, DNA gyrase and topoisomerase IV, which is another advantage of this antibiotic. Steric hinderance imposed by the attachment of a partner antibiotic or linker can inhibit interaction with the target protein. In the case of quinolones, this limitation can be compensated by interaction with a second target protein. Nevertheless, quinolones have two disadvantages: rapid resistance development due to spontaneous mutations of target enzymes, and limited availability of antibiotics showing the synergy effect with quinolones in combination therapy.
Among quinolone antibiotics, ciprofloxacin has been extensively used to construct dual-acting antibiotic hybrids (Table 1). Cephalosporin–ciprofloxacin hybrids have been developed (Evans et al., 2019), functioning as dual-acting antibiotics in the absence of β-lactamase, and as prodrugs in its presence. β-Lactamase-mediated hydrolysis of cephalosporins results in the intracellular release of ciprofloxacin, restoring antimicrobial efficacy (Evans et al., 2019). Accordingly, these compounds have exhibited good antimicrobial activity against New Delhi metallo-β-lactamase-expressing E. coli (Table 1).
The ciprofloxacin pharmacophore has also been used to synthesize ciprofloxacin–aminoglycoside hybrid compounds (Pokrovskaya et al., 2009; Shavit et al., 2017). A series of hybrids linking ciprofloxacin and neomycin via a 1,2,3-triazole moiety have been designed and synthesized (Pokrovskaya et al., 2009). Several of these hybrids are significantly more potent than the parent neomycin and showed good antimicrobial activity against neomycin-resistant E. coli (Table 1). More recently, kanamycin–ciprofloxacin hybrid compounds have been reported (Shavit et al., 2017). Their minimal inhibitory concentrations (MICs) against E. coli are comparable to those of neomycin–ciprofloxacin hybrids (Table 1). Notably, these dual-target hybrids have significantly lower resistance frequencies in E. coli than each individual drug or their combination (Shavit et al., 2017). Among the aminoglycosides, tobramycin shows concentration-dependent killing against P. aeruginosa at high concentrations by increasing outer membrane permeability through lipopolysaccharide destabilization (Bulitta et al., 2015; Dhiman et al., 2023; Herzog et al., 2012). Consequently, tobramycin–ciprofloxacin hybrids show strong synergy with several fluoroquinolones against multidrug-resistant P. aeruginosa by enhancing fluoroquinolone penetration (Dhiman et al., 2023; Gorityala et al., 2016). These findings indicate that tobramycin-based hybrids may represent a novel class of antibiotic potentiators.
A series of hybrid antibiotics containing ciprofloxacin and macrolides, such as erythromycin and telithromycin, have also been synthesized (Hutinec et al., 2010; Ma et al., 2019). These compounds have exhibited good antibacterial activity against Haemophilus influenzae, Mycoplasma pneumoniae, Moraxella catarrhalis, and E. coli (Table 1). A recent study reported a hybrid compound with a relatively low MIC (8 μg/ml) against erythromycin-resistant M. pneumoniae (Liu et al., 2022) (Table 1).
Fluoroquinolone–oxazolidinone hybrids are among the most extensively studied fluoroquinolone-based dual-acting antibiotic hybrids (Table 1), and several of these compounds have been reported (Hubschwerlen et al., 2003a, 2003b; Liu et al., 2019; Scaiola et al., 2019). Oxazolidinone antibiotics, such as linezolid and tedizolid, block the initiation of protein synthesis by binding to the peptidyl site of the ribosomal 50S subunit (Bozdogan and Appelbaum, 2004). However, owing to the activity of efflux pumps, most Gram-negative bacteria show intrinsic resistance to oxazolidinones (Schumacher et al., 2007). Interestingly, most of the synthesized fluoroquinolone–oxazolidinone hybrids generally exhibit good antibacterial activity against E. coli and H. influenzae (Table 1), but show only moderate activity against P. aeruginosa (Liu et al., 2019).
Another class of novel fluoroquinolone-based hybrids involves conjugation to the macrocyclic core of ansamycin antibiotics, such as rifampicin and kanglemycin (Peek et al., 2022; Yuan et al., 2020). These antibiotics block mRNA synthesis in bacteria by inhibiting bacterial DNA-dependent RNA polymerase (Calvori et al., 1965). Several of the resulting hybrids have demonstrated good antibacterial activity against E. coli, H. influenzae, and Helicobacter pylori (Table 1). Notably, one hybrid compound, TNP-2092, displayed a significantly low MIC (0.06 μg/ml) against Neisseria gonorrhoeae (Yuan et al., 2020).
Recently, ciprofloxacin-based hybrids fused to sulfonamides, which inhibit bacterial dihydropteroate synthetase in the folic acid pathway (Kratky et al., 2012), have been described (Ibrahim et al., 2022). These compounds exhibit good antibacterial activity against E. coli (Ibrahim et al., 2022). Another antifolate antibiotic, trimethoprim, which binds to dihydrofolate reductase and inhibits the reduction of dihydrofolic acid to tetrahydrofolic acid (Brogden et al., 1982), has been fused with ciprofloxacin (Karoli et al., 2012). One resulting hybrid compound has shown good antibacterial activity against P. aeruginosa and K. pneumoniae, but lacks activity against trimethoprim- and ciprofloxacin-resistant E. coli (Karoli et al., 2012).
Metronidazole is active against most anaerobic and microaerophilic bacteria, but lacks activity against most aerobic bacteria, including E. coli (Leitsch, 2019). Several quinolone-based hybrids incorporating metronidazole have been synthesized (Cui et al., 2014; Zhang et al., 2015), and most have shown good antibacterial activity against P. aeruginosa and E. coli (Table 1). Additionally, many of these hybrids exhibit favorable pharmacokinetic properties, with no obvious toxicity toward human hepatocyte cells (Cui et al., 2014).
Quinolone dimer, including quinolone–fluoroquinolone and fluoroquinolone–fluoroquinolone conjugates, have also been reported (Panda et al., 2015; Ross et al., 2015). However, quinolone–fluoroquinolone conjugates with amino acid linkers have shown poor antimicrobial activity against P. aeruginosa (Panda et al., 2015). Meanwhile, fluoroquinolone–fluoroquinolone conjugates with variable linkers—such as polyethylene glycol chains, basic amines, and aryl linkers—exhibited significantly low MIC values against E. coli and P. aeruginosa (0.06 and 0.125 μg/ml, respectively) (Ross et al., 2015).
In summary, various quinolone-containing antibiotic hybrids have been synthesized, and many have demonstrated significant good antibacterial activity against a range of Gram-negative bacteria (Table 1). Notably, fluoroquinolone-oxazolidinone and fluoroquinolone–rifampicin hybrids have progressed into clinical trials (Koh et al., 2023). However, these compounds are being evaluated primarily for the treatment of bacterial infections caused by Gram-positive bacteria (Koh et al., 2023). To properly assess the therapeutic potential of hybrid antibiotics, studies examining in vivo efficacy and safety using animal models for infection are required. However, these studies were not performed in most hybrid antibiotics in this section. Therefore, on the basis of the present study alone, the efficacy of hybrid antibiotics cannot be clearly determined.
In addition to quinolone-containing antibiotic hybrids, a wide range of hybrid compounds with diverse drug combinations have been described (Table 1). Polymyxins are membrane-targeting decalipopeptide antibiotics that disrupt bacterial membranes and interfere with proton motive forces (Velkov et al., 2013). Polymyxin B3-based hybrid compounds fused with tobramycin, an aminoglycoside antibiotic, have been reported (Domalaon et al., 2017). These hybrid compounds show moderate antibacterial activity against various Gram-negative pathogens, including E. coli, K. pneumoniae, A. baumannii, and P. aeruginosa (Table 1). Notably, they exhibit potent activity against clinical isolates of carbapenem-resistant and colistin-resistant P. aeruginosa and show strong synergy with minocycline, rifampicin, and vancomycin against multidrug-resistant P. aeruginosa (Domalaon et al., 2017). Polymyxin E-vancomycin hybrid conjugates have also been reported (van Groesen et al., 2021). These compounds showed very potent antibacterial activity against vancomycin-resistant Gram-positive pathogens but demonstrate only moderate activity against Gram-negative pathogens, including K. pneumoniae, A. baumannii, and P. aeruginosa (Table 1). Similarly, a vancomycin-based hybrid compound fused to nisin, a natural polycyclic antibacterial peptide, shows moderate antibacterial activity against K. pneumoniae and M. catarrhalis (Arnusch et al., 2008), likely due to the high molecular weight of vancomycin.
A rifamycin-based hybrid fused to nitroimidazole (TNP-2198) showed a significantly low MIC value (0.004 μg/ml) against H. pylori (Ma et al., 2022). The nitroimidazole moiety contains a nitro group and an imidazole ring, as found in metronidazole. TNP-2198 has also demonstrated potent activity against H. pylori strains resistant to both rifamycin and nitroimidazole (Table 1). This compound is currently undergoing Phase II clinical trials for the treatment of H. pylori and C. difficile infections as well as bacterial vaginosis (Koh et al., 2023; Ma et al., 2022).
Further, two antibiotic hybrids containing aminoglycoside antibiotics have also been synthesized (Findlay et al., 2012; Hanessian et al., 2011). Neomycin B hybrids are linked to triclosan, an antibacterial and antifungal agent that inhibits bacterial fatty acid synthesis. They show good antibacterial activity against E. coli, K. pneumoniae, and A. baumannii, but not against P. aeruginosa (Findlay et al., 2012). In contrast, sisomicin-based hybrid compounds fused with gentamicin, another aminoglycoside antibiotic, demonstrate good antibacterial activity against P. aeruginosa, along with E. coli and K. pneumoniae (Hanessian et al., 2011).
Linezolid-based hybrid compounds are fused to sparsomycin, a nucleoside analog of uracil that binds to the 50S ribosomal subunit and inhibits bacterial protein synthesis via peptidyl transferase inhibition (Ottenheijm et al., 1986). These compounds show moderate antibacterial activity against H. influenzae (Zhou et al., 2008). Another oxazolidinone-based hybrid linked to cephalosporin has also been described (Liu et al., 2018). To enhance cellular uptake of hybrid antibiotics, siderophores—high-affinity iron-chelating compounds (Schalk, 2025)—have been incorporated into these hybrid antibiotics. Siderophore-mediated active transport resulted in low MIC values against E. coli, A. baumannii, and P. aeruginosa (0.025, 0.4, and 0.4 μg/ml, respectively) (Liu et al., 2018). In the presence of β-lactamases, such as Acinetobacter-derived cephalosporinase-1 (ADC-1), intracellular release of oxazolidinone upon cephalosporin cleavage allows the freed oxazolidinone to inactivate its target. Consequently, the oxazolidinone–cephalosporin–siderophore conjugate demonstrated a relatively low MIC (6 μg/ml) against A. baumannii expressing ADC-1 (Table 1).
More recently, mupirocin-based hybrid compounds fused to holomycin, a dithiolopyrrolone antibiotic (Liras, 2014), have been reported (Johnson et al., 2024). Mupirocin is a pseudomonic acid antibiotic that inhibits isoleucine-tRNA ligase in bacteria, and is generally ineffective against the most common Gram-negative bacteria owing to its poor outer membrane penetration (Khoshnood et al., 2019). However, mupirocin–holomycin conjugates show enhanced antibacterial activity against E. coli compared with mupirocin alone (Johnson et al., 2024).
Although hybrid antibiotics and combination therapy have distinct pharmacokinetic and pharmacodynamic properties (Fig. 1), both have in common that they use two antibacterial moieties. Therefore, insights derived from combination therapy may aid in the development of effective hybrid antibiotics. In this context, we will explore combination therapy. The fractional inhibitory concentration index (FICI) is commonly used to measure the interactions between two antibiotics (Odds, 2003). FICI values of ≤ 0.5, 1, and ≥ 4 indicate synergy, no interaction, and antagonism, respectively (Odds, 2003). Although numerous antibiotic combinations with FICIs of ≤ 0.5 have been reported, the underlying molecular mechanisms responsible for their synergy have not often been elucidated. Here, we summarize recent studies that have revealed the molecular basis of synergistic interactions between antibiotic combinations (Table 2).
Combination therapy with trimethoprim and sulfamethoxazole, also known as cotrimoxazole, is widely used to treat various bacterial and certain fungal infections (Smilack, 1999). Surprisingly, the molecular basis of their synergistic effect has been clarified only recently (Minato et al., 2018). The two antibiotics inhibit sequential steps in tetrahydrofolate biosynthesis: trimethoprim blocks the conversion of dihydrofolate to tetrahydrofolate, whereas sulfamethoxazole inhibits dihydropteroate production from p-aminobenzoic acid and 6-hydroxymethyl-7,8-dihydropterin pyrophosphate (Minato et al., 2018). Generally, the potent synergy between the two antibiotics is believed to be caused by the sequential inhibition of adjacent biosynthetic steps, despite limited experimental evidence. A recent report showed that other antibiotic combinations inhibiting sequential steps in tetrahydrofolate biosynthesis do not demonstrate a synergistic effect; only a mixture of trimethoprim and sulfamethoxazole showed this synergy (FICI = 0.31) (Minato et al., 2018). Mechanically, sulfamethoxazole potentiates trimethoprim by limiting de novo dihydrofolate production, while trimethoprim enhances sulfamethoxazole activity by inhibiting dihydropterin pyrophosphate synthesis (Minato et al., 2018). These findings indicate that the synergy between the two antibiotics may be derived from delicate physiological mechanisms.
Nitrofurantoin is a nitrofuran antibiotic commonly used to treat uncomplicated urinary tract infections (Mahdizade Ari et al., 2023). It shows strong synergistic interactions with amikacin against E. coli (FICI = 0.375) (Ren et al., 2023). Amikacin is an aminoglycoside antibiotic that induces mistranslation of messenger RNA, leading to the accumulation of misfolded proteins (Mingeot-Leclercq et al., 1999). These mistranslated proteins cause bacterial envelope stress, constitutively activating the CpxAR two‐component system. Activation of Cpx signaling stimulates the expression of NfsAB, a major bacterial nitroreductase, through the SoxS/MarA regulons. Nitroreductases overexpression generates lethal reactive intermediates via nitroreduction and promotes prodrug activation of nitrofurantoin (Ren et al., 2023). These results show that stress-induced changes in gene expression can enhance the antibacterial activity of specific antibiotics.
Antibiotics uptake and efflux across bacterial membranes play a pivotal role in antibacterial efficacy, particularly in Gram-negative bacteria that block penetration of high-molecular weight antibiotics through their outer membrane (O'Shea and Moser, 2008). Several studies have reported that antibiotic synergy is mediated by augmented penetration of antibiotics (Table 2). For example, the nalidixic acid–tetracycline combination shows strong synergy against both multidrug-resistant E. coli (FICI = 0.25–0.5) and multidrug-resistant A. baumannii (FICI = 0.1875–0.5) (Gaurav et al., 2021). This synergy is mediated by the enhanced uptake and reduced efflux of tetracycline by nalidixic acid (Gaurav et al., 2021). Notably, the nalidixic acid–tetracycline combination did not show synergy against antibiotic-susceptible A. baumannii and E. coli isolates (Gaurav et al., 2021).
Flavomycin is a widely used antibiotic feed additive in the livestock industry that inhibits bacterial cell wall synthesis (Ostash et al., 2010). Colistin, also known as polymyxin E, is a polycationic cyclic polypeptide antibiotic that binds to lipopolysaccharides and phospholipids in the outer cell membrane of Gram-negative bacteria. This leads to disruption of the outer cell membrane (Li et al., 2006). The flavomycin–colistin combination shows strong synergy against both E. coli (FICI = 0.19–0.49) and mcr-1-positive E. coli (FICI = 0.023–0.375) (Huang et al., 2024). This synergy manifests as augmented penetration of flavomycin by colistin, leading to increased intracellular accumulation of flavomycin and improved bacterial killing (Huang et al., 2024).
Similarly, the novobiocin–colistin combination exhibits strong synergy against colistin-resistant A. baumannii (FICI = 0.129) and K. pneumoniae (FICI = 0.012) (May et al., 2017). Although this synergy is initially believed to result from enhanced novobiocin penetration by colistin, due to high molecular weight of novobiocin (612 Da), similar to flavomycin (1583 Da), subsequent studies have shown that it instead results from novobiocin-mediated potentiation of colistin activity (Mandler et al., 2018; Mattingly et al., 2020; May et al., 2017). Notably, novobiocin, an aminocoumarin antibiotic that inhibits bacterial DNA gyrase (Lawson and Stevenson, 2012), also binds and activates the ATPase LptB that powers lipopolysaccharide transport, which enhances polymyxin activity (Mandler et al., 2018; May et al., 2017). Novobiocin derivatives were synthesized to separate DNA gyrase inhibition from LptB stimulation. Although one of the derivatives retained only LptB-stimulatory activity, it continued to demonstrate strong synergy with polymyxin by binding to LptB and stimulating lipopolysaccharide transport (Mandler et al., 2018).
More recently, the synergistic mechanism underlying hygromycin A–macrolides combinations was revealed through structural and biochemical experiments (Chen et al., 2023). Hygromycin A, also known as totomycin, is a modified cinnamic acid antibiotic containing a furanose sugar and aminocyclitol moiety. It prevents the formation of new peptide bonds by targeting the catalytic peptidyl transferase center (PTC) of the 50S large ribosomal subunit, and selectively acts against Borrelia burgdorferi, the causative agent of Lyme disease (Leimer et al., 2021). Macrolides are a class of natural antibiotics with a large macrocyclic lactone ring attached to deoxy sugars (Zhanel et al., 2001). These antibiotics inhibit bacterial ribosomal translation by targeting the nascent peptide exit tunnel (NPET) of the 50S large ribosomal subunit (Vazquez-Laslop and Mankin, 2018). The hygromycin A–macrolides combination shows moderate synergy (FICI = 0.5) against B. burgdorferi (Table 2). Among the various PTC-targeting antibiotics, such as hygromycin A, clindamycin, and linezolid, only hygromycin A can simultaneously bind the 70S ribosome with NPET-targeting macrolides without a steric clash. Hygromycin binding slows the dissociation of macrolides from the ribosome, which potentiates the efficacy of macrolides in bacterial growth inhibition and early killing (Chen et al., 2023). Collectively, these reports demonstrate that antibiotic synergy is mediated by various complex and highly specific mechanisms.
Owing to the challenges associated with the development of novel antibiotics, alternative treatment strategies using currently available antibiotics have been extensively studied. The development of dual-acting hybrid antibiotics has been a long-standing research direction for improving the antibacterial efficacy of existing antibiotics. Among these hybrid compounds, fluoroquinolone–oxazolidinone, fluoroquinolone–rifampicin, and rifamycin–nitroimidazole hybrids have entered clinical trials (Table 1) (Koh et al., 2023). However, as most antibiotics do not have outer membrane-destabilizing activity exhibited by colistin, an increase of the molecular weight by covalent fusion of two antibiotics can block outer membrane penetration in Gram-negative bacteria. Consequently, most hybrid compounds currently in clinical trials have been tested primarily against Gram-positive bacteria. Only rifamycin–nitroimidazole hybrids are currently in Phase II clinical trials for the treatment of H. pylori infections (Koh et al., 2023). Therefore, combining antibiotics that exhibit strong synergy may provide some assistance in overcoming the unavoidable disadvantage of increased molecular weight. Unfortunately, the molecular mechanisms underlying the synergistic effects between currently available antibiotics remain poorly understood. Consequently, further efforts are required to elucidate these mechanisms. Although synergy between two antibiotics does not inherently compensate for impermeability or pharmacokinetic constraints caused by increased molecular weight, the pharmacodynamic effects conferred by synergy can positively affect the efficacy of hybrid antibiotics. Therefore, the elaborate design of dual-acting hybrid antibiotics informed by well-defined synergistic mechanisms can substantially enhance the successful development of hybrid antibiotics for the treatment of infections caused by Gram-negative bacteria. This perspective can be one of useful strategies for achieving the development of effective hybrid antibiotics. As physicochemical factors, including polarity, charge distribution, conformational flexibility, amphiphilicity, and susceptibility to efflux, are also important for uptake of hybrid antibiotics, modification of physicochemical properties or combination therapy with membrane-active antimicrobial adjuvant to enhance the efficacy of hybrid antibiotics is also one of useful strategies for developing effective hybrid antibiotics.
Fig. 1.
Comparison of hybrid antibiotics with combination therapy. Both strategies can broaden the spectrum of action, reduce antibiotic resistance, and induce synergistic effects. (A) Hybrid antibiotics containing a metabolically non-cleavable covalent linker demonstrate singular pharmacokinetic and pharmacodynamic properties. A representative example of hybrid antibiotics is shown. (B) Combination therapy demonstrates independent, and potentially unpredictable, pharmacokinetic and pharmacodynamic properties. A representative example of combination therapy is shown.
jm-2601004f1.jpg
Table 1.
Antibiotic hybrids against Gram-negative pathogens
Hybrid antibiotics Target pathogens MIC (mg/ml) MIC in the resistant mutant strains Development stage Comments Reference
Ciprofloxacin–cephalosporin E. coli 0.18 NDM-expressing E. coli: 0.05 In vitro bacteria assay Evans et al. (2019)
Ciprofloxacin–neomycin E. coli 0.75–6 Neomycin-resistant E. coli: 6 In vitro bacteria assay Pokrovskaya et al. (2009)
Ciprofloxacin–kanamycin E. coli 0.28–12 Kanamycin-resistant E. coli: 0.1–3.0 In vitro bacteria assay The hybrids had significantly lower resistance frequencies in E. coli than each drug alone or their combination mixture. Shavit et al. (2017)
Ciprofloxacin–tobramycin P. aeruginosa 4 ND In vitro bacteria assay The compound showed strong synergy with fluoroquinolone against multidrug-resistant P. aeruginosa, A. baumannii, and K. pneumoniae. Gorityala et al. (2016)
Ciprofloxacin–tobramycin P. aeruginosa 64 ND In vitro bacteria assay The compound showed strong synergy with ciprofloxacin, levofloxacin, and moxifloxacin against multidrug-resistant P. aeruginosa. Dhiman et al. (2023)
Ciprofloxacin–erythromycin H. influenzae 4 ND In vitro bacteria assay Hutinec et al. (2010)
Ciprofloxacin–erythromycin H. influenzae, M. catarrhalis H. influenzae: 2 ND In vitro bacteria assay Ma et al. (2019)
M. catarrhalis: 0.5
Quinolone–telithromycin H. influenzae, E. coli, K. pneumoniae H. influenzae: 2 ND In vitro bacteria assay Fan et al. (2020)
E. coli: 8
K. pneumoniae: 16
Quinolone–telithromycin H. influenzae, M. pneumoniae H. influenzae: 4 M. pneumoniae: 0.008 Erythromycin-resistant M. pneumoniae: 8 In vitro bacteria assay Liu et al. (2022)
Ciprofloxacin–oxazolidinone H. influenzae, E. coli H. influenzae: 0.06 ND In vitro bacteria assay Hubschwerlen et al. (2003a)
E. coli: 1
Ciprofloxacin–oxazolidinone H. influenzae, E. coli H. influenzae: 0.03 ND In vitro bacteria assay Hubschwerlen et al. (2003b)
E. coli: 0.5
Fluoroquinolone–oxazolidinone H. influenzae, E. coli, P. aeruginosa H. influenzae: 0.016 ND In vitro bacteria assay Liu et al. (2019)
E. coli: 1
P. aeruginosa: 16
Fluoroquinolone–oxazolidinone (cadazolid) E. coli E. coli: 8 ND Discontinued in 2018 after two Phase III trials Scaiola et al. (2019)
Fluoroquinolone–rifampicin H. influenzae, E. coli, N. gonorrhoeae, H. pylori H. influenzae: 0.012 ND Phase II for acute bacterial skin and skin structure infection Yuan et al. (2020)
E. coli: 0.25
N. gonorrhoeae: 0.06
H. pylori: 0.12
Fluoroquinolone–kanglemycin E. coli 4 ND In vitro bacteria assay Peek et al. (2022)
Ciprofloxacin–sulfonamide E. coli 0.013 ND In vitro bacteria assay Ibrahim et al. (2022)
Quinolone–trimethoprim P. aeruginosa, K. pneumoniae P. aeruginosa: 8–16 ND In vitro bacteria assay Karoli et al. (2012)
K. pneumoniae: 8
Quinolone–metronidazole P. aeruginosa, E. coli P. aeruginosa: 2 ND In vitro bacteria assay Several compounds showed appropriate ranges to pharmacokinetic behaviors and no obvious toxicity to human hepatocyte cells. Cui et al. (2014)
E. coli: 0.5
Quinolone–metronidazole P. aeruginosa, E. coli P. aeruginosa: 0.25 ND In vitro bacteria assay Zhang et al. (2015)
E. coli: 0.25
Quinolone–fluoroquinolone P. aeruginosa 312.5 ND In vitro bacteria assay Panda et al. (2015)
Ciprofloxacin dimer E. coli, P. aeruginosa P. aeruginosa: 0.125 ND In vitro bacteria assay Ross et al. (2015)
E. coli: 0.03
Polymyxin B3–tobramycin- E. coli, K. pneumoniae, A. baumannii, P. aeruginosa P. aeruginosa: 2 Carbapenem-resistant MDR/XDR P. aeruginosa: 2–16 In vitro bacteria assay The compound showed strong synergy with minocycline, rifampicin, and vancomycin against multidrug-resistant P. aeruginosa. Domalaon et al. (2017)
A. baumannii: 16 Colistin-resistant P. aeruginosa: 4–32
E. coli: 8
K. pneumoniae: 128
Polymyxin E–vancomycin (vancomyxin) K. pneumoniae, A. baumannii, P. aeruginosa K. pneumoniae: 8 ND In vitro bacteria assay van Groesen et al. (2021)
P. aeruginosa: 16
A. baumannii: 16
Vancomycin–nisin K. pneumoniae, M. catarrhalis K. pneumoniae: 16 ND In vitro bacteria assay Arnusch et al. (2008)
M. catarrhalis: 16
Rifamycin–nitroimidazole (TNP-2198) H. pylori H. pylori: 0.004 Rifamycin-resistant H. pylori: 0.5 Phase II for H. pylori, vaginosis and CDAD ongoing since 2021/2022 Ma et al. (2022)
Rifamycin- and metronidazole-resistant H. pylori: 0.5
Neomycin B–triclosan E. coli, K. pneumoniae, A. baumannii, P. aeruginosa E. coli: 0.25 ND In vitro bacteria assay Findlay et al. (2012)
K. pneumoniae: 1
P. aeruginosa: 64
A. baumannii: 8
Sisomicin–gentamicin E. coli, K. pneumoniae, P. aeruginosa E. coli: 0.5 ND In vitro bacteria assay Hanessian et al. (2011)
K. pneumoniae: 0.25
P. aeruginosa: 0.5
Linezolid–sparsomycin H. influenzae 4 ND In vitro bacteria assay Zhou et al. (2008)
Oxazolidinone–cephalosporin–siderophore E. coli, A. baumannii, P. aeruginosa E. coli: 0.025 ADC-1-expressing A. baumannii: 6 In vitro bacteria assay Liu et al. (2018)
A. baumannii: 0.4
P. aeruginosa: 0.4
Mupirocin–holomycin E. coli 64 ND In vitro bacteria assay Johnson et al. (2024)

bNot determined.

Table 2.
Combination therapies against Gram-negative pathogens
Combination antibiotics Target pathogens FICI Synergy mechanisms Clinical uses Comments Reference
Trimethoprim and sulfamethoxazole E. coli 0.31 Mutual potentiation (Sulfamethoxazole potentiates trimethoprim by limiting de novo dihydrofolate production and trimethoprim potentiates sulfamethoxazole activity through inhibition of dihydropterin pyrophosphate synthesis) Clinical use Minato et al. (2018)
Amikacin and nitrofurantoin E. coli, K. pneumoniae E. coli: 0.375 Amikacin induces bacterial envelope stress by introducing mistranslated proteins, thereby constitutively activating the CpxAR two‐component system. The activation of Cpx signaling stimulates the expression of bacterial major nitroreductases (NfsAB). Nitroreductases overexpression generates considerable quantity of lethal reactive intermediates via nitroreduction and promotes the prodrug activation of nitrofurantoin. No Ren et al. (2023)
K. pneumoniae: 0.5
Nalidixic acid and tetracycline Multidrug-resistant A. baumannii and E. coli Multidrug-resistant A. baumannii: 0.1875–0.5 Enhanced uptake and reduced efflux of tetracycline by nalidixic acid explain the basis of synergy between nalidixic acid and tetracycline. No Nalidixic acid and tetracycline combination did not display synergy against susceptible A. baumannii and E. coli isolates. Gaurav et al. (2021)
Flavomycin and colistin E. coli, mcr-1-positive E. coli E. coli: 0.19–0.49 The synergy is manifested as an augmented penetration of the E. coli OM by colistin, leading to increased intracellular accumulation of flavomycin and enhanced cell killing thereafter. No Huang et al. (2024)
Novobiocin and colistin Colistin-resistant A. baumannii and K. pneumoniae Colistin-resistant A. baumannii: 0.129 Novobiocin binds and activates the ATPase LptB that powers lipopolysaccharide transport, which enhances polymyxin activity. No May et al. (2017); Mandler et al. (2018); Mattingly et al. (2020)
Colistin-resistant K. pneumoniae: 0.012
Hygromycin A and macrolides B. burgdorferi 0.5 Hygromycin A cooperatively binds ribosomes with nascent peptide exit tunnel-targeting macrolides and slows down their dissociation, which potentiates macrolide’s efficacy in bacterial growth inhibition and early killing. No Chen et al. (2023)

bNot determined.

  • Antimicrobial Resistance Collaborators. 2022. Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. Lancet. 399: 629–655. ArticlePubMedPMC
  • Arnusch CJ, Bonvin AM, Verel AM, Jansen WT, Liskamp RM, et al. 2008. The vancomycin-nisin(1–12) hybrid restores activity against vancomycin resistant Enterococci. Biochemistry. 47: 12661–12663. ArticlePubMed
  • Bozdogan B, Appelbaum PC. 2004. Oxazolidinones: activity, mode of action, and mechanism of resistance. Int J Antimicrob Agents. 23: 113–119. ArticlePubMed
  • Brogden RN, Carmine AA, Heel RC, Speight TM, Avery GS. 1982. Trimethoprim: a review of its antibacterial activity, pharmacokinetics and therapeutic use in urinary tract infections. Drugs. 23: 405–430. ArticlePubMed
  • Bulitta JB, Ly NS, Landersdorfer CB, Wanigaratne NA, Velkov T, et al. 2015. Two mechanisms of killing of Pseudomonas aeruginosa by tobramycin assessed at multiple inocula via mechanism-based modeling. Antimicrob Agents Chemother. 59: 2315–2327. ArticlePubMedPMCLink
  • Butler MS, Blaskovich MA, Cooper MA. 2017. Antibiotics in the clinical pipeline at the end of 2015. J Antibiot (Tokyo). 70: 3–24. ArticlePubMedPDF
  • Calvori C, Frontali L, Leoni L, Tecce G. 1965. Effect of rifamycin on protein synthesis. Nature. 207: 417–418. ArticlePubMedPDF
  • Chen CW, Leimer N, Syroegin EA, Dunand C, Bulman ZP, et al. 2023. Structural insights into the mechanism of overcoming Erm-mediated resistance by macrolides acting together with hygromycin-A. Nat Commun. 14: 4196.ArticlePubMedPMCPDF
  • Cui SF, Peng LP, Zhang HZ, Rasheed S, Vijaya Kumar K, et al. 2014. Novel hybrids of metronidazole and quinolones: synthesis, bioactive evaluation, cytotoxicity, preliminary antimicrobial mechanism and effect of metal ions on their transportation by human serum albumin. Eur J Med Chem. 86: 318–334. ArticlePubMed
  • Dhiman S, Ramirez D, Li Y, Kumar A, Arthur G, et al. 2023. Chimeric tobramycin-based adjuvant TOB-TOB-CIP potentiates fluoroquinolone and β-lactam antibiotics against multidrug-resistant Pseudomonas aeruginosa. ACS Infect Dis. 9: 864–885. ArticlePubMedLink
  • Domalaon R, Yang X, Lyu Y, Zhanel GG, Schweizer F. 2017. Polymyxin B3-tobramycin hybrids with Pseudomonas aeruginosa-selective antibacterial activity and strong potentiation of rifampicin, minocycline, and vancomycin. ACS Infect Dis. 3: 941–954. ArticlePubMed
  • Evans LE, Krishna A, Ma Y, Webb TE, Marshall DC, et al. 2019. Exploitation of antibiotic resistance as a novel drug target: development of a β-lactamase-activated antibacterial prodrug. J Med Chem. 62: 4411–4425. ArticlePubMedPMC
  • Fan BZ, Hiasa H, Lv W, Brody S, Yang ZY, et al. 2020. Design, synthesis and structure-activity relationships of novel 15-membered macrolides: Quinolone/quinoline-containing sidechains tethered to the C-6 position of azithromycin acylides. Eur J Med Chem. 193: 112222.ArticlePubMed
  • Findlay B, Zhanel GG, Schweizer F. 2012. Neomycin-phenolic conjugates: polycationic amphiphiles with broad-spectrum antibacterial activity, low hemolytic activity and weak serum protein binding. Bioorg Med Chem Lett. 22: 1499–1503. ArticlePubMed
  • Gaurav A, Gupta V, Shrivastava SK, Pathania R. 2021. Mechanistic insights into synergy between nalidixic acid and tetracycline against clinical isolates of Acinetobacter baumannii and Escherichia coli. Commun Biol. 4: 542.ArticlePubMedPMCPDF
  • Gorityala BK, Guchhait G, Fernando DM, Deo S, McKenna SA, et al. 2016. Adjuvants based on hybrid antibiotics overcome resistance in Pseudomonas aeruginosa and enhance fluoroquinolone efficacy. Angew Chem Int Ed Engl. 55: 555–559. ArticlePubMedLink
  • Hanessian S, Maianti JP, Matias RD, Feeney LA, Armstrong ES. 2011. Hybrid aminoglycoside antibiotics via Tsuji palladium-catalyzed allylic deoxygenation. Org Lett. 13: 6476–6479. ArticlePubMed
  • Herzog IM, Green KD, Berkov-Zrihen Y, Feldman M, Vidavski RR, et al. 2012. 6''-Thioether tobramycin analogues: towards selective targeting of bacterial membranes. Angew Chem Int Ed Engl. 51: 5652–5656. ArticlePubMedPMC
  • Huang Y, Zhu Y, Yue HY, Liu YY, Deng LM, et al. 2024. Flavomycin restores colistin susceptibility in multidrug-resistant Gram-negative bacteria. mSystems. 9: e00109-24.ArticlePubMedPMCLink
  • Hubschwerlen C, Specklin JL, Baeschlin DK, Borer Y, Haefeli S, et al. 2003a. Structure-activity relationship in the oxazolidinone-quinolone hybrid series: influence of the central spacer on the antibacterial activity and the mode of action. Bioorg Med Chem Lett. 13: 4229–4233. Article
  • Hubschwerlen C, Specklin JL, Sigwalt C, Schroeder S, Locher HH. 2003b. Design, synthesis and biological evaluation of oxazolidinone-quinolone hybrids. Bioorg Med Chem. 11: 2313–2319. Article
  • Hutinec A, Derek M, Lazarevski G, Sunjic V, Paljetak HC, et al. 2010. Novel 8a-aza-8a-homoerythromycin—4''-(3-substituted-amino)propionates with broad spectrum antibacterial activity. Bioorg Med Chem Lett. 20: 3244–3249. ArticlePubMed
  • Ibrahim NM, Fahim SH, Hassan M, Farag AE, Georgey HH. 2022. Design and synthesis of ciprofloxacin-sulfonamide hybrids to manipulate ciprofloxacin pharmacological qualities: Potency and side effects. Eur J Med Chem. 228: 114021.ArticlePubMed
  • Johnson RM, Li K, Chen X, Morgan GL, Aube J, et al. 2024. The hybrid antibiotic thiomarinol A overcomes intrinsic resistance in Escherichia coli using a privileged dithiolopyrrolone moiety. ACS Infect Dis. 10: 582–593. ArticlePubMedPMCLink
  • Karoli T, Mamidyala SK, Zuegg J, Fry SR, Tee EH, et al. 2012. Structure aided design of chimeric antibiotics. Bioorg Med Chem Lett. 22: 2428–2433. ArticlePubMed
  • Khoshnood S, Heidary M, Asadi A, Soleimani S, Motahar M, et al. 2019. A review on mechanism of action, resistance, synergism, and clinical implications of mupirocin against Staphylococcus aureus. Biomed Pharmacother. 109: 1809–1818. ArticlePubMed
  • Koh AJJ, Thombare V, Hussein M, Rao GG, Li J, et al. 2023. Bifunctional antibiotic hybrids: A review of clinical candidates. Front Pharmacol. 14: 1158152.ArticlePubMedPMC
  • Kratky M, Vinsova J, Volkova M, Buchta V, Trejtnar F, et al. 2012. Antimicrobial activity of sulfonamides containing 5-chloro-2-hydroxybenzaldehyde and 5-chloro-2-hydroxybenzoic acid scaffold. Eur J Med Chem. 50: 433–440. ArticlePubMed
  • Lawson DM, Stevenson CE. 2012. Structural and functional dissection of aminocoumarin antibiotic biosynthesis: a review. J Struct Funct Genomics. 13: 125–133. ArticlePubMedPDF
  • Lee CR, Cho IH, Jeong BC, Lee SH. 2013. Strategies to minimize antibiotic resistance. Int J Environ Res Public Health. 10: 4274–4305. ArticlePubMedPMC
  • Leimer N, Wu X, Imai Y, Morrissette M, Pitt N, et al. 2021. A selective antibiotic for Lyme disease. Cell. 184: 5405–5418.e16. ArticlePubMedPMC
  • Leitsch D. 2019. A review on metronidazole: an old warhorse in antimicrobial chemotherapy. Parasitology. 146: 1167–1178. ArticlePubMed
  • Li J, Nation RL, Turnidge JD, Milne RW, Coulthard K, et al. 2006. Colistin: the re-emerging antibiotic for multidrug-resistant Gram-negative bacterial infections. Lancet Infect Dis. 6: 589–601. ArticlePubMed
  • Liras P. 2014. Holomycin, a dithiolopyrrolone compound produced by Streptomyces clavuligerus. Appl Microbiol Biotechnol. 98: 1023–1030. ArticlePubMedPDF
  • Liu XP, Lv W, Zhao F, Ding J, Zhang JR, et al. 2022. Design and synthesis of novel macrolones bridged with linkers from 11,12-positions of macrolides. Bioorg Med Chem Lett. 68: 128761.ArticlePubMed
  • Liu R, Miller PA, Vakulenko SB, Stewart NK, Boggess WC, et al. 2018. A synthetic dual drug sideromycin induces Gram-negative bacteria to commit suicide with a Gram-positive antibiotic. J Med Chem. 61: 3845–3854. ArticlePubMed
  • Liu L, Shao L, Li J, Cui H, Li B, et al. 2019. Synthesis, antibacterial activities, mode of action and acute toxicity studies of new oxazolidinone-fluoroquinolone hybrids. Molecules. 24: 1641.ArticlePubMedPMC
  • Ma Z, He S, Yuan Y, Zhuang Z, Liu Y, et al. 2022. Design, synthesis, and characterization of TNP-2198, a dual-targeted rifamycin-nitroimidazole conjugate with potent activity against microaerophilic and anaerobic bacterial pathogens. J Med Chem. 65: 4481–4495. ArticlePubMedPMC
  • Ma CX, Lv W, Li YX, Fan BZ, Han X, et al. 2019. Design, synthesis and structure-activity relationships of novel macrolones: hybrids of 2-fluoro 9-oxime ketolides and carbamoyl quinolones with highly improved activity against resistant pathogens. Eur J Med Chem. 169: 1–20. ArticlePubMed
  • Mahdizade Ari M, Dashtbin S, Ghasemi F, Shahroodian S, Kiani P, et al. 2023. Nitrofurantoin: properties and potential in treatment of urinary tract infection: a narrative review. Front Cell Infect Microbiol. 13: 1148603.ArticlePubMedPMC
  • Mandler MD, Baidin V, Lee J, Pahil KS, Owens TW, et al. 2018. Novobiocin enhances polymyxin activity by stimulating lipopolysaccharide transport. J Am Chem Soc. 140: 6749–6753. ArticlePubMedPMC
  • Mattingly AE, Cox KE, Smith R, Melander RJ, Ernst RK, et al. 2020. Screening an established natural product library identifies secondary metabolites that potentiate conventional antibiotics. ACS Infect Dis. 6: 2629–2640. ArticlePubMedPMC
  • May JM, Owens TW, Mandler MD, Simpson BW, Lazarus MB, et al. 2017. The antibiotic novobiocin binds and activates the ATPase that powers lipopolysaccharide transport. J Am Chem Soc. 139: 17221–17224. ArticlePubMedPMC
  • Minato Y, Dawadi S, Kordus SL, Sivanandam A, Aldrich CC, et al. 2018. Mutual potentiation drives synergy between trimethoprim and sulfamethoxazole. Nat Commun. 9: 1003.ArticlePubMedPMCPDF
  • Mingeot-Leclercq MP, Glupczynski Y, Tulkens PM. 1999. Aminoglycosides: activity and resistance. Antimicrob Agents Chemother. 43: 727–737. ArticlePubMedPMCLink
  • Odds FC. 2003. Synergy, antagonism, and what the chequerboard puts between them. J Antimicrob Chemother. 52: 1.ArticlePubMed
  • O'Shea R, Moser HE. 2008. Physicochemical properties of antibacterial compounds: implications for drug discovery. J Med Chem. 51: 2871–2878. ArticlePubMed
  • Ostash B, Doud E, Fedorenko V. 2010. The molecular biology of moenomycins: towards novel antibiotics based on inhibition of bacterial peptidoglycan glycosyltransferases. Biol Chem. 391: 499–504. ArticlePubMed
  • Ottenheijm HC, van den Broek LA, Ballesta JP, Zylicz Z. 1986. Chemical and biological aspects of sparsomycin, an antibiotic from Streptomyces. Prog Med Chem. 23: 219–268. ArticlePubMed
  • Panda SS, Liaqat S, Girgis AS, Samir A, Hall CD, et al. 2015. Novel antibacterial active quinolone-fluoroquinolone conjugates and 2D-QSAR studies. Bioorg Med Chem Lett. 25: 3816–3821. ArticlePubMed
  • Peek J, Koirala B, Brady SF. 2022. Synthesis and evaluation of dual-action kanglemycin-fluoroquinolone hybrid antibiotics. Bioorg Med Chem Lett. 57: 128484.ArticlePubMed
  • Pokrovskaya V, Baasov T. 2010. Dual-acting hybrid antibiotics: a promising strategy to combat bacterial resistance. Expert Opin Drug Discov. 5: 883–902. ArticlePubMed
  • Pokrovskaya V, Belakhov V, Hainrichson M, Yaron S, Baasov T. 2009. Design, synthesis, and evaluation of novel fluoroquinolone-aminoglycoside hybrid antibiotics. J Med Chem. 52: 2243–2254. ArticlePubMed
  • Ren H, Zhong Z, Zhou S, Wei Y, Liang Y, et al. 2023. CpxA/R-controlled nitroreductase expression as target for combinatorial therapy against uropathogens by promoting reactive oxygen species generation. Adv Sci (Weinh). 10: 2300938. ArticlePubMedPMC
  • Ross AG, Benton BM, Chin D, De Pascale G, Fuller J, et al. 2015. Synthesis of ciprofloxacin dimers for evaluation of bacterial permeability in atypical chemical space. Bioorg Med Chem Lett. 25: 3468–3475. ArticlePubMed
  • Scaiola A, Leibundgut M, Boehringer D, Caspers P, Bur D, et al. 2019. Structural basis of translation inhibition by cadazolid, a novel quinoxolidinone antibiotic. Sci Rep. 9: 5634.ArticlePubMedPMCPDF
  • Schalk IJ. 2025. Bacterial siderophores: diversity, uptake pathways and applications. Nat Rev Microbiol. 23: 24–40. ArticlePubMedPDF
  • Schumacher A, Trittler R, Bohnert JA, Kummerer K, Pages JM, et al. 2007. Intracellular accumulation of linezolid in Escherichia coli, Citrobacter freundii and Enterobacter aerogenes: role of enhanced efflux pump activity and inactivation. J Antimicrob Chemother. 59: 1261–1264. ArticlePubMed
  • Shavit M, Pokrovskaya V, Belakhov V, Baasov T. 2017. Covalently linked kanamycin-ciprofloxacin hybrid antibiotics as a tool to fight bacterial resistance. Bioorg Med Chem. 25: 2917–2925. ArticlePubMed
  • Smilack JD. 1999. Trimethoprim-sulfamethoxazole. Mayo Clin Proc. 74: 730–734. ArticlePubMed
  • Theuretzbacher U. 2020. Dual-mechanism antibiotics. Nat Microbiol. 5: 984–985. ArticlePubMedPDF
  • Thomas CM, Nielsen KM. 2005. Mechanisms of, and barriers to, horizontal gene transfer between bacteria. Nat Rev Microbiol. 3: 711–721. ArticlePubMedPDF
  • van Groesen E, Slingerland CJ, Innocenti P, Mihajlovic M, Masereeuw R, et al. 2021. Vancomyxins: vancomycin-polymyxin nonapeptide conjugates that retain anti-Gram-positive activity with enhanced potency against Gram-negative strains. ACS Infect Dis. 7: 2746–2754. ArticlePubMedPMCLink
  • Vazquez-Laslop N, Mankin AS. 2018. How macrolide antibiotics work. Trends Biochem Sci. 43: 668–684. ArticlePubMedPMC
  • Velkov T, Roberts KD, Nation RL, Thompson PE, Li J. 2013. Pharmacology of polymyxins: new insights into an 'old' class of antibiotics. Future Microbiol. 8: 711–724. ArticlePubMedPMC
  • Yuan Y, Wang X, Xu X, Liu Y, Li C, et al. 2020. Evaluation of a dual-acting antibacterial agent, TNP-2092, on gut microbiota and potential application in the treatment of gastrointestinal and liver disorders. ACS Infect Dis. 6: 820–831. ArticlePubMed
  • Zhanel GG, Dueck M, Hoban DJ, Vercaigne LM, Embil JM, et al. 2001. Review of macrolides and ketolides: focus on respiratory tract infections. Drugs. 61: 443–498. ArticlePubMed
  • Zhang L, Kumar KV, Geng RX, Zhou CH. 2015. Design and biological evaluation of novel quinolone-based metronidazole derivatives as potent Cu2+ mediated DNA-targeting antibacterial agents. Bioorg Med Chem Lett. 25: 3699–3705. ArticlePubMed
  • Zhou J, Bhattacharjee A, Chen S, Chen Y, Duffy E, et al. 2008. Design at the atomic level: design of biaryloxazolidinones as potent orally active antibiotics. Bioorg Med Chem Lett. 18: 6175–6178. ArticlePubMed

Figure & Data

References

    Citations

    Citations to this article as recorded by  

      • ePub LinkePub Link
      • Cite this Article
        Cite this Article
        export Copy Download
        Close
        Download Citation
        Download a citation file in RIS format that can be imported by all major citation management software, including EndNote, ProCite, RefWorks, and Reference Manager.
        Format:
        • RIS — For EndNote, ProCite, RefWorks, and most other reference management software
        • BibTeX — For JabRef, BibDesk, and other BibTeX-specific software
        Include:
        • Citation for the content below
        Recent trends in dual-acting hybrid antibiotics and combination therapies against Gram-negative pathogens
        J. Microbiol. 2026;64(3):e2601004  Published online March 31, 2026
        DOI: https://doi.org/10.71150/jm.2601004
        Close
      • XML DownloadXML Download
      Figure
      Recent trends in dual-acting hybrid antibiotics and combination therapies against Gram-negative pathogens
      Image
      Fig. 1. Comparison of hybrid antibiotics with combination therapy. Both strategies can broaden the spectrum of action, reduce antibiotic resistance, and induce synergistic effects. (A) Hybrid antibiotics containing a metabolically non-cleavable covalent linker demonstrate singular pharmacokinetic and pharmacodynamic properties. A representative example of hybrid antibiotics is shown. (B) Combination therapy demonstrates independent, and potentially unpredictable, pharmacokinetic and pharmacodynamic properties. A representative example of combination therapy is shown.

      Fig. 1.

      Recent trends in dual-acting hybrid antibiotics and combination therapies against Gram-negative pathogens
      Hybrid antibiotics Target pathogens MIC (mg/ml) MIC in the resistant mutant strains Development stage Comments Reference
      Ciprofloxacin–cephalosporin E. coli 0.18 NDM-expressing E. coli: 0.05 In vitro bacteria assay Evans et al. (2019)
      Ciprofloxacin–neomycin E. coli 0.75–6 Neomycin-resistant E. coli: 6 In vitro bacteria assay Pokrovskaya et al. (2009)
      Ciprofloxacin–kanamycin E. coli 0.28–12 Kanamycin-resistant E. coli: 0.1–3.0 In vitro bacteria assay The hybrids had significantly lower resistance frequencies in E. coli than each drug alone or their combination mixture. Shavit et al. (2017)
      Ciprofloxacin–tobramycin P. aeruginosa 4 ND In vitro bacteria assay The compound showed strong synergy with fluoroquinolone against multidrug-resistant P. aeruginosa, A. baumannii, and K. pneumoniae. Gorityala et al. (2016)
      Ciprofloxacin–tobramycin P. aeruginosa 64 ND In vitro bacteria assay The compound showed strong synergy with ciprofloxacin, levofloxacin, and moxifloxacin against multidrug-resistant P. aeruginosa. Dhiman et al. (2023)
      Ciprofloxacin–erythromycin H. influenzae 4 ND In vitro bacteria assay Hutinec et al. (2010)
      Ciprofloxacin–erythromycin H. influenzae, M. catarrhalis H. influenzae: 2 ND In vitro bacteria assay Ma et al. (2019)
      M. catarrhalis: 0.5
      Quinolone–telithromycin H. influenzae, E. coli, K. pneumoniae H. influenzae: 2 ND In vitro bacteria assay Fan et al. (2020)
      E. coli: 8
      K. pneumoniae: 16
      Quinolone–telithromycin H. influenzae, M. pneumoniae H. influenzae: 4 M. pneumoniae: 0.008 Erythromycin-resistant M. pneumoniae: 8 In vitro bacteria assay Liu et al. (2022)
      Ciprofloxacin–oxazolidinone H. influenzae, E. coli H. influenzae: 0.06 ND In vitro bacteria assay Hubschwerlen et al. (2003a)
      E. coli: 1
      Ciprofloxacin–oxazolidinone H. influenzae, E. coli H. influenzae: 0.03 ND In vitro bacteria assay Hubschwerlen et al. (2003b)
      E. coli: 0.5
      Fluoroquinolone–oxazolidinone H. influenzae, E. coli, P. aeruginosa H. influenzae: 0.016 ND In vitro bacteria assay Liu et al. (2019)
      E. coli: 1
      P. aeruginosa: 16
      Fluoroquinolone–oxazolidinone (cadazolid) E. coli E. coli: 8 ND Discontinued in 2018 after two Phase III trials Scaiola et al. (2019)
      Fluoroquinolone–rifampicin H. influenzae, E. coli, N. gonorrhoeae, H. pylori H. influenzae: 0.012 ND Phase II for acute bacterial skin and skin structure infection Yuan et al. (2020)
      E. coli: 0.25
      N. gonorrhoeae: 0.06
      H. pylori: 0.12
      Fluoroquinolone–kanglemycin E. coli 4 ND In vitro bacteria assay Peek et al. (2022)
      Ciprofloxacin–sulfonamide E. coli 0.013 ND In vitro bacteria assay Ibrahim et al. (2022)
      Quinolone–trimethoprim P. aeruginosa, K. pneumoniae P. aeruginosa: 8–16 ND In vitro bacteria assay Karoli et al. (2012)
      K. pneumoniae: 8
      Quinolone–metronidazole P. aeruginosa, E. coli P. aeruginosa: 2 ND In vitro bacteria assay Several compounds showed appropriate ranges to pharmacokinetic behaviors and no obvious toxicity to human hepatocyte cells. Cui et al. (2014)
      E. coli: 0.5
      Quinolone–metronidazole P. aeruginosa, E. coli P. aeruginosa: 0.25 ND In vitro bacteria assay Zhang et al. (2015)
      E. coli: 0.25
      Quinolone–fluoroquinolone P. aeruginosa 312.5 ND In vitro bacteria assay Panda et al. (2015)
      Ciprofloxacin dimer E. coli, P. aeruginosa P. aeruginosa: 0.125 ND In vitro bacteria assay Ross et al. (2015)
      E. coli: 0.03
      Polymyxin B3–tobramycin- E. coli, K. pneumoniae, A. baumannii, P. aeruginosa P. aeruginosa: 2 Carbapenem-resistant MDR/XDR P. aeruginosa: 2–16 In vitro bacteria assay The compound showed strong synergy with minocycline, rifampicin, and vancomycin against multidrug-resistant P. aeruginosa. Domalaon et al. (2017)
      A. baumannii: 16 Colistin-resistant P. aeruginosa: 4–32
      E. coli: 8
      K. pneumoniae: 128
      Polymyxin E–vancomycin (vancomyxin) K. pneumoniae, A. baumannii, P. aeruginosa K. pneumoniae: 8 ND In vitro bacteria assay van Groesen et al. (2021)
      P. aeruginosa: 16
      A. baumannii: 16
      Vancomycin–nisin K. pneumoniae, M. catarrhalis K. pneumoniae: 16 ND In vitro bacteria assay Arnusch et al. (2008)
      M. catarrhalis: 16
      Rifamycin–nitroimidazole (TNP-2198) H. pylori H. pylori: 0.004 Rifamycin-resistant H. pylori: 0.5 Phase II for H. pylori, vaginosis and CDAD ongoing since 2021/2022 Ma et al. (2022)
      Rifamycin- and metronidazole-resistant H. pylori: 0.5
      Neomycin B–triclosan E. coli, K. pneumoniae, A. baumannii, P. aeruginosa E. coli: 0.25 ND In vitro bacteria assay Findlay et al. (2012)
      K. pneumoniae: 1
      P. aeruginosa: 64
      A. baumannii: 8
      Sisomicin–gentamicin E. coli, K. pneumoniae, P. aeruginosa E. coli: 0.5 ND In vitro bacteria assay Hanessian et al. (2011)
      K. pneumoniae: 0.25
      P. aeruginosa: 0.5
      Linezolid–sparsomycin H. influenzae 4 ND In vitro bacteria assay Zhou et al. (2008)
      Oxazolidinone–cephalosporin–siderophore E. coli, A. baumannii, P. aeruginosa E. coli: 0.025 ADC-1-expressing A. baumannii: 6 In vitro bacteria assay Liu et al. (2018)
      A. baumannii: 0.4
      P. aeruginosa: 0.4
      Mupirocin–holomycin E. coli 64 ND In vitro bacteria assay Johnson et al. (2024)
      Combination antibiotics Target pathogens FICI Synergy mechanisms Clinical uses Comments Reference
      Trimethoprim and sulfamethoxazole E. coli 0.31 Mutual potentiation (Sulfamethoxazole potentiates trimethoprim by limiting de novo dihydrofolate production and trimethoprim potentiates sulfamethoxazole activity through inhibition of dihydropterin pyrophosphate synthesis) Clinical use Minato et al. (2018)
      Amikacin and nitrofurantoin E. coli, K. pneumoniae E. coli: 0.375 Amikacin induces bacterial envelope stress by introducing mistranslated proteins, thereby constitutively activating the CpxAR two‐component system. The activation of Cpx signaling stimulates the expression of bacterial major nitroreductases (NfsAB). Nitroreductases overexpression generates considerable quantity of lethal reactive intermediates via nitroreduction and promotes the prodrug activation of nitrofurantoin. No Ren et al. (2023)
      K. pneumoniae: 0.5
      Nalidixic acid and tetracycline Multidrug-resistant A. baumannii and E. coli Multidrug-resistant A. baumannii: 0.1875–0.5 Enhanced uptake and reduced efflux of tetracycline by nalidixic acid explain the basis of synergy between nalidixic acid and tetracycline. No Nalidixic acid and tetracycline combination did not display synergy against susceptible A. baumannii and E. coli isolates. Gaurav et al. (2021)
      Flavomycin and colistin E. coli, mcr-1-positive E. coli E. coli: 0.19–0.49 The synergy is manifested as an augmented penetration of the E. coli OM by colistin, leading to increased intracellular accumulation of flavomycin and enhanced cell killing thereafter. No Huang et al. (2024)
      Novobiocin and colistin Colistin-resistant A. baumannii and K. pneumoniae Colistin-resistant A. baumannii: 0.129 Novobiocin binds and activates the ATPase LptB that powers lipopolysaccharide transport, which enhances polymyxin activity. No May et al. (2017); Mandler et al. (2018); Mattingly et al. (2020)
      Colistin-resistant K. pneumoniae: 0.012
      Hygromycin A and macrolides B. burgdorferi 0.5 Hygromycin A cooperatively binds ribosomes with nascent peptide exit tunnel-targeting macrolides and slows down their dissociation, which potentiates macrolide’s efficacy in bacterial growth inhibition and early killing. No Chen et al. (2023)
      Table 1. Antibiotic hybrids against Gram-negative pathogens

      Not determined.

      Table 2. Combination therapies against Gram-negative pathogens

      Not determined.

      Table 1.

      Table 2.

      Journal of Microbiology : Journal of Microbiology
      © The Microbiological Society of Korea.
      TOP