Samantha A. Meenach

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 (IONPs) 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 IONP 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 IONPs. 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, heating capabilities and biocompatibility. The particles had a hydrated diameter of 52nm, were stable in phosphate buffered saline solution and media with 10% v/v fetal bovine serum over at least 12h, and generated enough heat to raise solution temperatures well into the hyperthermia range (41-45°C) when exposed to an AMF, owing to an average specific absorption rate of 83.5Wg(-1). Cytotoxicity 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 IONPs 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 with cisplatin or MMH treatments alone.

Nanocomposite microparticles (nCmP) for the delivery of tacrolimus in the treatment of pulmonary arterial hypertension.

Tacrolimus (TAC) has exhibited promising therapeutic potential in the treatment of pulmonary arterial hypertension (PAH); however, its application is prevented by its poor solubility, instability, poor bioavailability, and negative systemic side effects. To overcome the obstacles of using TAC for the treatment of PAH, we developed nanocomposite microparticles (nCmP) for the pulmonary delivery of tacrolimus in the form of dry powder aerosols. These particles can provide targeted pulmonary delivery, improved solubility of tacrolimus, the potential of penetration through mucus barrier, and controlled drug release. In this system, tacrolimus-loaded polymeric nanoparticles (NP) were prepared via emulsion solvent evaporation and nCmP were prepared by spray drying these NP with mannitol. The NP were approximately 200nm in diameter with narrow size distribution both before loading into and after redispersion from nCmP. The NP exhibited smooth, spherical morphology and the nCmP were raisin-like spheres. High encapsulation efficacy was achieved both in the encapsulation of tacrolimus in NP and that of NP in nCmP. nCmP exhibited desirable aerosol dispersion properties, allowing them to deposit into the deep lung regions for effective drug delivery. A549 cells were used as in vitro models to demonstrate the non-cytotoxicity of TAC nCmP. Overall, the designed nCmP have the potential to aid in the delivery of tacrolimus for the treatment of PAH.

Tumor-penetrating aerosol nanocomposite microparticles for the treatment of lung cancer

One of the major public health problems in the Unites States is cancer, where over 1 in 4 deaths is due to this disease each year. As of 2010, lung and bronchus cancers are the leading types of cancer-related deaths in all ages for males and females in the US [1]. The chemotherapeutic treatment most often applied for lung cancer is intravenous paclitaxel (PTX) in the form of Taxol. Methods used to treat respiratory diseases such as asthma and chronic obstructive pulmonary disease (COPD) involve the use of aerosols devices. The first use of aerosolized chemotherapy was reported in 1968 [2]. The advantage of using aerosols treatment is the targeting directly to the lungs and the reduction of systemic side effects provoked by high doses of drug in the body. In this project we have developed a dry powder nanocomposite microparticle (nCmP) aerosol containing PTX-loaded nanoparticles comprised of acetalated dextran (Ac-Dex). Ac-Dex is easily biodegradable and results in fast release of therapeutic agents under acidic conditions as seen in tumor tissues [3]. In addition to the drug, a tumor-penetrating peptide (iRDG) was conjugated to Ac-Dex to help the targeting and penetration into the inner layers of solid tumors. Finally, we formulated nCmPs in mannitol via spray drying. The effectiveness of the complex drug produced was tested in lung cancer cells (A459 lung adenocarcinoma) in two-dimensional (2-D) and three-dimensional (3-D) cell culture studies.

Optimization of Acetalated Dextran–Based Nanocomposite Microparticles for Deep Lung Delivery of Therapeutics via Spray-Drying

Nanocomposite microparticle (nCmP) systems exhibit promising potential in the application of therapeutics for pulmonary drug delivery. This work aimed at identifying the optimal spray-drying condition(s) to prepare nCmP with specific drug delivery properties including small aerodynamic diameter, effective nanoparticle (NP) redispersion upon nCmP exposure to an aqueous solution, high drug loading, and low water content. Acetalated dextran (Ac-Dex) was used to form NPs, curcumin was used as a model drug, and mannitol was the excipient in the nCmP formulation. Box-Behnken design was applied using Design-Expert software for nCmP parameter optimization. NP ratio (NP%) and feed concentration (Fc) are significant parameters that affect the aerodynamic diameters of nCmP systems. NP% is also a significant parameter that affects the drug loading. Fc is the only parameter that influenced the water content of the particles significantly. All nCmP systems could be completely redispersed into the parent NPs, indicating that none of the factors have an influence on this property within the design range. The optimal spray-drying condition to prepare nCmP with a small aerodynamic diameter, redispersion of the NPs, low water content, and high drug loading is 80% NP%, 0.5% Fc, and an inlet temperature lower than 130°C.

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.

Coadministration of a tumor-penetrating peptide improves the therapeutic efficacy of paclitaxel in a novel air-grown lung cancer 3D spheroid model

Three‐dimensional (3 D) cell culture platforms are increasingly being used in cancer research and drug development since they mimic avascular tumors in vitro. In this study, we focused on the development of a novel air‐grown multicellular spheroid (MCS) model to mimic in vivo tumors for understanding lung cancer biology and improvement in the evaluation of aerosol anticancer therapeutics. 3 D MCS were formed using A549 lung adenocarcinoma cells, comprising cellular heterogeneity with respect to different proliferative and metabolic gradients. The growth kinetics, morphology and 3 D structure of air‐grown MCS were characterized by brightfield, fluorescent and scanning electron microscopy. MCS demonstrated a significant decrease in growth when the tumor‐penetrating peptide iRGD and paclitaxel (PTX) were coadministered as compared with PTX alone. It was also found that when treated with both iRGD and PTX, A549 MCS exhibited an increase in apoptosis and decrease in clonogenic survival capacity in contrast to PTX treatment alone. This study demonstrated that coadministration of iRGD resulted in the improvement of the tumor penetration ability of PTX in an in vitro A549 3 D MCS model. In addition, this is the first time a high‐throughput air‐grown lung cancer tumor spheroid model has been developed and evaluated.

In Vitro Pulmonary Cell Culture in Pharmaceutical Inhalation Aerosol Delivery: 2-D, 3-D, and In Situ Bioimpactor Models.

BACKGROUND The use of non-invasive inhaled aerosols for pulmonary drug delivery continues to grow. This is due to the many unique advantages this delivery route offers for the treatment of both local and systemic diseases. The physicochemical properties of the formulated drugs as well as the physiology of the lungs play a key role in both the deposition and absorption of the particles. The airway and the alveolar epithelium are targets for the treatment of respiratory diseases. However, particles have to overcome biological barriers before they reach their target and produce an effect. METHODS In vitro aerosol dispersion performance (i.e. aerodynamic size and aerodynamic size distribution) of inhalable particles is quantified by inertial impaction, as required by regulatory agencies for an investigational pharmaceutical inhalation aerosol formulation to be approved for use in patients as a marketed pharmaceutical product. Using inertial impaction in conjunction with cell cultures of various pulmonary cells in situ as bioimpactors has unique aspects in correlating aerodynamic properties with pulmonary cellular behavior including viability and uptake. These can be as co-culture or in single culture, as 3-D multicellular spheroids or 2-D cellular monolayer using different conditions to grow them, such as air-liquid interface culture (ALI) or in liquid covered culture (LCC). RESULTS evaluation of the currently available in vitro models and the challenges in developing reliable cellular tools to predict the deposition of inhalable particles in the lungs as a function of aerodynamic particle properties is presented in the manuscript. CONCLUSION The mechanistic aerodynamic and biophysical properties of inhaled aerosol particles on the entire respiratory tract at the cellular level based on aerodynamic size and aerodynamic size distribution will be better understood with the development of in vitro methods which are described in this work.

Dry powders based on mucus-penetrating nanocomposite microparticles for pulmonary delivery of antibiotics

Pulmonary antibiotic delivery is increasingly recommended as maintenance therapy for cystic fibrosis (CF) patients with chronic Pseudomonas aeruginosa infection. However, the abnormally thick and sticky mucus present in the respiratory tract of CF patients impairs efficient mucus penetration and limits the range of antibiotics for inhalation treatment. To overcome the obstacles of pulmonary antibiotic delivery, we have developed nanocomposite microparticles (nCmP) for inhalation of antibiotics in the form of dry powder aerosols, which will achieve targeted delivery, rapid mucus penetration, and controlled drug release. Azithromycin and rapamycin loaded nanoparticles were prepared via nanoprecipitation. nCmPs based on mannitol carrier were fabricated by spray drying. Scanning electron microscopy and dynamic light scattering showed 200 nm diameter nanoparticles entrapped in 1 - 2 μm sized microparticles with smooth morphology. In vitro release testing showed that both drug loaded particles have sustained drug release over 12 hours. Redispersity testing indicated that nanoparticles can be well redispersed after being released from mannitol carriers.

Tumor-penetrating acetalated dextran nanoparticles capable of tandem delivery of agents for the treatment of lung cancer

The overall survival rate for patients with lung cancer is still low and many affected patients are not eligible for the first-line treatments (surgery, chemotherapy, and radiation) for non-small cell lung cancer (NSCLC) due to severe side effects. Paclitaxel (PTX) and cisplatin (CDDP) are two of the most commonly utilized drugs in the treatment of NSCLC. These drugs will be encapsulated in tumor-penetrating polymeric nanoparticles (NP) for application in the treatment of lung cancer. Due to the limitations in the NP system itself where particles are often unable to penetrate into the tumor parenchyma to deliver its dose of drug, little has been seen in terms of an increase in clinical outcome for cancer patients treated with nanoparticles although the significant effort that has gone into the study of chemotherapeutic-loaded nanoparticles. Therefore, there is an imperative need for the development of system capable of tumor penetration. Our aim is to develop and optimize peptide-conjugated polymer nanoparticles which can deliver tandem anticancer agents capable of enhancing the targeting and treatment of NSCLC. Acetalated dextran (Ac-Dex) will be used to encapsulate both PTX and CDDP in NPs which in turn will be conjugated with the tumor-penetrating peptide iRGD. This multifunctional particle will not only release PTX and CDDP in tandem, but will be capable of tumor penetration through a mechanism imparted by the peptide. The parameters of the emulsion-based NP system (size, shape, drug loading, peptide-conjugation, cytotoxicity, penetration into NSCLC tumor spheroids, etc.) have been optimized to ensure effective targeting and delivery.

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.