Inhalable PEGylated Phospholipid Liposheres Containing Paclitaxel for Targeted Pulmonary Delivery for Lung Cancer Applications

Despite the significant advances in the treatment of lung cancer, it is a disease that still signifies poor prognosis due to the challenges in implementation of treatment. Targeted pulmonary inhalation drug delivery offers many advantages for lung cancer patients in comparison to conventional systemic chemotherapy. These include the potential to deliver local therapeutically effective concentrations of drug directly to the lung, minimized side effects due to limited systemic delivery, and ease of use for the patient. Inhalable dry powder formulations of nanoparticles and microparticles (lipospheres) containing a chemotherapeutic are advantageous in their ability to deliver drug deep in the lung via optimally sized particles, higher local drug dose delivery, and long-term storage capability. In this work, novel advanced spray-dried inhalable PEGylated phospholipid liposphere powders containing the chemotherapeutic paclitaxel were successfully designed and produced via dilute organic solution advanced spray drying under various conditions. Fixed ratios of dipalmatoylphosphatidylcholine (DPPC) and dipalmatoylphosphatidylethanolamine poly(ethylene glycol) (DPPE-PEG) at three different polymeric chain lengths were combined with various ratios of paclitaxel in a dilute methanol solution. Upon optimization of the spray drying conditions (e.g. pump rate), the physicochemical characterization of the particles was completed. Scanning electron microscopy (SEM) images showed the spherical particle morphology of the inhalable particles. The size of the particles was statistically analyzed using these images SigmaScan software, and these particles were determined to be 600 nm – 1.2 μm in diameter, which is optimal for efficient targeting of the deep lung alveolar and small airway regions for enhanced local deposition. Differential scanning calorimetry (DSC) and X-ray powder diffraction (XRPD) were performed to analyze solid-state transitions and long-range molecular order, respectively, and allowed for the confirmation of the presence of phospholipid bilayers and/or paclitaxel and their phase transition behavior. The water content of the particles was very low as quantified analytically via Karl Fisher titration. The composition of the particles was confirmed using attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. Confocal Raman microspectroscopy was employed in chemical imaging to assess particle composition and miscibility. The amount of paclitaxel loaded into the particles was analyzed via high performance liquid chromatography (HPLC), and their aerosol performance was evaluated using the Next Generation Impactor (NGI) and an approved dry powder inhaler (DPI) device for human use to determine the emitted dose, respirable dose, fine particle fraction and mass median aerodynamic diameter. Overall, these results demonstrate this novel therapeutic nanomedicine platform as one capable of effectively delivering paclitaxel directly to the lung in high local concentration for the treatment of lung cancer

SYNTHESIS AND CHARACTERIZATION OF MAGNETIC HYDROGEL NANOCOMPOSITES FOR CANCER THERAPY APPLICATIONS

Currently, cancer is the second leading cause of death in the United States. Conventional cancer treatment includes chemotherapy, radiation, and surgical resection, but unfortunately, all of these methods have significant drawbacks. Hyperthermia, the heating of cancerous tissues to between 41 and 45°C, has been shown to improve the efficacy of cancer therapy when used in conjunction with irradiation and/or chemotherapy. In this work, a novel method for remotely administering heat is presented. This method involves heating of tumor tissue using hydrogel nanocomposites containing magnetic nanoparticles which can be remotely heated upon exposure to an external alternating magnetic field (AMF). The iron oxide nanoparticles contained in the hydrogel nanocomposites are able to heat via an AMF due to Brownian and Neel relaxation processes. The administration of hyperthermia via hydrogel nanocomposites allows for local delivery of heat to tumor tissue while also providing a drug depot to deliver chemotherapeutic agents. Both in vivo and in vitro studies have demonstrated that numerous chemotherapeutic agents, when used in conjunction with hyperthermia, show improved efficacy in treating cancer Various magnetic hydrogel nanocomposites were synthesized and characterized for this work including poly(ethylene glycol) (PEG)-based hydrogels, which were studied due to their inherent biocompatibility and “stealth” properties, as well as, poly(β-amino ester) (PBAE)-based hydrogels which have tailorable degradation properties. The PEG hydrogels were investigated for their temperature-responsiveness swelling, mechanical strength, heating capabilities, biocompatibility, ability to kill M059K glioblastoma cells via thermoablation, and the ability to deliver paclitaxel, a chemotherapeutic agent. PBAE hydrogels were also characterized for their degradation and swelling properties, ability to heat upon exposure to an AMF, biocompatibility, mechanical strength, and ability to deliver paclitaxel in a controlled fashion. Additionally, multiple cancer cell lines were exposed to a combination of paclitaxel and heat (at 42.5 °C) in vitro and it was shown that A539 lung carcinoma cells exhibit higher cytotoxicity when exposed to both heat and paclitaxel than either treatment alone. Overall, magnetic hydrogel nanocomposites are promising materials that can be utilized for the multi-modality treatment of cancer through the synergistic delivery of both heat and chemotherapeutic agents

Inhalable PEGylated Phospholipid Lipospheres Containing Paclitaxel for Trageted Pulmonary Delivery for Lung Cancer Applications

Despite the significant advances in the treatment of lung cancer, it is a disease that still signifies poor prognosis due to the challenges in implementation of treatment. Targeted pulmonary inhalation drug delivery offers many advantages for lung cancer patients in comparison to conventional systemic chemotherapy. These include the potential to deliver local therapeutically effective concentrations of drug directly to the lung, minimized side effects due to limited systemic delivery, and ease of use for the patient. Inhalable dry powder formulations of nanoparticles and microparticles (lipospheres) containing a chemotherapeutic are advantageous in their ability to deliver drug deep in the lung via optimally sized particles, higher local drug dose delivery, and long-term storage capability. In this work, novel advanced spray-dried inhalable PEGylated phospholipid liposphere powders containing the chemotherapeutic paclitaxel were successfully designed and produced via dilute organic solution advanced spray drying under various conditions. Fixed ratios of dipalmatoylphosphatidylcholine (DPPC) and dipalmatoylphosphatidylethanolamine poly(ethylene glycol) (DPPE-PEG) at three different polymeric chain lengths were combined with various ratios of paclitaxel in a dilute methanol solution. Upon optimization of the spray drying conditions (e.g. pump rate), the physicochemical characterization of the particles was completed. Scanning electron microscopy (SEM) images showed the spherical particle morphology of the inhalable particles. The size of the particles was statistically analyzed using these images SigmaScan software, and these particles were determined to be 600 nm – 1.2 μm in diameter, which is optimal for efficient targeting of the deep lung alveolar and small airway regions for enhanced local deposition. Differential scanning calorimetry (DSC) and X-ray powder diffraction (XRPD) were performed to analyze solidstate transitions and long-range molecular order, respectively, and allowed for the confirmation of the presence of phospholipid bilayers and/or paclitaxel and their phase transition behavior. The water content of the particles was very low as quantified analytically via Karl Fisher titration. The composition of the particles was confirmed using attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. Confocal Raman microspectroscopy was employed in chemical imaging to assess particle composition and miscibility. The amount of paclitaxel loaded into the particles was analyzed via high performance liquid chromatography (HPLC), and their aerosol performance was evaluated using the Next Generation Impactor (NGI) and an approved dry powder inhaler (DPI) device for human use to determine the emitted dose, respirable dose, fine particle fraction and mass median aerodynamic diameter. Overall, these results demonstrate this novel therapeutic nanomedicine platform as one capable of effectively delivering paclitaxel directly to the lung in high local concentration for the treatment of lung cancer

Synthesis and Characterization of Nanocomposite Microparticles (nCmP) for the Treatment of Cystic Fibrosis-Related Infections

Purpose: Pulmonary antibiotic delivery is recommended as maintenance therapy for cystic fibrosis (CF) patients who experience chronic infections. However, abnormally thick and sticky mucus present in the respiratory tract of CF patients impairs mucus penetration and limits the efficacy of inhaled antibiotics. To overcome the obstacles of pulmonary antibiotic delivery, we have developed nanocomposite microparticles (nCmP) for the inhalation application of antibiotics in the form of dry powder aerosols. Methods: Azithromycin-loaded and rapamycin-loaded polymeric nanoparticles (NP) were prepared via nanoprecipitation and nCmP were prepared by spray drying and the physicochemical characteristics were evaluated. Results: The nanoparticles were 200 nm in diameter both before loading into and after redispersion from nCmP. The NP exhibited smooth, spherical morphology and the nCmP were corrugated spheres about 1 μm in diameter. Both drugs were successfully encapsulated into the NP and were released in a sustained manner. The NP were successfully loaded into nCmP with favorable encapsulation efficacy. All materials were stable at manufacturing and storage conditions and nCmP were in an amorphous state after spray drying. nCmP demonstrated desirable aerosol dispersion characteristics, allowing them to deposit into the deep lung regions for effective drug delivery. Conclusions: The described nCmP have the potential to overcome mucus-limited pulmonary delivery of antibiotics

Synthesis and Characterization of CREKA-Conjugated Iron Oxide Nanoparticles for Hyperthermia Applications

One of the current challenges in the systemic delivery of nanoparticles in cancer therapy applications is the lack of effective tumor localization. Iron oxide nanoparticles coated with crosslinked dextran were functionalized with the tumor homing peptide CREKA, which binds to fibrinogen complexes in the extracellular matrix of tumors. This allows for the homing of these nanoparticles to tumor tissue. The iron oxide nanoparticle core allows for particle heating upon exposure to an alternating magnetic field (AMF) while the dextran coating stabilizes the particles in suspension and decreases the cytotoxicity of the system. Magnetically mediated hyperthermia (MMH) allows for the heating of tumor tissue to increase the efficacy of traditional cancer treatments using the iron oxide nanoparticles. While MMH provides the opportunity for localized heating, this method is currently limited by the lack of particle penetration into tumor tissue, even after effective targeted delivery to the tumor site. The CREKA-conjugated nanoparticles presented were characterized for their size, stability, biocompatibility, and heating capabilities. The particles were stable in PBS and media over at least twelve hours, had a hydrated diameter of 52 nm, and generated enough heat to raise solution temperatures well into the hyperthermia range (41 – 45 °C) when exposed to an AMF. Biocompatibility studies demonstrated that the particles have low cytotoxicity over long exposure times at low concentrations. A fibrinogen clotting assay was used to determine the binding affinity of CREKA-conjugated particles, which was significantly greater than the binding affinity of dextran, only coated iron oxide nanoparticles demonstrating the potential for this particle system to effectively home to a variety of tumor locations. Finally, it was shown that in vitro MMH increased the effects of cisplatin compared to cisplatin or MMH treatments alone

Characterization and Aerosol Dispersion Performance of Spray-Dried Chemotherapeutic PEGylated Phospholipid Particles for Dry Powder Inhalation Delivery in Lung Cancer

Pulmonary inhalation chemotherapeutic drug delivery offers many advantages for lung cancer patients in comparison to conventional systemic chemotherapy. Inhalable particles are advantageous in their ability to deliver drug deep in the lung by utilizing optimally sized particles and higher local drug dose delivery. In this work, spray-dried and co-spray dried inhalable lung surfactant-mimic PEGylated lipopolymers as microparticulate/nanoparticulate dry powders containing paclitaxel were rationally designed via organic solution advanced spray drying (no water) in closed-mode from dilute concentration feed solution. Dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylethanolamine poly(ethylene glycol) (DPPE-PEG) with varying PEG chain length were mixed with varying amounts of paclitaxel in methanol to produce co-spray dried microparticles and nanoparticles. Scanning electron microscopy showed the spherical particle morphology of the inhalable particles. Thermal analysis and X-ray powder diffraction confirmed the retention of the phospholipid bilayer structure in the solid-state following spray drying, the degree of solid-state molecular order, and solid-state phase transition behavior. The residual water content of the particles was very low as quantified analytically Karl Fisher titration. The amount of paclitaxel loaded into the particles was quantified which indicated high encapsulation efficiencies (43-99%). Dry powder aerosol dispersion performance was measure in vitro using the Next Generation Impactor™ (NGI™) coupled with the Handihaler® dry powder inhaler device and showed mass median aerodynamic diameters in the range of 3.4 – 7μm. These results demonstrate that this novel microparticulate/nanoparticulate chemotherapeutic PEGylated phospholipid inhalation aerosol platform has great potential in lung cancer drug delivery

Design, Physicochemical Characterization, and Optimization of Organic Solution Advanced Spray-Dried Inhalable Dipalmitoylphosphatidylcholine (DPPC) and Dipalmitoylphosphatidylethanolamine Poly(Ethylene Glycol) (DPPE-PEG) Microparticles and Nanoparticles for Targeted Respiratory Nanomedicine Delivery as Dry Powder Inhalation Aerosols

Novel advanced spray-dried and co-spray-dried inhalable lung surfactant-mimic phospholipid and poly(ethylene glycol) (PEG)ylated lipopolymers as microparticulate/nanoparticulate dry powders of biodegradable biocompatible lipopolymers were rationally formulated via an organic solution advanced spray-drying process in closed mode using various phospholipid formulations and rationally chosen spray-drying pump rates. Ratios of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylethanolamine PEG (DPPE-PEG) with varying PEG lengths were mixed in a dilute methanol solution. Scanning electron microscopy images showed the smooth, spherical particle morphology of the inhalable particles. The size of the particles was statistically analyzed using the scanning electron micrographs and SigmaScan® software and were determined to be 600 nm to 1.2 μm in diameter, which is optimal for deep-lung alveolar penetration. Differential scanning calorimetry (DSC) and powder X-ray diffraction (PXRD) were performed to analyze solid-state transitions and long-range molecular order, respectively, and allowed for the confirmation of the presence of phospholipid bilayers in the solid state of the particles. The residual water content of the particles was very low, as quantified analytically via Karl Fischer titration. The composition of the particles was confirmed using attenuated total-reflectance Fourier-transform infrared (ATR-FTIR) spectroscopy and confocal Raman microscopy (CRM), and chemical imaging confirmed the chemical homogeneity of the particles. The dry powder aerosol dispersion properties were evaluated using the Next Generation Impactor™ (NGI™) coupled with the HandiHaler® dry powder inhaler device, where the mass median aerodynamic diameter from 2.6 to 4.3 μm with excellent aerosol dispersion performance, as exemplified by high values of emitted dose, fine particle fraction, and respirable fraction. Overall, it was determined that the pump rates defined in the spray-drying process had a significant effect on the solid-state particle properties and that a higher pump rate produced the most optimal system. Advanced dry powder inhalers of inhalable lipopolymers for targeted dry powder inhalation delivery were successfully achieved

Development of three-dimensional lung multicellular spheroids in air- and liquid-interface culture for the evaluation of anticancer therapeutics

Three-dimensional (3D) lung multicellular spheroids (MCS) in liquid-covered culture (LCC) and air-interface culture (AIC) conditions have both been developed for the evaluation of aerosol anticancer therapeutics in solution and aerosols, respectively. The MCS were formed by seeding lung cancer cells on top of collagen where they formed spheroids due to the prevalence of cell-to-cell interactions. LCC MCS were exposed to paclitaxel (PTX) in media whereas AIC MCS were exposed to dry powder PEGylated phospholipid aerosol microparticles containing paclitaxel. The difference in viability for 2D versus 3D culture for both LCC and AIC was evaluated along with the effects of the particles on lung epithelium via transepithelial electrical resistance (TEER) measurements. For LCC and AIC conditions, the 3D spheroids were more resistant to treatment with higher IC50 values for A549 and H358 cell lines. TEER results initially indicated a decrease in resistance upon drug or particle exposure, however, these values increased over the course of several days indicating the ability of the cells to recover. Overall, these studies offer a comprehensive in vitro evaluation of aerosol particles used in the treatment of lung cancer while introducing a new method for culturing lung cancer MCS in both LCC and AIC conditions

Development and Physicochemical Characterization of Acetalated Dextran Aerosol Particle Systems for Deep Lung Delivery

Biocompatible, biodegradable polymers are commonly used as excipients to improve the drug delivery properties of aerosol formulations, in which acetalated dextran (Ac-Dex) exhibits promising potential as a polymer in various therapeutic applications. Despite this promise, there is no comprehensive study on the use of Ac-Dex as an excipient for dry powder aerosol formulations. In this study, we developed and characterized pulmonary drug delivery aerosol microparticle systems based on spray-dried Ac-Dex with capabilities of (1) delivering therapeutics to the deep lung, (2) targeting the particles to a desired location within the lungs, and (3) releasing the therapeutics in a controlled fashion. Two types of Ac-Dex, with either rapid or slow degradation rates, were synthesized. Nanocomposite microparticle (nCmP) and microparticle (MP) systems were successfully formulated using both kinds of Ac-Dex as excipients and curcumin as a model drug. The resulting MP were collapsed spheres approximately 1μm in diameter, while the nCmP were similar in size with wrinkled surfaces, and these systems dissociated into 200nm nanoparticles upon reconstitution in water. The drug release rates of the Ac-Dex particles were tuned by modifying the particle size and ratio of fast to slow degrading Ac-Dex. The pH of the environment was also a significant factor that influenced the drug release rate. All nCmP and MP systems exhibited desirable aerodynamic diameters that are suitable for deep lung delivery (e.g. below 5μm). Overall, the engineered Ac-Dex aerosol particle systems have the potential to provide targeted and effective delivery of therapeutics into the deep lung

Dry powders based on mucus-penetrating nanoparticles entrapped microparticles for pulmonary delivery of Tobramycin

Pulmonary drug delivery system is increasingly recommended as maintenance therapy to prolong the interval between pulmonary exacerbations and to slow the progression of lung disease in cystic fibrosis (CF) patients with chronic P. aeruginosa infection due to its capability to achieve high drug concentrations at the site of infection and to minimize the risk of systemic toxicity. The most common used inhaled Tobramycin formation so far is nebulization such as Tobi® and Bramitob® which are regarded as inconvenient due to the long administration time and limited portability for chronic drug therapy in daily life of patients. The only dry powder formulation of Tobramycin is based on PulmoSphereTM technology, which has many advantages over nebulizers including faster delivery, easy use, portability, reduced need for cleaning and room temperature storage. Yet a lack of proof exists to indicate their efficient mucus penetration, which is the major obstacle for pulmonary drug delivery. To overcome the shortcomings of established pulmonary antibiotic delivery, we proposed the use of mucus-penetrating nanoparticles entrapped microparticles (so-called nanocomposite microparticles) combining the advantages of both nanoparticles and microparticles. The nanoparticles were comprised of the anti-biotic tobramycin encapsulated in the polymer acetalated dextran (Ac-Dex) and PVA coating, which enables the system to penetrate the mucus and to release drug in controlled rate. The nanoparticles were then entrapped in microparticles using advanced organic spray drying techniques which can improve the targeted delivery of the drug. This system will enlighten the dry powder based antibiotic delivery providing a desirable alternative way for inhalation therapy