Development of anti-infective therapy against intracellular pathogens using targeted particulate delivery systems

Abstract

Pathogens from bacterial class: Salmonella, Mycobacterium tuberculosis, Listeria can survive and replicate intracellularly. While some pathogens remain cytosolic others such as Salmonella and Mycobacterium tuberculosis enhance their protection by forming vacuoles. The pathogens modulate the host cell’s antibacterial response and prevent clearance by lysosomes to suit their intracellular survival. Conventional antimicrobial agents cannot breach the bacterial defenses to attain required concentration required for killing the bacteria. The development of growing resistance in these intracellular pathogens is a direct consequence of insufficient antibiotic concentrations reaching such a shielded intracellular site. Hence, we present our approach to delivering antibiotics to intracellular vacuoles of pathogens to augment and improve existing antibacterial therapies. An oral nanoparticle-based antibiotic carrier was synthesized and evaluated as a therapy against Salmonella infection. The surface modification of the nanoparticles by lipid and polyelectrolyte coating was carried out to study targeting and delivery of ciprofloxacin into the Salmonella-infected cells. The lipid coating was carried out by sonicating liposomes with MSN, while polyelectrolyte coating on MSN was carried out by layer-by-layer technique. The nano-formulations were characterized by physical and chemical instrumental methods. In-vitro studies were carried out on macrophage and epithelial cells infected with Salmonella. The lipid coating improved the biocompatibility of the particle and surprisingly co-localized with the intracellular Salmonella. Arginine was functionalized on the polyelectrolyte-coated particle to exploit enhanced arginine requirements of Salmonella-infected cells. In-vitro experiments with ciprofloxacin-loaded Arg-MSN (Cip Arg-MSN) exhibited two-fold higher antibacterial activity with compared to free ciprofloxacin. We also observed an increase in the Reactive Nitrogen species (RNS) upon Arg-MSN administration. Salmonella burden in the infected organs such as spleen, liver, and MLN (mesenteric lymph nodes) and the mortality index decreased significantly after administration with lipid-coated and arginine-coated antibiotic carriers. The coordinated effect of improved antibiotic delivery, intracellular targeting, and production of reactive nitrogen species resulted in an improved therapeutic design against Salmonella infection. PLGA-MSN composite particle was evaluated as an antibiotic carrier for treating tuberculosis. Both hydrophobic and hydrophilic antibiotics could be efficiently loaded into this hybrid particle system. This composite particle was designed to be nebulized into the lung to achieve higher drug bioavailability. The composite particle was loaded with rifampicin and vancomycin and investigated to treat mice infected with Mycobacterium tuberculosis H37Rv strain. The particle treated mice showed nearly ten times lower MTB burden in lungs compared to nebulized antibiotic solutions studied in both, daily dosing and weekly dosing treatment schedules. The bacterial burden in major organs, pathological changes during infection and mortality index of the infected mice improved significantly using the developed particulate therapies against Salmonella and MTB infections. Thus, we present two interlinked strategies: one, by the therapeutic design of the antibiotic carrier and second, by an efficient delivery route to maximize therapeutic efficacy against intracellular pathogens