Inhalable Macromolecular Prodrugs of Ciprofloxacin for the Treatment of Pulmonary Intracellular Bacterial Infections

Abstract

Alveolar intracellular bacterial infections, such as those caused by Burkholderia pseudomallei and Francisella tularensis, are one of the most challenging infectious disease settings where the appropriate treatment of these pulmonary pathogens remains an important unmet clinical need. In this thesis, we outline the design and development of an inhalable macromolecular prodrug technology against airborne F. tularensis (pulmonary tularemia). Francisella is an infectious bacterium with a high global burden, historical precedence of being weaponized for biological warfare, and is currently classified by the U.S. Centers for Disease Control and Prevention as a Tier 1 bioagent capable of mass public harm and casualty. The current standards of care for treating respiratory infections of F. tularensis relies on rigorous prolonged applications of oral and intravenous antibiotics. In greater than forty-percent of cases, these therapies clinically fail to clear the bacterial infection due to poor pulmonary biodistribution and sustained localized drug concentrations causing a high-rate of fatality and disease relapse. Direct pulmonary drug delivery offers a unique opportunity to control drug concentrations at the site of bacterial persistence, and is gaining popularity as an attractive strategy for the treatment of pulmonary infections. Inhalable free-drug dispersions and liposomal based antibiotic formulations have been clinically exploited against bacteria such as Pseudomonas aeruginosa in cystic fibrosis patients, however, with limited success due to poor pulmonary pharmacokinetics brought on by rapid drug release, low drug encapsulation efficiencies, and complex formulations procedures that affect manufacturing scalability and reproducibility. To address these limitations, we have synthesized polymeric prodrugs with high-drug densities of a model antibiotic, ciprofloxacin, from polymerizable drug monomers that can provide rapid extended therapy against respiratory tularemia and reduce drug dosing. Our macromolecular ciprofloxacin prodrugs deliver sustained release of antibiotics via ester hydrolysis from engineered chemically-labile drug-linkers, with the ability to control drug release kinetics with the choice of various linkers and polymer architecture. Specifically, ciprofloxacin polymeric prodrugs derived from a phenolic ester modified drug linker showed faster hydrolysis kinetics in human serum with 50% of the drug released within 5d, whereas constructs with an alkyllic ester linkage demonstrated similar release over 22d. Using a quantitative LC-MS approach, this difference in drug release was also captured in vivo with pulmonary drug half-lives of 9.3h and 15.6h from the phenol and alkyllic ester macromolecular prodrugs, respectively. Establishing appropriate in vivo antibiotic pharmacokinetic-pharmacodynamic profiles is critical for promoting therapeutic efficacy. We observed that having a slower alkyl-ester modified drug linker within these unimeric copolymer morphologies although provides improved stability, it fails to meet specific PK-PD thresholds necessary for efficacy. This was further supported by evaluations using lethal aerosol exposures of F. tularensis in murine challenge models where we demonstrated enhanced survival rates with the endotrachaelly administered fast-releasing macromolecular prodrugs compared to slower-releasing variants and free drug controls. Modifying the polymer architecture provided an alternative approach to alter and control drug release profiles. Systematic hydrolysis studies in human serum showed that diblock copolymers where the drug was segregated to a second hydrophobic segment had considerably slower release kinetics compared to the molecularly soluble unimeric species. This was independent of the drug-linker suggesting that degree of solvation near the esters is important to achieve and can vary the hydrolysis rates. Current iterations of these macromolecular prodrugs extend from these observations to create more complex architectures such as mannosylated radiant star nanoparticles that can actively target and bind alveolar macrophages, and has improved drug release kinetics, compared to the previous micelle-forming diblock copolymers. In addition, the incorporation of mannose targeting chemistry can provide potential dose-sparing properties to overcome drug resistance. The manufacturing of these drug conjugates deviates from formulation based approaches and focuses on creative small-molecule synthetic strategies providing a highly-modular technology where the final macromolecular therapeutic can be engineered from a library of drugs, drug-linkers, and corresponding hydrophilic targeting moieties or solubilizers. These constructs characteristically afford an expanded and more versatile drug repertoire from conventional delivery vehicles, and the success of these macromolecular prodrugs stems from providing tunable, individualized drug release kinetics that control for in vivo PK parameters such as Cmax and AUC. This thesis establishes inhalable macromolecular prodrugs synthesized from polymerizable prodrug monomers as a promising modular platform technology that can not only treat aerosolized F. tularensis but may also be applicable for other invasive alveolar intracellular bacteria.