RESUMEN
Eradication of Mycobacterium tuberculosis (MTB) infection requires daily administration of combinations of rifampin (RIF), isoniazid [isonicotinylhydrazine (INH)], pyrazinamide, and ethambutol, among other drug therapies. To facilitate and optimize MTB therapeutic selections, a mononuclear phagocyte (MP; monocyte, macrophage, and dendritic cell)-targeted drug delivery strategy was developed. Long-acting nanoformulations of RIF and an INH derivative, pentenyl-INH (INHP), were prepared, and their physicochemical properties were evaluated. This included the evaluation of MP particle uptake and retention, cell viability, and antimicrobial efficacy. Drug levels reached 6 µg/10(6) cells in human monocyte-derived macrophages (MDMs) for nanoparticle treatments compared with 0.1 µg/10(6) cells for native drugs. High RIF and INHP levels were retained in MDM for >15 d following nanoparticle loading. Rapid loss of native drugs was observed in cells and culture fluids within 24 h. Antimicrobial activities were determined against Mycobacterium smegmatis (M. smegmatis). Coadministration of nanoformulated RIF and INHP provided a 6-fold increase in therapeutic efficacy compared with equivalent concentrations of native drugs. Notably, nanoformulated RIF and INHP were found to be localized in recycling and late MDM endosomal compartments. These were the same compartments that contained the pathogen. Our results demonstrate the potential of antimicrobial nanomedicines to simplify MTB drug regimens.
Asunto(s)
Antituberculosos/farmacología , Endosomas/efectos de los fármacos , Macrófagos/efectos de los fármacos , Nanopartículas , Células Cultivadas , Endosomas/metabolismo , Humanos , Macrófagos/metabolismo , Fracciones Subcelulares/metabolismoRESUMEN
The drug delivery platform for folic acid (FA)-coated nanoformulated ritonavir (RTV)-boosted atazanavir (FA-nanoATV/r) using poloxamer 407 was developed to enhance cell and tissue targeting for a range of antiretroviral drugs. Such formulations would serve to extend the drug half-life while improving the pharmacokinetic profile and biodistribution to reservoirs of human immunodeficiency virus (HIV) infection. To this end, we now report enhanced pharmacokinetics and drug biodistribution with limited local and systemic toxicities of this novel nanoformulation. The use of FA as a targeting ligand for nanoATV/r resulted in plasma and tissue drug concentrations up to 200-fold higher compared to equimolar doses of native drug. In addition, ATV and RTV concentrations in plasma from mice on a folate-deficient diet were up to 23-fold higher for mice administered FA-nanoATV/r than for mice on a normal diet. Compared to earlier nanoATV/r formulations, FA-nanoATV/r resulted in enhanced and sustained plasma and tissue ATV concentrations. In a drug interaction study, ATV plasma and tissue concentrations were up to 5-fold higher in mice treated with FA-nanoATV/r than in mice treated with FA-nanoATV alone. As observed in mice, enhanced and sustained plasma concentrations of ATV were observed in monkeys. NanoATV/r was associated with transient local inflammation at the site of injection. There were no systemic adverse reactions associated with up to 10 weeks of chronic exposure of mice or monkeys to FA-nanoATV/r.
Asunto(s)
Fármacos Anti-VIH/farmacocinética , Portadores de Fármacos/farmacocinética , Ácido Fólico/química , Nanoestructuras , Oligopéptidos/farmacocinética , Piridinas/farmacocinética , Ritonavir/farmacocinética , Animales , Fármacos Anti-VIH/sangre , Sulfato de Atazanavir , Esquema de Medicación , Portadores de Fármacos/química , Composición de Medicamentos , Evaluación Preclínica de Medicamentos , Ácido Fólico/metabolismo , Alimentos Formulados , Semivida , Humanos , Inyecciones Intramusculares , Macaca mulatta , Masculino , Ratones , Ratones Endogámicos C57BL , Nanoestructuras/química , Oligopéptidos/sangre , Poloxámero/química , Piridinas/sangre , Ritonavir/sangre , Distribución TisularRESUMEN
Long-acting injectable nanoformulated antiretroviral therapy (nanoART) was developed with the explicit goal of improving medicine compliance and for drug targeting of viral tissue reservoirs. Prior nanoART studies completed in humanized virus-infected mice demonstrated sustained antiretroviral responses. However, the pharmacokinetics (PK) and tissue distribution of nanoART were not characterized. To this end, the PK and tissue distribution of nanoformulated atazanavir (ATV) and ritonavir (RTV) injected subcutaneously or intramuscularly in mice and monkeys were evaluated. Fourteen days after injection, ATV and RTV levels were up to 13-, 41-, and 4,500-fold higher than those resulting from native-drug administration in plasma, tissues, and at the site of injection, respectively. At nanoART doses of 10, 50, 100, and 250 mg/kg of body weight, relationships of more- and less-than-proportional increases in plasma and tissue levels with dose increases were demonstrated with ATV and RTV. Multiple-dose regimens showed serum and tissue concentrations up to 270-fold higher than native-drug concentrations throughout 8 weeks of study. Importantly, nanoART was localized in nonlysosomal compartments in tissue macrophages, creating intracellular depot sites. Reflective data were obtained in representative rhesus macaque studies. We conclude that nanoART demonstrates blood and tissue antiretroviral drug levels that are enhanced compared to those of native drugs. The sustained and enhanced PK profile of nanoART is, at least in part, the result of the sustained release of ATV and RTV from tissue macrophases and at the site of injection.
Asunto(s)
Fármacos Anti-VIH/farmacocinética , Inhibidores de la Proteasa del VIH/farmacocinética , Oligopéptidos/farmacocinética , Piridinas/farmacocinética , Ritonavir/farmacocinética , Animales , Fármacos Anti-VIH/administración & dosificación , Fármacos Anti-VIH/sangre , Terapia Antirretroviral Altamente Activa , Sulfato de Atazanavir , Esquema de Medicación , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , Inhibidores de la Proteasa del VIH/administración & dosificación , Inhibidores de la Proteasa del VIH/sangre , Macaca mulatta , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Oligopéptidos/administración & dosificación , Oligopéptidos/sangre , Piridinas/administración & dosificación , Piridinas/sangre , Ritonavir/administración & dosificación , Ritonavir/sangre , Distribución TisularRESUMEN
Macrophages serve as vehicles for the carriage and delivery of polymer-coated nanoformulated antiretroviral therapy (nanoART). Although superior to native drug, high drug concentrations are required for viral inhibition. Herein, folate-modified ritonavir-boosted atazanavir (ATV/r)-encased polymers facilitated macrophage receptor targeting for optimizing drug dosing. Folate coating of nanoART ATV/r significantly enhanced cell uptake, retention and antiretroviral activities without altering cell viability. Enhanced retentions of folate-coated nanoART within recycling endosomes provided a stable subcellular drug depot. Importantly, up to a five-fold enhanced plasma and tissue drug levels followed folate-coated formulation injection in mice. Folate polymer encased ATV/r improves nanoART pharmacokinetics bringing the technology one step closer to human use. FROM THE CLINICAL EDITOR: This team of authors describes a novel method for macrophage folate receptor-targeted antiretroviral therapy. Atazanvir entry, retention, and antiretroviral activities were superior using the presented method, and so was its biodistribution, enabling a more efficient way to address human immunodeficiency virus infections, with a hoped for clinical application in the near future.
Asunto(s)
Fármacos Anti-VIH/uso terapéutico , Receptores de Folato Anclados a GPI/metabolismo , Ácido Fólico/uso terapéutico , Infecciones por VIH/tratamiento farmacológico , VIH-1/efectos de los fármacos , Macrófagos/metabolismo , Oligopéptidos/uso terapéutico , Piridinas/uso terapéutico , Animales , Fármacos Anti-VIH/administración & dosificación , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacocinética , Sulfato de Atazanavir , Células Cultivadas , Sistemas de Liberación de Medicamentos , Ácido Fólico/administración & dosificación , Ácido Fólico/análogos & derivados , Ácido Fólico/farmacocinética , Infecciones por VIH/metabolismo , Humanos , Macrófagos/efectos de los fármacos , Macrófagos/virología , Masculino , Ratones , Ratones Endogámicos BALB C , Oligopéptidos/administración & dosificación , Oligopéptidos/química , Oligopéptidos/farmacocinética , Piridinas/administración & dosificación , Piridinas/química , Piridinas/farmacocinética , Distribución TisularRESUMEN
AIM: While the therapeutic potential for current long-acting (LA) antiretroviral therapy (ART) is undeniable, ligand-decorated nanoformulated LA-ART could optimize drug delivery to viral reservoirs. The development of decorated ART hinges, however, on formulation processes and manufacture efficiencies. To this end, we compared manufacture and purification techniques for ligand-decorated antiretroviral drug nanocrystals. MATERIALS & METHODS: Ligand-decorated nanoparticle manufacturing was tested using folic acid (FA) nanoformulated cabotegravir. RESULTS: Direct manufacturing of FA-cabotegravir resulted in stable particles with high drug loading and monocyte-macrophage targeting. A one step 'direct' scheme proved superior over differential centrifugation or tangential flow filtration facilitating particle stability and preparation simplicity and efficiency. CONCLUSION: Direct manufacturing of FA nanoparticles provides a path toward large-scale clinical grade manufacturing of cell-targeted LA-ART.
Asunto(s)
Antirretrovirales/administración & dosificación , Sistemas de Liberación de Medicamentos , Infecciones por VIH/tratamiento farmacológico , Nanopartículas/administración & dosificación , Animales , Antirretrovirales/química , Modelos Animales de Enfermedad , Ácido Fólico/administración & dosificación , Ácido Fólico/química , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/patogenicidad , Humanos , Macrófagos/efectos de los fármacos , Ratones , Nanopartículas/química , Piridonas/administración & dosificación , Piridonas/química , Distribución Tisular/efectos de los fármacosRESUMEN
Long-acting nanoformulated antiretroviral therapy (nanoART) can sustain plasma drug levels and improve its biodistribution. Cell targeted-nanoART can achieve this and bring drug efficiently to viral reservoirs. However, whether such improvements affect antiretroviral responses remains unknown. To these ends, we tested folic acid (FA)-linked poloxamer407-coated ritonavir-boosted atazanavir (FA-nanoATV/r) nanoparticles for their ability to affect chronic HIV-1 infection in humanized mice. Following three, 100mg/kg FA-nanoATV/r intramuscular injections administered every other week to infected animals, viral RNA was at or below the detection limit, cell-associated HIV-1p24 reduced and CD4+ T cell counts protected. The dosing regimen improved treatment outcomes more than two fold from untargeted nanoATV/r. We posit that these nanoformulations have potential for translation to human use.
Asunto(s)
Antirretrovirales/farmacocinética , Preparaciones de Acción Retardada/farmacocinética , Portadores de Fármacos/metabolismo , Receptores de Folato Anclados a GPI/metabolismo , Nanomedicina/métodos , Nanopartículas/metabolismo , Animales , Antirretrovirales/administración & dosificación , Sulfato de Atazanavir/administración & dosificación , Sulfato de Atazanavir/farmacocinética , Recuento de Linfocito CD4 , Preparaciones de Acción Retardada/administración & dosificación , Portadores de Fármacos/administración & dosificación , Ácido Fólico/administración & dosificación , Ácido Fólico/metabolismo , Infecciones por VIH/tratamiento farmacológico , Humanos , Inyecciones Intramusculares , Ratones SCID , Nanopartículas/administración & dosificación , Poloxámero/administración & dosificación , Poloxámero/metabolismo , ARN Viral/sangre , Ritonavir/administración & dosificación , Ritonavir/farmacocinética , Resultado del Tratamiento , Carga ViralRESUMEN
Regimen adherence, systemic toxicities, and limited drug penetrance to viral reservoirs are obstacles limiting the effectiveness of antiretroviral therapy (ART). Our laboratory's development of the monocyte-macrophage-targeted long-acting nanoformulated ART (nanoART) carriage provides a novel opportunity to simplify drug-dosing regimens. Progress has nonetheless been slowed by cumbersome, but required, pharmacokinetic (PK), pharmacodynamics, and biodistribution testing. To this end, we developed a small magnetite ART (SMART) nanoparticle platform to assess antiretroviral drug tissue biodistribution and PK using magnetic resonance imaging (MRI) scans. Herein, we have taken this technique a significant step further by determining nanoART PK with folic acid (FA) decorated magnetite (ultrasmall superparamagnetic iron oxide [USPIO]) particles and by using SMART particles. FA nanoparticles enhanced the entry and particle retention to the reticuloendothelial system over nondecorated polymers after systemic administration into mice. These data were seen by MRI testing and validated by comparison with SMART particles and direct evaluation of tissue drug levels after nanoART. The development of alendronate (ALN)-coated magnetite thus serves as a rapid initial screen for the ability of targeting ligands to enhance nanoparticle-antiretroviral drug biodistribution, underscoring the value of decorated magnetite particles as a theranostic tool for improved drug delivery.
Asunto(s)
Antirretrovirales/farmacocinética , Imagen por Resonancia Magnética/métodos , Nanopartículas de Magnetita , Alendronato/química , Animales , Sulfato de Atazanavir/farmacocinética , Dextranos , Sistemas de Liberación de Medicamentos/métodos , Ácido Fólico/química , Ácido Fólico/farmacocinética , Macrófagos/efectos de los fármacos , Nanopartículas de Magnetita/química , Masculino , Ratones , Ratones Endogámicos BALB C , Terapia Molecular Dirigida/métodos , Sistema Mononuclear Fagocítico/efectos de los fármacos , Nanomedicina/métodos , Reproducibilidad de los Resultados , Distribución TisularRESUMEN
Long-acting nanoformulated antiretroviral therapy (nanoART) induces a range of innate immune migratory, phagocytic and secretory cell functions that perpetuate drug depots. While recycling endosomes serve as the macrophage subcellular depots, little is known of the dynamics of nanoART-cell interactions. To this end, we assessed temporal leukocyte responses, drug uptake and distribution following both intraperitoneal and intramuscular injection of nanoformulated atazanavir (nanoATV). Local inflammatory responses heralded drug distribution to peritoneal cell populations, regional lymph nodes, spleen and liver. This proceeded for three days in male Balb/c mice. NanoATV-induced changes in myeloid populations were assessed by fluorescence-activated cell sorting (FACS) with CD45, CD3, CD11b, F4/80, and GR-1 antibodies. The localization of nanoATV within leukocyte cell subsets was determined by confocal microscopy. Combined FACS and ultra-performance liquid chromatography tandem mass-spectrometry assays determined nanoATV carriages by cell-based vehicles. A robust granulocyte, but not peritoneal macrophage nanoATV response paralleled zymosan A treatment. ATV levels were highest at sites of injection in peritoneal or muscle macrophages, dependent on the injection site. The spleen and liver served as nanoATV tissue depots while drug levels in lymph nodes were higher than those recorded in plasma. Dual polymer and cell labeling demonstrated a nearly exclusive drug reservoir in macrophages within the liver and spleen. Overall, nanoART induces innate immune responses coincident with rapid tissue macrophage distribution. Taken together, these works provide avenues for therapeutic development designed towards chemical eradication of human immunodeficiency viral infection.
Asunto(s)
Sulfato de Atazanavir/administración & dosificación , Portadores de Fármacos/administración & dosificación , Inhibidores de la Proteasa del VIH/administración & dosificación , Nanopartículas/administración & dosificación , Animales , Sulfato de Atazanavir/farmacocinética , Química Farmacéutica , Preparaciones de Acción Retardada/farmacocinética , Portadores de Fármacos/farmacocinética , Sistemas de Liberación de Medicamentos , Infecciones por VIH/tratamiento farmacológico , Inhibidores de la Proteasa del VIH/farmacocinética , Humanos , Inmunidad Innata , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Microscopía Confocal , Distribución TisularRESUMEN
Long-acting nanoformulated antiretroviral therapy (nanoART) that targets monocyte-macrophages could improve the drug's half-life and protein-binding capacities while facilitating cell and tissue depots. To this end, ART nanoparticles that target the folic acid (FA) receptor and permit cell-based drug depots were examined using pharmacokinetic and pharmacodynamic (PD) tests. FA receptor-targeted poloxamer 407 nanocrystals, containing ritonavir-boosted atazanavir (ATV/r), significantly increased drug bioavailability and PD by five and 100 times, respectively. Drug particles administered to human peripheral blood lymphocyte reconstituted NOD.Cg-Prkdc(scid)Il2rg(tm1Wjl)/SzJ mice and infected with HIV-1ADA led to ATV/r drug concentrations that paralleled FA receptor beta staining in both the macrophage-rich parafollicular areas of spleen and lymph nodes. Drug levels were higher in these tissues than what could be achieved by either native drug or untargeted nanoART particles. The data also mirrored potent reductions in viral loads, tissue viral RNA and numbers of HIV-1p24+ cells in infected and treated animals. We conclude that FA-P407 coating of ART nanoparticles readily facilitates drug carriage and antiretroviral responses.
Asunto(s)
Receptor 1 de Folato/metabolismo , Nanopartículas/química , Oligopéptidos/farmacología , Piridinas/farmacología , Ritonavir/farmacología , Animales , Antirretrovirales/farmacología , Antígenos CD/metabolismo , Sulfato de Atazanavir , Química Farmacéutica , Citometría de Flujo , Proteína p24 del Núcleo del VIH/metabolismo , Humanos , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Oligopéptidos/farmacocinética , Poloxámero/química , Piridinas/farmacocinética , Ritonavir/farmacocinética , Bazo/efectos de los fármacos , Bazo/metabolismo , Linfocitos T/inmunología , Distribución Tisular/efectos de los fármacosRESUMEN
AIM: Drug toxicities, compliance and penetrance into viral reservoirs have diminished the efficacy of long-term antiretroviral therapy (ART) for treatment of HIV infection. Cell-targeted nanoformulated ART was developed to improve disease outcomes. However, rapid noninvasive determination of drug biodistribution is unrealized. To this end, small magnetite ART (SMART) nanoparticles can provide assessments of ART biodistribution by MRI. MATERIALS & METHODS: Poly(lactic-co-glycolic acid), 1,2-distearoyl-sn-glycero-3-phosphocholine- and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-(methoxy-PEG 2000)-encased particles were synthesized with atazanavir (ATV) and magnetite. Uptake and retention of ATV and magnetite administered at 3:1 ratios (weight/weight) were determined in human monocyte-derived macrophages and mice. RESULTS: SMART particles were taken up and retained in macrophages. In mice, following parenteral SMART injection, magnetite and drug biodistribution paralleled one another with MRI signal intensity greatest in the liver and spleen at 24 h. Significantly, ATV and magnetite levels correlated. CONCLUSION: SMART can permit rapid assessment of drug tissue concentrations in viral reservoirs.