Antibiotic Hybrids: the Next Generation of Agents and Adjuvants against Gram-Negative Pathogens?

Abstract

The global incidence of drug-resistant Gram-negative bacillary infections has been increasing, and there is a dire need to develop novel strategies to overcome this problem. Intrinsic resistance in Gram-negative bacteria, such as their protective outer membrane and constitutively overexpressed efflux pumps, is a major survival weapon that renders them refractory to current antibiotics. Several potential avenues to overcome this problem have been at the heart of antibiotic drug discovery in the past few decades. We review some of these strategies, with emphasis on antibiotic hybrids either as stand-alone antibacterial agents or as adjuvants that potentiate a primary antibiotic in Gram-negative bacteria. Antibiotic hybrid is defined in this review as a synthetic construct of two or more pharmacophores belonging to an established agent known to elicit a desired antimicrobial effect. The concepts, advances, and challenges of antibiotic hybrids are elaborated in this article. Moreover, we discuss several antibiotic hybrids that were or are in clinical evaluation. Mechanistic insights into how tobramycin-based antibiotic hybrids are able to potentiate legacy antibiotics in multidrug-resistant Gram-negative bacilli are also highlighted. Antibiotic hybrids indeed have a promising future as a therapeutic strategy to overcome drug resistance in Gram-negative pathogens and/or expand the usefulness of our current antibiotic arsenal.

Keywords: antibacterial; antibiotic; antimicrobial; efflux; hybrid; permeability.

FIG 1

Dual membrane of Gram-negative bacteria. The periplasmic space that contains a thin peptidoglycan layer is enclosed by the outer membrane and the inner membrane. The asymmetric OM has an inner leaflet composed of phospholipids and an outer leaflet with abundant LPS. The LPS is sectioned mainly into hydrophobic lipid A (the structure is expanded in the dashed red box), the hydrophilic core oligosaccharides (blue hexagons), and the hydrophilic O antigen (black diamonds). However, there is some degree of variation among Gram-negative bacteria. The lipid A structure typically has two negatively charged phosphate groups stabilized by a divalent cation (such as Mg²⁺) bridge between adjacent lipid A phosphate groups that imparts structural stability to the OM. Both the inner and outer leaflets of the IM consist mainly of hydrophobic phospholipids. The difference in molecular compositions between the OM and IM results in their orthogonal sieving properties.

FIG 2

Two different pharmacophoric domains attached covalently by a linker domain. The lability of the linker determines the type of antibiotic hybrid generated. A linker that can be enzymatically degraded (preferably by only bacterium-specific enzymes) gives rise to two functional pharmacophoric entities that are thus used in the antibiotic hybrid prodrug strategy. A linker that is inert to enzymatic degradation is used to hold the two pharmacophoric domains together in the antibiotic hybrid drug strategy.

FIG 3

β-Lactam hydrolysis of cephalosporin followed by nonenzymatic release of a leaving group (LG). Enzymatic hydrolysis may be due to β-lactamases, such as serine- or metallo-based β-lactamases, or β-lactam target enzyme transpeptidases. Most hybrid prodrugs utilize this mechanism to release another antibiotic as a leaving group. R, any molecular substituent; E, enzyme.

FIG 4

β-Lactam hydrolysis of penicillin that contains an installed leaving group at position 6 via an S-aminosulfenimine (A) or a vinyl ester (B) linkage. The mechanisms for the nonenzymatically driven release of the leaving group for panels A and B were elucidated thorough NMR experiments (185, 186). Enzymatic hydrolysis may be due to β-lactamases, such as serine or metallo-based β-lactamases (not depicted above), or the β-lactam target enzyme transpeptidase.

FIG 5

Examples of antibiotic hybrid prodrugs: a cefamandole derivative linked to omadine (hybrid 1), desacetylcephalothin linked to the alanine racemase inhibitor chloroalanyl dipeptide (hybrid 2), desacetylcephalothin linked to triclosan-NB2001 (hybrid 3), and desacetylcefotaxime linked to fleroxacin-Ro 23-9424 (hybrid 4). The majority of antibiotic hybrid prodrugs consist of a cephalosporin.

FIG 6

Electron transfer of hydrolyzed Ro 23-9424 results in the formation of hydrolyzed desacetylcefotaxime and fleroxacin by-products. Ro 23-9424 (hybrid 4) initially acts as a cephalosporin, followed by the release of a functional fluoroquinolone as a product of β-lactam hydrolysis. The highlighted red circle is the hydrolyzed β-lactam ring.

FIG 7

Examples of antibiotic hybrid drugs that are active against Gram-negative pathogens: trimethoprim linked to ciprofloxacin–BP-4Q-002 (hybrid 5), the TyrRS inhibitor 3-(2-fluorophenyl)furan-2(5H)-one linked to ciprofloxacin (hybrid 6), the flavonoid naringenin linked to ciprofloxacin (hybrid 7), neomycin B linked to ciprofloxacin via an aromatic triazole linker (hybrid 8), and neomycin B linked to ciprofloxacin with a hydroxyl group-containing aliphatic triazole linker (hybrid 9). Most antibiotic hybrid drugs consist of a fluoroquinolone.

FIG 8

Structure of cefiderocol (hybrid 10), previously known as S-649266, derived by linking ceftazidime to the siderophore catechol 2-chloro-3,4-dihydroxybenzoic acid. This β-lactam–siderophore hybrid possesses potent antibacterial activity against metallo-β-lactamase-producing Gram-negative bacilli.

FIG 9

Examples of tobramycin-based hybrids: tobramycin linked to ciprofloxacin (hybrid 11), tobramycin linked to moxifloxacin (hybrid 12), and tobramycin linked to a lysine peptoid mimic (hybrid 13). All three tobramycin-based hybrids contain a 12-carbon-long aliphatic (C₁₂) hydrocarbon linker.

FIG 10

Examples of tobramycin-efflux pump hybrids: tobramycin linked to 1-(1-naphthylmethyl)-piperazine (NMP) (hybrid 14), tobramycin linked to paroxetine (hybrid 15), and tobramycin linked to the dibasic peptide (DBP) analog d-Ala–d-hPhe–aminoquinoline (MC-04,124) (hybrid 16). All three tobramycin-based hybrids contain a 12-carbon-long aliphatic (C₁₂) hydrocarbon linker.

FIG 11

Potentiation of legacy antibiotics against P. aeruginosa PAO1 by tobramycin-based hybrids: tobramycin-ciprofloxacin hybrid 11 (Tob-Cip), tobramycin-lysine peptoid hybrid 13 (Tob-Lys), and tobramycin-NMP hybrid 14 (Tob-NMP). Tobramycin-lysine peptoid and tobramycin-NMP could not potentiate colistin and were not tested with ciprofloxacin.

FIG 12

Proposed mechanism(s) of action of tobramycin-based antibiotic hybrids.

FIG 13

Examples of advanced hybrid antibiotics undergoing clinical trials: cadazolid (hybrid 17) and TNP-2092, also known as CBR-2092 (hybrid 18). Cadazolid was derived by fusing ciprofloxacin and tedizolid (with overlapping pharmacophores), while TNP-2092 comprises ciprofloxacin and rifampin-derived pharmacophores. These hybrids display limited antibacterial activity against Gram-negative bacilli.