IA 2015 | Lectures
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Tuberculosis is a major cause of illness in low and middle income countries and remains a public health problem in all countries of the world including the Asia Pacific region. Although, there has been substantial progress against the disease since 1990, the gains have been unevenly distributed and, in some countries in our region, there has been very little progress.
Several features of the disease, in particularly, the capacity of the organism to enter a latent phase, make it particularly challenge to control and eradicate it. Major challenges include barriers to case detection and commencement on treatment, the long duration of treatment required to prevent disease relapse, the large pool of latently infected individuals in the world and the difficulty in effectively treating this infection on a population-wide scale, and the emergence of drug-resistant disease that is increasingly difficult and expensive to effectively treat.
Advances in therapy for TB disease and for latent infection in the absence of disease require drugs that can be delivered at an affordable price at scale, that have low toxicity, that are effective against latent organisms, and that are distributed in the lung and systemically. In addition, formulations targeting TB disease need to be active against replicating organisms and to form part of a multi-drug regimen that is designed to prevent the emergence of resistance.
Several methods have been proposed to simulate in vitro the dissolution of inhaled powder drugs in the lungs after inhalation. A first goal of these in vitro methods is to be able to rank various powder formulations according to their different solubilities. A more advanced goal is to already in vitro be able to predict the pharmacokinetic behavior of powder formulations in clinical studies. The purpose of the presentation is to explore some major conditions for simulating the in vivo pulmonary dissolution/absorption process in an in vitro system. An important complication of the of the in vivo dissolution process is that it constitutes only the first step of a multiple-step process starting with deposition of a drug particle on the airway mucosa ending with the released solute appearing in the systemic circulation. The dissolution step is likely rate-controlling only for rather low-soluble substances. In other cases dissolution is a coupled transport process also dependent on the subsequent absorption through the air/blood barrier as well as the removal of the solute with the pulmonary capillary circulation. In order to simulate the in vivo dissolution and absorption process in vitro it is likely that a diffusion barrier is needed in the simulation model. Another design parameter that needs to be established is whether the dissolution medium should perfuse the dissolution compartment as a stirred static batch or perfuse the dissolution compartment in a single-pass dynamic mode. The latter dynamic mode is more akin to the in vivo situation where systemic removal of solute is high and blood returning to the lungs has rather low concentrations of inhaled solutes.
The controversies about electronic cigarettes (EC) or electronic nicotine delivery devices (ENDS) have become greater and more complex with the rapid increase in EC without nicotine (ECON). ECON is not a tobacco product but ECON use is considered as smoking.
The first controversy is whether ENDS is efficacious for smoking cessation or reduction. The question can be answered by randomized controlled trials (RCTs) comparing ENDS with placebo, nicotine replacement therapy (NRT) or medications on smokers with quit intention. Emerging evidence from RCT is likely to show some cessation efficacy. However, it would not resolve the related controversy about ENDS promotion as a smoking cessation aid. At the population level, studies have shown that ENDS use is not related to smoking cessation or is negatively associated with quitting. The main concern is that ENDS may delay or prevent smokers from total abstinence and deterring the use of proven-effective cessation treatments including counselling.
The second controversy is about harm reduction. ENDS deliver less chemicals and carcinogens than cigarettes. ENDS may be a “healthier” alternative if smokers replace cigarettes with ENDS and the nicotine intake remains the same. People against ENDS say that ENDS cannot replace cigarettes at the population level, and are against the marketing message and strategies of “harm reduction”. Studies have reported high dual use rates. Most smokers do not switch from cigarettes to ENDS completely, but use ENDS in smokefree environments to circumvent smokefree laws. ENDS marketing with minimal regulations will lead to more problems before realizing the benefits from “harm reduction”. One problem is the rapid increase in ENDS use worldwide. This re-normalization of smoking behavior may lead to increased cigarette smoking in never smokers and reduced smoking cessation in current smokers.
The third controversy is to what extent ENDS should be regulated. A total ban includes banning of manufacture, trade (import and export), sale and purchase, possession and use of ENDS. Most governments are facing challenges and conflicting views from tobacco control advocates, public health professionals, the tobacco and ENDS and related industry and the public.
The fourth and certainly not the least is about ECON, which are being heavily promoted especially if there is a loophole in the law. ECON with numerous flavors and fashionable designs are claimed to be safe. ECON promotions are similar to cigarette marketing in the pre-ad ban era, targeting young people and smokers with health concerns. The safety and long-term health effects of ECON use is uncertain. Apart from the harmful chemicals detected from ECON, the composition of the smoke being inhaled or released into the environment is mostly unknown and highly variable.
What are NOT controversial are that (1) the ENDS and ECON industry is expanding and the Big Tobacco companies have involved in the ENDS business; (2) the debate about ENDS continues among tobacco control advocates and others; (3) Our understanding of the dynamic situations and future trends is grossly inadequate; and (4) much more monitoring and surveillance, research and evaluation are warranted.
Inhaled insulin has been shown to be safe and effective in over 120 clinical trials in more than 10,000 patients. The second FDA-approved inhaled insulin product, Afrezza® is now available in the US. The first product, Exubera® was approved in the US and Europe in 2006 but withdrawn by Pfizer for business reasons in 2008. Most patients prefer inhaled insulin to injections.
Dance Biopharm is developing an aqueous-based inhaled insulin product, Dance 501, consisting of a sterile, preservative-free, aqueous liquid formulation of recombinant human insulin stored in a separate sterile dispenser and a small handheld electronic inhaler. The inhaler has consumer electronic design that offers a platform for advanced electronic capabilities. The patient dispenses a specific number of drops of a human recombinant insulin formulation from the separate, sterile, multi-dose container (available in different strengths). Exclusive to the delivery of Dance Biopharm’s preservative free formulations, the innovator breath-actuated liquid inhaler silently produces consistently-sized small droplets generated during optimal inspiratory flows providing efficient and reliable delivery of insulin into the lungs as the patient breathes slowly and comfortably for a few breaths. A unique system guides inspiratory flow through dose completion with minimal to no cough reflex.
Based on human gamma scintigraphy studies with a 200 μl payload, the efficiency of delivery from the Dance 501 device to the lungs is 71.5±6.5% (n=6) with an emitted dose at 82.6±3.5% of which 86.5±3.5% is deposited in the lungs with a peripheral to central lung deposition ratio of 2.5±0.8. The product is absorbed more rapidly than injected insulins but also has a longer acting component providing both a bolus and a basal component to its overall pharmacokinetic profile. The blood insulin concentration profile seen following the inhalation from Dance 501 is similar to that produced by a healthy pancreas in normal individuals consuming typical complex meals. We believe Dance 501 will become a user-friendly needle free alternative to currently available therapies.
With the increasing incidence of asthma and COPD (chronic obstructive pulmonary disease) worldwide over recent and coming years there is a growing interest and need for inhaled medications in the Asian market as well as other parts of the globe. This publication describes the development of a salbutamol powder formulation coupled to a simple low cost dry powder inhaler capsule device, Twister®. The study encompassed several stages including selection of suitable excipients grades, capsule types, powder mixing processes and finally acceptable aerosol performance to meet current Chinese Pharmacopeia market requirements.
An initial study to screen for the most suitable excipients involved looking at several lactose grades, including Lactohale versus Respitose grades with different granulometry and obtained either from milling or from sieving. After mixing the Salbutamol sulphate API (active pharmaceutical ingredient) with various lactose blends, their BCU (blend content uniformity) was determined and the mixtures with the most consistent blends were selected for further development.
Optimization of the powder mixing process was studied using a Turbula® low shear blender and several key parameters were investigated including blending speed and time, the use of fines of lactose, the necessity to carry out a pre-blending of lactose, with the most efficient blending conditions for this particular blend being found to be 90 rpm during 60 minutes after a first pre-blending of lactose.
Inhalation powder capsules can play a critical role in capsule based DPI performance and for this development various capsule grades were studied and parameters such as the capsule material were investigated with HPMC capsules determined as being the most compatible for this particular powder and device combination.
Finally various aerosol performance parameters were determined including ease of filling, DCU (dose content uniformity) and FPM (fine particle mass) and the performance of the selected powders, process and capsules were determined and found to be in line with the expectations of Asian regulatory requirements such as the Chinese Pharmacopeia. The performance of the developed powder formulation in combination with the capsule device was studied with different inhalation profiles in order to predict its real use by patients as well as different device resistance levels, and the results were deemed to be acceptable.
The presentation will also cover the advantages and disadvantages of both MDIs and DPIs and some of the key challenges faced when developing these two inhalation dosage forms from the device perspective.
In summary, this case study details how it is possible to develop a targeted low cost DPI device by investigating in sequence all of the key elements that contribute to the critical performance attributes of the final powder in combination with the device. The final stage is to test the developed combination product against the regulatory requirements of the intended market
Most inhaled toxic or therapeutic aerosols (e.g., cigarette smoke or pulmonary drugs) consist of multiple nano-to-micro-scale solid or liquid particles with dissolved or embedded compounds, as well as associated vapors. After inhalation of toxic particles or drug-aerosols, the transport and conversion phenomena may lead to local particle/vapor deposition and possibly species-mass transfer into systemic regions. In the case of micro-dosimetry administration, it is of interest to achieve direct drug-delivery, i.e., optimal targeting to pre-determined sites in the lung. Of special interest in this study are the hygroscopic growth of nano-sized multi-component droplets and the droplet-vapor interactions during their transport through the pulmonary route.
Existing computational fluid-particle dynamics (CF-PD) models neglect the interactions between droplets and vapors during their transport. However, those mechanisms are essential for an improved understanding and a better prediction of local aerosol deposition in human lungs. Focusing on electronic cigarette aerosols, we developed a comprehensive and accurate CF-PD model which is capable of simultaneously analyzing, for any set of inhalation conditions, multi-component droplet-vapor airflow interaction dynamics with evaporation and condensation effects in models of human respiratory systems.
The experimentally validated numerical simulation results include detailed transport, deposition and absorption data for different components with different volatilities in both vapor and liquid forms in the electronic cigarette aerosols, including the coupled dynamics of droplet-size change in an idealized human upper lung airway model. Results indicate that hygroscopic growth of droplets significantly influences their deposition patterns by enhancing inertial impaction and limiting the Brownian motion effect. It also indicates that the ambient relative humidity, initial droplet diameter, and initial droplet composition are parameters that can significantly influence the droplet-growth ratio and hence local deposition pattern. For nicotine, glycerol and PG, the two-way droplet-vapor phase change is not negligible for accurate deposition predictions. Using the standard steady-state puffing parameters (27.5 ml/s for 2 seconds based on the Canadian Intensive Smoking Protocol), 8.40% of glycerol, 14.56% of PG, and 9.88% of nicotine can be absorbed by the human upper lung airway during the consumption of one electronic cigarette. Local deposition and absorption rate for different species in both vapor and liquid forms are also provided. These results can be used for the evaluation of potential health-risk effects of toxic aerosols, as well as the efficacy of drug-aerosol delivery.
Inhalation products are one of the most complicated to develop as a follow-on or generic product. This is due to the fact that equivalence must be shown in terms of the device, in vitro performance, as well as clinical safety and efficacy. This presentation will provide attendees with the understanding of the current and potential future requirements for these products for US FDA, EMA and Health Canada. These requirements have evolved drastically over the last 20 years as the scientific understanding behind the efficacy and safety of inhalation products has progressed. Despite these changes, a greater effort toward regulatory and scientific harmonization is needed, and demonstration of equivalent clinical efficacy and safety needs to become more robust and straightforward. Clinical development case studies will be presented along with common errors and pitfalls to avoid for several inhalation products for asthma and COPD. Practical considerations to better planning, conduct, and analyses of these studies will be presented.
There are a huge range of materials used in a pMDI container closure system. This includes the primary container, the canister and the valve but also secondary packing such as foil over wraps, labels etc. This presentation discusses briefly the main points to consider when selecting materials and what impact these decisions could have. These include:
• Physical properties
o Simple understanding of materials. More detail on the main types of materials used in pMDIs will be discussed. These include plastic rubbers and metals.
o Manufacturing considerations. For pMDIs there can be very tight tolerances on manufacturing components to achieve the required dose. Selecting materials that can be manufactured to these tolerances is very important.
• Supplier information
o This introduces the information issued by the IPAC-RS on the baseline requirements that suppliers of materials for inhalation devices should supply. It also covers aspects of general supplier information, e.g. quality system, change management etc.
• Chemical compatibility
o Sorption of API. This can be very important for low dose pMDI formulations.
o Extractables and leachables. What are, where do they come from and why are the regulators so interested in them, especially for pMDIs.
o Current regulatory expectations. How the guidances and regulations have changed and adapted over time along with the differences between EU and US.
o Finally the limits around extractables and leachables, how low to go in determining the identification of extractables and leachables.
Throughout the lifecycle of a pharmaceutical product decisions are made to select, qualify and control the materials that are in direct contact with the dosage form. Orally inhaled and nasal drug products (OINDP) are considered high risk dosage forms due to the route of administration and likelihood of component-dosage form interaction. OINDP packaging and delivery systems consist of a variety of polymeric, elastomeric and metal critical components, some of which are in direct contact with the drug formulation and/or patient tissues during storage and use of the drug product, and most of which are integral and necessary to the product’s proper functioning. OINDP manufacturers and global regulatory authorities therefore hold the materials and components used to a high standard of quality. A systematic approach to developing a robust risk management program for a given product has been developed. The IPAC-RS Materials working group has developed specific chemical risk evaluation tools. Examples of their use in developing comprehensive studies to assess material safety and compatibility will be discussed.
The presentation provides an insight into Boehringer Ingelheim’s experiences with extractables and leachables for inhalation products.
It is based on the experiences that are related to the development of a dry powder inhalation system, where several polymer components had to be selected and to be qualified. In addition, the experiences have been confirmed with other inhalation product development.
Based on this case study, the following learnings are shared:
- It is essential to have an open exchange with the supplier of the polymer brand.
- Written long-term supply agreements that exactly define the quality of the polymer brands are important and need to be adhered to.
- Changes in the supply chain (including N-1, N-2 suppliers) are likely to affect the extractable profile. Examples are shown where such changes in the supply chain resulted either in major additional extractable peaks or in significant changes in the amounts of known extractables. In such cases, the changes in the supply chain cannot be accepted without further qualification work.
- To characterize the extractable and the leachable profile comprehensively, components made from many different polymerization campaigns must be investigated.
- Extractables may convert into unexpected leachables. This may be due to reaction with the formulation and lead to (e.g.) hydrolytic fragmentation; or it may be a reaction with oxygen. Examples are Ph. Eur. plastic additive 09 and Ph. Eur. plastic additive 12, as identified in Ph. Eur. monograph 3.1.13.
- Leachables may originate from polymers (including sealing polymers in blisters), but also from manufacturing equipment (e.g. metal surfaces, filters for liquids).
- Specifications for extractables and leachables will be diverse for a global product. Often, US-FDA has the most detailed and most stringent specifications in order to fully control the supply and manufacturing chain and to avoid patient exposure to unqualified leachables. The European Union, in contrast, may rely on the Ph. Eur. monographs on plastic materials.
The presentation will provide examples for different approaches to specification setting. For GMP reasons, it is recommended to apply the most stringent specifications worldwide.
- Toxicological assessment of extractables and of leachables should consider not only maximum observed levels but maximum specified levels. The need for a toxicological qualification depends on the daily uptake of extractables and leachables. The daily uptake is a function of the number of doses administered per day, the mass of a dose and the concentration of the leachable in a dose.
Extractables and leachable work never stops. Any lifecycle change (e.g. due to complaints, stop of polymer brand production, or changes in the supply chain including N-1, N-2 supplie
Lung infections with tuberculosis (TB), non-tuberculous mycobacteria (NTM), melioidosis and Q-fever are a public health problem globally and also in several Asian countries, as well as infections with Pseudomonas aeruginosa (PA) that cause higher incidence of morbidity and mortality in non-cystic fibrosis (CF) bronchiectasis (NCFBE), chronic obstructive pulmonary diseases (COPD), and, in Japan, in diffuse panbronchiolitis. Liposomes have the potential to improve inhaled antibiotic therapy by providing a sustained high-concentration of antibiotic in the lungs and thus reducing the need for multiple administrations each day, improving the airway tolerability, and reducing systemic exposure to minimize the likelihood and severity of systemic side effects. The liposomal encapsulation significantly enhances the antibiotic activity in the biofilms formed by organisms such as PA and NTM. Furthermore, microorganisms such as mycobacteria can be harboured by pulmonary macrophages and liposomal encapsulation enhances the phagocytosis and killing of these intracellular infections. Inhaled once-daily formulations of liposomal ciprofloxacin, Lipoquin® (Aradigm), and a combination of free ciprofloxacin and Lipoquin, termed Pulmaquin® (Aradigm), have been evaluated in a number of animal models of infectious diseases and in human clinical trials with positive outcomes: tularemia, plague, Q-fever, CF, and NCFBE.
Different release rates may be preferred dependent upon the host, the pathogen and its location within the lungs. Therefore, we have developed strategies to modify the release rate profile of the Lipoquin formulation without changing the lipid composition: 1. by addition of surfactant after osmotic swelling in a hypotonic environment to create a more permeable membrane; and 2. by freeze-thaw to transform the drug into a nanocrystalline state inside the liposomes. These formulations were characterized by cryo-TEM analysis, pH, dynamic light scattering, in vitro release (IVR) assay, aerosol particle size distribution and the effect of mesh nebulization (PARI eFlow) on functionality.
The addition of certain types of non-ionic surfactants to Lipoquin under specific conditions led to formulations which had significantly faster release properties than Lipoquin. The mean vesicle size increased only modestly and the formulations retained their vesicular structure. The formulations also retained their properties after nebulization and were stable after 2 years refrigerated storage.
The addition of sucrose or trehalose to the liposomal ciprofloxacin formulation stabilized the liposomes to freeze-thaw. After freeze-thaw, ciprofloxacin in the liposome vesicles formed nanocrystals which resulted in significantly slower release profiles. The shape and length of the nanocrystals and the release profiles of ciprofloxacin from the formulation could be adjusted by varying the amount of surfactant. These formulations were robust to nebulization, with retention of nanocrystals within the vesicles, and formed aerosols with a volume median diameter of 3.8-3.9 µm and geometric standard deviation of 1.7.
These new methods have the potential to personalize therapy for individual patients or groups of patients by modulating the release properties of liposomally encapsulated drugs.
Pirfenidone (PFD) is clinically used for treatment of idiopathic pulmonary fibrosis; however, previous clinical studies demonstrated that orally-taken PFD often caused phototoxic skin responses (51.4% of patients) and severe gastrointestinal (GI) dysfunctions (49.1%). Dry powder inhalation system of PFD might be efficacious for controlling distribution of PFD in skin and GI, possibly leading to reduced systemic side-effects.
The present study aimed to develop novel respirable powder formulation of PFD (PFD-RP) for minimizing systemic exposure of PFD and maximizing topical effects in pulmonary tissues. To clarify the phototoxic potential, photobiochemical properties of PFD were examined with focus on UV absorption, generation of reactive oxygen species (ROS), lipid peroxidation, and DNA impairment. In vivo phototoxicity testing was conducted after oral administration of PFD (30–160 mg/kg). PFD-RP was prepared with a jet-mill, and physicochemical properties were characterized in terms of appearance, particle size distribution, and in vitro inhalation performance. In vivo pharmacological effects of PFD-RP were also assessed in experimental rat asthma/COPD models. Pharmacokinetic and safety studies on PFD formulations were carried out after oral and intratracheal administrations. PFD exhibited intense UVA/B absorption with molar extinction coefficient of 1,290 M-1･cm-1 and generated ROS, including singlet oxygen and superoxide, after exposure to simulated sunlight (Xe lamp, 250 W/m2). These photochemical data suggested the potent photoreactivity of PFD, which might be key trigger for phototoxic reactions. Based on the in vitro phototoxic assessments, PFD exhibited potent photoirritant risk, although in vitro photogenotoxic reaction of pirfenidone was negligible. Photostability testing demonstrated that PFD-RP, as well as PFD powder, were markedly more photostable than PFD solution. Laser diffraction and cascade impactor analysis of newly developed PFD-RP, consisting of jet-milled PFD and lactose carriers, suggested its high dispersion and in vitro inhalation performance. Inhaled PFD-RP (0.3 mg-PFD/rat) could suppress antigen-evoked inflammatory events in experimental asthma/COPD model rats as evidenced by reduced infiltration of inflammatory cells and decrease of inflammation-related biomarkers in the pulmonary tissues. The PFD-RP at a pharmacologically effective dose could also reduce phototoxic skin responses and GI dysfunctions of PFD because of its lower systemic exposure than that after oral administration of PFD at the phototoxic dose (160 mg/kg). From these findings, the PFD-based inhalation therapy would be an attractive alternative to current oral therapy of PFD with a better safety margin for clinical treatment of idiopathic pulmonary fibrosis.
Colistin is the last-line therapy for the treatment of lung infections caused by multi-drug resistant Gram-negative ‘superbugs’. Unfortunately intravenous administration of colistin can cause severe nephrotoxicity in up to 60% patients. This study aimed to develop an inhaled combination dry powder formulation of colistin and rifapentine for the treatment of respiratory infections.
The combination formulation was produced by spray-drying rifapentine particles suspended in an aqueous colistin solution. The combination dry powder had enhanced antimicrobial activities against planktonic and biofilm cultures of Pseudomonas aeruginosa, with both minimum inhibitory concentration (MIC) and minimum biofilm inhibitory concentration (MBIC) values (2 and 4 mg/L, respectively) being half that of pure colistin (MIC 4 mg/L and MBIC 8 mg/L) and one-sixteenth that of pure rifapentine (MIC 32 mg/L and MBIC 64 mg/L). High aerosol performance, as measured via an Aerolizer device, was observed with emitted doses > 89% and fine particle fraction (FPF) total > 76%. The proportion of submicron particles of rifapentine particles was minimised by the attachment of colistin which increased the overall particle mass and aerodynamic size distribution. Using the spray-drying method described here, stable particles of amorphous colistin and crystalline rifapentine were distributed homogenously in each stage of the impinger. Unlike the colistin alone formulation, no deterioration in aerosol performance was found for the combination powder when exposed to a high relative humidity of 75%. In our previous study, surface coating by rifampicin contributed to the moisture protection of colistin. Here, a novel approach with a new mechanism was proposed whereby moisture protection was attributed to the carrier effect of elongated crystalline rifapentine particles which minimised contact between hygroscopic colistin particles. This inhaled combination antibiotic formulation with enhanced aerosol dispersion efficiency and in vitro efficacy could become a superior treatment for respiratory infections.
Nowadays, in order to overcome the poor patient compliance of conventional injection administration, suspension pressurized metered dose inhalers (pMDIs) have been introduced into the frontier of protein/peptides administration. However, since water, organic solvent and hydrofluoroalkanes (HFAs, the widely used propellant) act as co-solvent for majority of suspension pMDIs containing water-soluble protein/peptides, it is vital to ensure the drugs could disperse homogeneously and meanwhile keep away from hydrolysis in the system. Otherwise, the preparation would be physically and/or chemically unstable, leading to poor bioavailability and even side-effects.
To achieve these two aims (homogeneous dispersion and non-hydrolysis), the authors chose the water-soluble yet susceptible-to-hydrolysis peptide drug salmon calcitonin (sCT) whose major indication is osteoporosis as a model drug, and explored a novel bottom-up method (i.e., lipid vesicle inversion) to prepare suspension pMDIs containing anhydrous reverse micelle based water-soluble protein/peptides nanoparticles (ARM-NPs).
Tertiary butanol-dissolved lecithin (oil phase) was added into the sCT/ lyoprotectant aqueous solution (water phase) to form double molecular layers lipid vesicles. After freeze-drying the lipid vesicles, the obtained lyophilized powder was inverted to ARM-NPs due to the addition of anhydrous ethanol. It was observed that: the ARM-NPs had uniform shape and dispersed homogeneously in the system (particle size: 147.7±2.00 nm; polydispersity index: 0.152±0.021); Moreover, it had an anhydrous drug core surrounded by lipids, perfectly preventing sCT from hydrolysis. Besides, a hypothesis was raised to explain the mechanism of the formation of ARM-NPs on the basis of differential scanning calorimetry and phase diagram analysis. Ultimately, the ARM-NPs were filled by HFAs as a propellant to prepare the suspension pMDI containing sCT.
Preparation evaluation, namely 24h-stability, appearance, total presses per bottle, content of drug per press, aerosol distribution and drug leakage rate was conducted to the pMDI. The results indicated that the suspension pMDI was stable, uniform and opalescent, with satisfactory deposition ratios, which was a promising pulmonary drug delivery system of sCT. Nevertheless, further investigation about the pharmacodynamics and pharmacokinetics profile of the preparation was necessary before promoting this novel method into other water-soluble protein/peptides and therefore applying clinically.
The World Health Organisation estimates 9 million people developed Tuberculosis (TB) in 2012 globally, with 1.3 million fatalities in the same year. Current approaches involve the administration of antibiotics, either using oral tablets or intravenous infusions for long periods of time. These approaches result in risk of side effects, poor patient compliance, and thus limited therapeutic effectiveness. The lung offers a fantastic portal for both the local and systemic delivery for a number of medicines and addresses patient needs for therapies that are quick and convenient to administer.
Rifapentine is a semisynthetic cyclopentyl rifamycin antibiotic, generally prescribed for the treatment of TB, with a serum half-life several times longer than rifampicin, which can provide less frequent dosing in comparison with standard dosing regimens. Compared to the more commonly used rifampicin, rifapentine is several times more active against Mycobacterium tuberculosis, both in vitro and in vivo. Unfortunately, the shared mechanism of action among rifamycins leads to extensive cross-resistance among members of this class of drugs. There is evidence that increased dosing frequency of rifapentine has the potential to substantially accelerate treatment of active TB to just three months or less. Studies performed in mice found positive results, while the lack of treatment shortening in guinea pigs foreshadowed similar failures in a related multi-centre human clinical trial.
There are several factors that may explain the limited efficacy of rifapentine in humans. These include high rifapentine plasma protein binding (98%), variation in rifapentine bioavailability with food intake and induction of its own metabolism. In our study, a novel dry powder formulation of rifapentine was produced and its pharmacokinetics in healthy mice was compared after intratracheal (IT) and intraperitoneal (IP) administration. Results showed that inhaled delivery of rifapentine, used as a primary or adjunct therapy, has the potential to selectively enhance therapeutic efficacy at the pulmonary site of infection whilst minimising systemic exposure and related toxicity.
One of the strategies of tuberculosis treatment is to deliver high doses of drug to the lung. Little data is available on the effects of administering these high doses of drug to the lung and their dissolution in the lung. Inhaled drugs first meet a lung surfactant monolayer which is spread on an aqueous sub-phase overlying the respiratory epithelium. While undissolved drug particles undergo alveolar phagocytosis, the drug needs to cross the lung surfactant barrier and then dissolve in the very small amount of lung fluid in order for it to be absorbed. This aqueous fluid contains high concentration of lung surfactants which are stored as lamellar bodies after secretion by type II alveolar cells. These lamellar bodies then unravel in the alveolar space, forming tubular myelins which are similar to that of multi-lamellar liposomes. The principal component of lung surfactant is 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). It is not clear how lung surfactants interact with high doses of anti-TB drugs and influence drug dissolution.
Using a Langmuir Blodgett trough, the interactions of intraconazole (INZ) and rifanpicine (RIF) with lung surfactant monolayer were determined. These drugs were present in the aqueous sub-phase at concentrations up to 1 mg/ml. The collapse pressure (π) of the DPPC monolayer dropped significantly from 58 mN/m to 38 mN/m when INZ was present at a concentration of 1.0 mg/ml in the subphase. Although similar collapse pressure decrease was observed with RIF, the collapse pressure-area isotherm expansion and the increase in the area per DPPC molecule were greater with RIF than INZ. The area per DPPC molecule at solid phase increased from 51 Å2 to 80 Å2 at 0.5 mg/ml of RIF. In contrast, the area per DPPC molecule initially increased to 71 Å2 at 200 µg/ml of INZ and then decreased to 68 Å2 at 1 mg/ml of INZ.
The mechanisms of interaction of these two drugs were different. While RIF was speculated to participate in monolayer formation, INZ may have interacted with DPPC causing them to leave the monolayer. When INZ at 1 mg/mL was centrifuged with 500 µg/mL DPPC in water, only ~ 4% of INZ was found in the supernatant layer indicating a strong interaction between INZ and DPPC. The dissolution of INZ generally increased with increasing DPPC concentration and in the presence of multi-lamellar vesicles of DPPC when the dissolution study was conducted by Franz Diffusion cells using different concentrations of DPPC (0.01 %w/v, 0.025% w/v, 0.05%w/v) and multi-lamellar liposomes(0.05% w/w) in the dissolution media, phosphate buffer medium (PBS, pH = 7.4).
The study concluded that the presence of high concentration of anti-TB drugs significantly decreases the collapse pressure of the DPPC monolayer which may facilitate the drug particles to reach the aqueous sub-phase by disrupting the lung surfactant monolayer barrier. Lung surfactant and its nanostructures such as liposomes help increase the dissolution of drugs probably by solubilisation.
The goal of this effort was twofold: to prepare rifampicin (RFP)/PLGA nanoparticles-containing microspheres (MS) using the 4-fluid nozzle spray drier and investigate in vivo uptake of RFP by alveolar macrophages, and to develop a nanoparticle delivery system of a highly water-soluble OCT313 against alveolar macrophages using a novel two-solution mixing spray nozzle.
PLGA was used as a carrier for nanoparticles. Mannitol (MAN) was used as a carrier for the MS. The RFP/MAN MS and (RFP/PLGA)/MAN MS were administrated intratracheally to rats and the in vivo retention of RFP in the lung and the uptake of RFP in alveolar macrophages were studied. Fluorescent calsein nanoparticles coated with surfactant ER-290 (SOLID) were prepared by a water-in-oil emulsion method. SOLID-nanoparticles containing mannitol microspheres (SOLID/MAN) were prepared by a spray drying method using a two-solution mixing spray nozzle (solution 1: the SOLID suspension in isopropanol, solution 2: MAN aqueous solution). Fluorescence intensity of calsein in alveolar macrophages was measured. The SOLID/MAN microspheres using OTC313 were also prepared.
In the case of (RFP/PLGA)/MAN MS, RFP remained in the lung after 4hr. The amount of RFP uptake in alveolar macrophages after 4 hr was larger than that after 1 hr. In an in vivo study of uptake of calsein after pulmonary administration of the SOLID/MAN microspheres by alveolar macrophages, the uptake of calsein from the SOLID/MAN microspheres was larger than that from the calsein microspheres prepared by spray drying a calsein aqueous solution.
Pulmonary tuberculosis (TB) is a leading killer worldwide and multi drug resistant (MDR) tuberculosis is reaching epidemic proportions. Newer antibiotics are scarce and are often more expensive and more toxic. Due to emergence of MDR and XDR TB, there is an urgent need for new drugs and delivery systems for TB. Nitric oxide possesses wide-ranging antimicrobial activity and is effective against pathogenic mycobacterium in in-vitro studies. Nitric oxide is a free radical which is highly unstable and reactive with a short half-life (3-5 sec in vivo) making it challenging to introduce it to a biological system in a controlled manner for therapeutic purpose. The potential of NO as an antimicrobial agent is dependent on the availability of an appropriate delivery system which can carry and deliver the required payload of nitric oxide to the targeted site. NO donors may provide an alternative approach but to stabilize NO generated from bare compounds is difficult. Bare NO-donor compounds cannot provide a sustained supply of NO over a long time span. MP containing NO donors may provide an unconventional novel approach for sustained delivery of NO and may represent a novel paradigm in TB therapy.
We developed and aerodynamically characterized clinically used nitric oxide donors such as Isosorbide Mononitrate (ISMN), Sodium Nitroprusside (SNP) and Diethylenetriamine NO adduct (DETA/NO) with and without standard anti-TB drugs [Isonizid and Rifabutin] in inhalable PLGA [poly(lactic-co-glycolic acid)] microparticle (MP) delivery systems to provide sustained delivery of NO to macrophages. The efficacy of MP in a mycobacterium infected macrophage and the therapeutic efficiency in a TB infected animal model was determined.
MPs were produced using a co-current, laboratory-scale spray dryer. Inhalable MPs were prepared suitable for deep lung deposition, i.e. within a respirable size range (1-10 μm) with an aerodynamic diameter of ~2.5 μm. The in vitro deposition of the DPI MPs containing ISMN, SNP and DETA/NO-MPs was investigated using a Mercer seven-stage cascade impactor. The activity of MP containing NO donors against intracellular Mtb was discernible in vitro in a dose and time dependent manner without significant toxicity. DETA-MP, equivalent to a drug concentration of 10, 50 and 100 μg/ml, showed significant anti mycobacterial effects after 6, 12, 24, 48 and 72 h of incubation. Similarly, SNP-MP demonstrated efficient killing at the highest dose of 100 μg/ml while ISMN-MP showed little anti mycobacterial activity at the same dose. The lower doses (10 and 50 μg/ml) of both MPs and soluble drugs did not show significant inhibition. The animal efficacy results reveals that MPs containing DETA-MP exerted significant anti TB activity against virulent Mtb in vivo but efficacy was feeble with SNP-MP as compared to oral and inhaled anti-tuberculosis drug therapy. Results suggest that at very low concentrations, NO releasing MPs did not show efficacy by direct free radical mechanism but instead activate cells via phagosome-lysosome fusion.
These results display the advantage of pulmonary delivering of NO via MP for anti-TB application in combination with DOTS therapy.
Magnetic nanoparticles (MNPs) have approved medical uses as a contrast agent for MRI and induction of hyperthermia for the treatment of testicular cancer. Taken together, MNP could serve as a mode of diagnosis as well as therapy, i.e. a theranostic agent. Indeed, most experimental efforts have been directed towards the use of iron oxide particles for site specific cancer detection and therapy. MNPs are typically composed of small crystallites (<100 nm), consisting of a single magnetic domain. With an applied magnetic field, the induced magnetization is reversible, which is characteristic of superparamagnetic behavior. The small size and associated random diffusion also creates a fluctuating magnetic field, which makes MNPs effective contrast agents for MRI. The application of an alternating magnetic field causes MNPs to undergo heating, through Brownian relaxation and/or Néel relaxation. This heat generation is sufficient to increase the temperature in vivo and thereby achieve cell death, which is referred to as magnetic hyperthermia.
Here, we present our studies involving the use of MNP for the treatment of lung cancer in animal models. EGFR-targeted, inhalable SPIO nanoparticles were synthesized and characterized for targeting lung tumor cells as well as for magnetic hyperthermia-mediated antitumor efficacy in a mouse orthotopic model of NSCLC. Our results show that EGFR targeting enhances tumor retention of SPIO nanoparticles. Further, magnetic hyperthermia treatment using targeted SPIO nanoparticles resulted in significant inhibition of in vivo lung tumor growth. We also have shown that the extent and mechanism of hyperthermia-induced cell kill is highly dependent on the aggregation state of SPIO nanoparticles. Well-dispersed nanoparticles induced apoptosis, similar to that observed with conventional hyperthermia. Sub-micron size aggregates, on the other hand, induced temperature-dependent autophagy through generation of oxidative stress. Micron size aggregates caused rapid membrane damage and acute cell kill, likely due to physical motion of the aggregates in alternating magnetic field. MNPs, having a magnetite core of 12 nm, were used to induce magnetic hyperthermia in A549 and MDA-MB-231 tumor cells. Multiple assays for cancer stem cells (CSC), including side population phenotype, aldehyde dehydrogenase expression, mammosphere formation, and in vivo xenotransplantation, indicated that magnetic hyperthermia reduced or, in some cases, eliminated the CSC subpopulation in treated cells. Our studies show that magnetic hyperthermia has pleiotropic effects, inducing acute necrosis in some cells while stimulating reactive oxygen species generation and slower cell kill in others. More recently, we have examined the properties of asymmetric MNP, which had aspect ratios of 1.8, 3.4 and 6.6, and all displayed magnetic hysteresis with corresponding saturation magnetization values of 65, 47, and 26 emu/g respectively. While these particles are not satisfactory for magnetic hyperthermia, they have promise for causing cell death by a mechanical mechanism involving physical rotation or vibration of the particles. Such particles would be effective MRI contrast agents, and a single particle could conceivable induce cell, thereby serving as an effective theranostic agent.
Curcumin (diferuloylmethane), the active constitute from turmeric extract, has gained momentum as the next generation of “magic drug” for the treatment of various chronic conditions such as cardiovascular diseases, neurodegenerative disorders and cancer diseases. Curcumin possesses high selectivity towards cancer cells compared to healthy cells, which results in negligible systemic toxicity even at higher administration doses. However, due to the rapid clearance and low bioavailability of curcumin in vivo, its application for cancer therapies has been hampered. Our team has used a solvent and anti-solvent precipitation method to synthesize curcumin nanoparticles (Cur-NPs) with high mono-dispersity and low agglomeration characteristics. By controlling preparation conditions (i.e. concentration of stabilizers and temperature), nanoparticles with tunable size distributions (30 to 200 nm) have been synthesized. In addition, the physical and chemical stability of Cur-NPs such as particle size, polydispersity and curcumin content was unchanged after 4 weeks of incubation. The degradation of Cur-NPs was minimal (<10%) compared to raw curcumin (>80%) after 24 hrs. Aerosol deposition results using the next generation impactor (NGI) showed that Cur-NPs with different particle sizes could be aerosolised with fine particle fraction (FPF) of >40%.
The in vitro study showed that engineered Cur-NPs had higher cytotoxicity effect against lung cancer cell line (A549 and Calu-3) compared to raw curcumin. The cytotoxicity effect was size-dependent, with the smaller nanoparticles exerting superior activity. Higher cytotoxic effect was further supported by higher internalization of Cur-NPs into the cancer cells. Qualitative results from confocal laser microscopy showed that Cur-NPs (30 nm) was internalized specifically into the nucleus and cytoplasm. Larger Cur-NPs were less efficiently internalized by the cells and were accumulated preferentially in the cytoplasm. Interestingly, curcumin either in the nanoparticle or solubilised form were not toxic to healthy lung cells (Beas-2B).
The effects of Cur-NPs in regulating the inflammation and oxidative stress in alveolar macrophage (AM) were also investigated, as these factors play an important role in promoting lung cancer. The different surface charges present on the Cur-NPs affected the cytotoxicity in AM. Positively charged Cur-NPs induced higher cytotoxicity compared to negatively and neutral particles. The same was observed for the anti-oxidant activity, which include nitric oxide production and radical scavenging activity. For the anti-inflammatory activity of Cur-NPs, particles with positive charges were the most effective followed with negative and neutral charged particles. The observed differences in activities could be due to the varying affinities of the AM towards Cur-NPs. Qualitative and quantitative intracellular uptake demonstrated that positively charged Cur-NPs had the greatest interaction with AM. As a conclusion, using an appropriate particle engineering and formulation approach, Cur-NPs could potentially be developed as the next–generation drug for lung cancer treatment.
Lung cancer is an important worldwide public health problem. Surgical resection is the preferred treatment for Stage I and Stage II lung cancers; whereas more advanced lung cancers (Stages III and IV) are treated with chemotherapy and/or radiation therapy. Despite recent advances in diagnosis and treatment, conventional methods of treatment fail to provide a cure in more than 80% of patients with lung cancer.
Within the changing paradigm of lung cancer diagnosis and treatment, chemotherapy has remained as an important part of the therapeutic armamentarium. In addition to its established role for treatment of advanced (Stages III and IV) lung cancers, adjuvant systemic chemotherapy is also being employed after surgical resection of early (Stage II) lung cancers. The major disadvantage of systemically administered chemotherapy is that drug toxicity limits the dose that could be safely administered, and cancers recur because they become refractory to the effects of the drugs.
In several cancers, regional chemotherapy has been successfully employed to administer higher and more effective doses of chemotherapy. Aerosolized chemotherapy provides a similar regional approach to the treatment of lung cancer, especially for non-small cell lung cancer (NSCLC) and multifocal or metastatic tumors. Aerosolized chemotherapy has the potential to increase exposure of tumors to higher levels of drugs while reducing their systemic toxicity and represents a fertile area for further investigation. Several methods of targeting inhaled agents to the site of the tumor(s) within the lung are being investigated to protect the normal lung tissues from local toxic effects. These approaches include: regional targeting by introduction of aerosol at different volumes of inhalation; enhanced condensational growth; utilizing intracorporeal nebulization catheters for aerosol delivery; use of external magnetic fields; or targeting receptors on cancer cells with specific ligands. Concurrent development of liposomal and nanoparticle formulations seeks to improve the safety and residence time of chemotherapeutic agents within the lung. With these techniques, chemotherapy levels achieved in the lungs could be safely increased several-fold while minimizing systemic toxicity.
Early phase clinical trials have shown that inhaled chemotherapy is effective for treatment of lung cancers, and it could play an important role as an adjunct to other forms of treatment. Further investigations of several modalities that target aerosols to lung cancers, safeguard the health of care providers, and protect the environment are needed to establish optimal protocols for inhaled chemotherapy.
Aerosol Isoprenaline and sodium cromoglycate dry powder inhaler were available in Chinese market since early 1960s. After more than 50 years of development, inhaled drugs market in China has grown into a period of steady increasing. The main reasons are:
1. The prevalence rate of asthma and COPD in China is keep increasing. For instance, Asthma prevalence rate of Children in Shanghai is now as high as 7.57%. The sales volume of anti-asthmatic drugs was 2.051 billion (RMB) in 2009, and increased to 6.174 billion in 2014.
2. Physicians and patients have more awareness of inhalation therapy, and are willing to accept it as a measure to prevent and control asthma and COPD.
3. Due to the promotion of GINA and GOLD, number of patients receiving inhaled corticosteroids during remission of asthma and COPD are increasing gradually.
4. In the process of eliminating CFC to protect the ozone layer, acceptance level of dry powder inhaler is rising up.
5. With the popularity of atomization devices, atomizing inhalation therapy has been accepted by many hospitals/institutions, patients and their families. Nebulizer market is growing rapidly.
6. Individual income increased, and along with the improvement of health care system, inhalation therapy is now affordable to a larger number of population.
Due to the atmospheric pollution (which is difficult to control in short term), sluggish Tobacco control, and invasion of foreign allergens, we can expect that the prevalence rate of asthma and COPD in China will keep demonstrating a upward trend. Now, inhaled drug accounts for only 55% of anti-asthmatic drugs in China, while the world level is 80%. Factors like GINA and GOLD promotion, increasing popularity of inhalation drug knowledge and medical coverage improvement, etc. all will lead inhaled drug market to continue expanding.
In order to keep pace with the development of preparations for inhalation and ensure the quality of the products, relative general chapters in the upcoming edition of the Chinese Pharmacopoeia have been revised. ACI and NGI methods are introduced to conduct aerodynamic assessment of fine particles. Test for the uniformity of delivered dose is going to be required in the routine test. In this presentation all these new requirements for the quality control of preparations for inhalation will be discussed, and also other requirements during the development of preparations for inhalation will be reviewed.