Identification and Characterization of Critical Processing Parameters in the Fabrication of Double-Emulsion Poly(lactic-co-glycolic) Acid Microparticles.

In the past several decades, polymeric microparticles (MPs) have emerged as viable solutions to address the limitations of standard pharmaceuticals and their corresponding delivery methods. While there are many preclinical studies that utilize polymeric MPs as a delivery vehicle, there are limited FDA-approved products. One potential barrier to the clinical translation of these technologies is a lack of understanding with regard to the manufacturing process, hindering batch scale-up. To address this knowledge gap, we sought to first identify critical processing parameters in the manufacturing process of blank (no therapeutic drug) and protein-loaded double-emulsion poly(lactic-co-glycolic) acid MPs through a quality by design approach. We then utilized the design of experiments as a tool to systematically investigate the impact of these parameters on critical quality attributes (e.g., size, surface morphology, release kinetics, inner occlusion size, etc.) of blank and protein-loaded MPs. Our results elucidate that some of the most significant CPPs impacting many CQAs of double-emulsion MPs are those within the primary or single-emulsion process (e.g., inner aqueous phase volume, solvent volume, etc.) and their interactions. Furthermore, our results indicate that microparticle internal structure (e.g., inner occlusion size, interconnectivity, etc.) can heavily influence protein release kinetics from double-emulsion MPs, suggesting it is a crucial CQA to understand. Altogether, this study identifies several important considerations in the manufacturing and characterization of double-emulsion MPs, potentially enhancing their translation.

In silico identification and synthesis of a multi-drug loaded MOF for treating tuberculosis.

Conventional drug delivery systems have been applied to a myriad of active ingredients but may be difficult to tailor for a given drug. Herein, we put forth a new strategy, which designs and selects the drug delivery material by considering the properties of encapsulated drugs (even multiple drugs, simultaneously). Specifically, through an in-silico screening process of 5109 MOFs using grand canonical Monte Carlo simulations, a customized MOF (referred as BIO-MOF-100) was selected and experimentally verified to be biologically stable, and capable of loading 3 anti-Tuberculosis drugs Rifampicin+Isoniazid+Pyrazinamide at 10% + 28% + 23% wt/wt (total > 50% by weight). Notably, the customized BIO-MOF-100 delivery system cleared naturally Pyrazinamide-resistant Bacillus Calmette-Guérin, reduced growth of virulent Erdman infection in macaque macrophages 10-100-fold compared to soluble drugs in vitro and was also significantly reduced Erdman growth in mice. These data suggest that the methodology of identifying-synthesizing materials can be used to generate solutions for challenging applications such as simultaneous delivery of multiple, small hydrophilic and hydrophobic molecules in the same molecular framework.

Microfluidic Systems For Manufacturing of Microparticle-Based Drug-Delivery Systems: Design, Construction, and Operation.

Particles synthesized from biodegradable polymers hold great potential as controlled drug delivery systems. Continuous flow platforms based on microfluidics offer attractive advantages over conventional batch-emulsification techniques for the scalable fabrication of drug-loaded particles with controlled physicochemical properties. However, widespread utilization of microfluidic technologies for the manufacturing of drug-loaded particles has been hindered largely by the lack of practical guidelines toward cost-effective development and reliable operation of microfluidic systems. Here, we present a framework for rational design and construction of microfluidic systems using commercially available components for high-throughput production of uniform biodegradable particles encapsulating drugs. We also demonstrate successful implementation of this framework to devise a robust microfluidic system that is capable of producing drug-carrying particles with desired characteristics. The guidelines provided in this study will likely help broaden the applicability of microfluidic technologies for the synthesis of high-quality, drug-loaded biodegradable particles.

Local induction of regulatory T cells prevents inflammatory bone loss in ligature-induced experimental periodontitis in mice.

Periodontitis (periodontal disease) is a highly prevalent disease, affecting over 65 million adults in the United States alone. Characterized by an overburden of invasive bacteria, gum inflammation and plaque buildup, over time, these symptoms can result in severe loss of gingival tissue attachment, bone resorption and even tooth loss. Although current treatments (local antibiotics and scaling and root planing procedures) target the bacterial dysbiosis, they do not address the underlying inflammatory imbalance in the periodontium. In the healthy steady state, the body naturally combats destructive, imbalanced inflammatory responses through regulatory pathways mediated by cells such as regulatory T cells (Tregs). Consequently, we hypothesized that local enrichment of regulatory lymphocytes (Tregs) could restore local, immunological homeostasis and prevent the main outcome of bone loss. Accordingly, we locally delivered a combination of TGFß, Rapamycin, and IL2 microspheres in a ligature-induced murine periodontitis model. Herein, we have demonstrated this preventative treatment decreases alveolar bone loss, increases the local ratio of Tregs to T effector cells and changes the local microenvironment's expression of inflammatory and regenerative markers. Ultimately, these Treg-inducing microspheres appear promising as a method to improve periodontitis outcomes and may be able to serve as a platform delivery system to treat other inflammatory diseases.

Advances in controlled drug delivery to the sinonasal mucosa.

Controlled drug delivery is a valuable strategy for increasing local therapeutic concentrations in a sustained manner, particularly in locations that are difficult to access. One such target is the sinonasal mucosa, which can be chronically inflamed in patients with rhinitis or rhinosinusitis resulting in diminished quality of life, significant healthcare expenses, and multiple co-morbidities. While numerous medical therapies with daily administration are available, anatomical, physiological, and patient adherence barriers can limit their therapeutic efficacy. As such, there has been considerable development of biomaterial-based systems that can locally deliver anti-inflammatory, antibiotic, decongestant, and antihistamine medications over an extended duration. This review aims to highlight advances in such biomaterial-based systems for sinonasal delivery. Delivery vehicles including nasal packs, dressings, sinus stents, polymeric meshes, nanoparticles, microparticles, and in situ hydrogels are reviewed. Benefits of these vehicles are discussed, as well as their limitations, which, recently, has motivated the development of combination systems that leverage desirable properties of their individual components to enhance therapeutic delivery. Finally, discussion is provided on the potential of combination delivery vehicles, which can provide greater control of the duration of therapeutic release, as well as the ability to encapsulate multiple therapies, provide mechanical support, or conform to the mucosa. The future clinical use of controlled release systems with these attributes could have a transformative impact on improving treatment of difficult-to-control chronic diseases of the sinonasal mucosa.

A ready-to-use, thermoresponsive, and extended-release delivery system for the paranasal sinuses.

A drug delivery system for the paranasal sinuses consisting of a freeze-dried thermoresponsive hydrogel with degradable microspheres, called FD-TEMPS (Freeze Dried-Thermogel, Extended-release Microsphere-based delivery to the Paranasal Sinuses), was developed. Glass transition temperatures (Tg') of the maximally freeze concentrated solutions consisting of poly(N-isopropylacrylamide) (pNIPAAm) and polyethylene glycol (PEG) were determined by differential scanning calorimetry, which informed optimization of the thermogel formulation. By replacing low molecular weight (MW) PEG (200 Da) with a higher MW PEG (2000 Da), the resulting freeze-dried gel exhibited a brittle texture, porous structure, and low residual moisture (< 3% measured by thermal gravimetric analysis). When combined with poly(lactic-co-glycolic acid) microspheres (PLGA MSs) and freeze dried, the complete system (FD-TEMPS) exhibited enhanced shelf-stability. Specifically, the smooth, spherical morphology of the MSs and initial release kinetics were maintained following 6 weeks of storage under ambient conditions. Furthermore, FD-TEMPS remained in place after application to a simulated mucosal surface, suggesting that it could be more uniformly distributed along the sinonasal mucosa in vivo. Freeze drying enables this delivery system to be stored as a ready-to-use product for better ease of clinical translation without compromising the thermoresponsive or sustained release characteristics that would enable local delivery of therapeutics to the sinonasal mucosa.

CCL2 loaded microparticles promote acute patency in silk-based vascular grafts implanted in rat aortae.

Cardiovascular disease is the leading cause of death worldwide, often associated with coronary artery occlusion. A common intervention for arterial blockage utilizes a vascular graft to bypass the diseased artery and restore downstream blood flow; however, current clinical options exhibit high long-term failure rates. Our goal was to develop an off-the-shelf tissue-engineered vascular graft capable of delivering a biological payload based on the monocyte recruitment factor C-C motif chemokine ligand 2 (CCL2) to induce remodeling. Bi-layered silk scaffolds consisting of an inner porous and outer electrospun layer were fabricated using a custom blend of Antherea Assama and Bombyx Mori silk (lyogel). Lyogel silk scaffolds alone (LG), and lyogel silk scaffolds containing microparticles (LGMP) were tested. The microparticles (MPs) were loaded with either CCL2 (LGMP+) or water (LGMP-). Scaffolds were implanted as abdominal aortic interposition grafts in Lewis rats for 1 and 8 weeks. 1-week implants exhibited patency rates of 50% (7/14), 100% (10/10), and 100% (5/5) in the LGMP-, LGMP+, and LG groups, respectively. The significantly higher patency rate for the LGMP+ group compared to the LGMP- group (p = 0.0188) suggests that CCL2 can prevent acute occlusion. Immunostaining of the explants revealed a significantly higher density of macrophages (CD68+ cells) within the outer vs. inner layer of LGMP- and LGMP+ constructs but not in LG constructs. After 8 weeks, there were no significant differences in patency rates between groups. All patent scaffolds at 8 weeks showed signs of remodeling; however, stenosis was observed within the majority of explants. This study demonstrated the successful fabrication of a custom blended silk scaffold functionalized with cell-mimicking microparticles to facilitate controlled delivery of a biological payload improving their in vivo performance. STATEMENT OF SIGNIFICANCE: This study outlines the development of a custom blended silk-based tissue-engineered vascular graft (TEVG) for use in arterial bypass or replacement surgery. A custom mixture of silk was formulated to improve biocompatibility and cellular binding to the tubular scaffold. Many current approaches to TEVGs include cells that encourage graft cellularization and remodeling; however, our technology incorporates a microparticle based delivery platform capable of delivering bioactive molecules that can mimic the function of seeded cells. In this study, we load the TEVGs with microparticles containing a monocyte attractant and demonstrate improved performance in terms of unobstructed blood flow versus blank microparticles. The acellular nature of this technology potentially reduces risk, increases reproducibility, and results in a more cost-effective graft when compared to cell-based options.

Local delivery strategies to restore immune homeostasis in the context of inflammation.

Immune homeostasis is maintained by a precise balance between effector immune cells and regulatory immune cells. Chronic deviations from immune homeostasis, driven by a greater ratio of effector to regulatory cues, can promote the development and propagation of inflammatory diseases/conditions (i.e., autoimmune diseases, transplant rejection, etc.). Current methods to treat chronic inflammation rely upon systemic administration of non-specific small molecules, resulting in broad immunosuppression with unwanted side effects. Consequently, recent studies have developed more localized and specific immunomodulatory approaches to treat inflammation through the use of local biomaterial-based delivery systems. In particular, this review focuses on (1) local biomaterial-based delivery systems, (2) common materials used for polymeric-delivery systems and (3) emerging immunomodulatory trends used to treat inflammation with increased specificity.

Compatibility of a Thermoresponsive and Controlled Release System for Promoting Sinonasal Cilia Regeneration.

The current clinical goal for managing chronic rhinosinusitis (CRS), a heterogenous disease of the paranasal sinuses, is to control inflammation, yet adjunct therapies that promote mucosal regeneration can improve the long-term health of the upper airways. The small natural openings to the sinuses, however, limit the efficacy of traditional drug delivery methods (i.e., nasal sprays and irrigation). Accordingly, a conformable thermoresponsive and controlled release system ("TEMPS", Thermogel, Extended-release Microsphere-based delivery to the Paranasal Sinuses) is developed. The poly(lactic-co-glycolic acid) microsphere component enables the encapsulation of numerous therapeutics, such as retinoic acid (RA), an analog of vitamin A (VA). Studies in CRS patients and preclinical models have shown that aqueous RA or VA gels promoted the differentiation of ciliated cells and improved mucosal healing following repeat applications. In the present study, TEMPS is designed for the controlled release of RA such that a single dose of RA-TEMPS delivers bioactive drug for at least 30 days. Furthermore, as TEMPS will be in direct contact with sinonasal tissue, its compatibility with ciliated human nasal epithelium is explored. After ex vivo incubation in thermogel for 24 h, cilia motility is maintained, providing evidence that TEMPS can be compatible for application along the sinonasal epithelium.

Advances in Multiplexed Paper-Based Analytical Devices for Cancer Diagnosis: A Review of Technological Developments.

Cancer is one of the leading causes of death worldwide producing estimated cost of $161.2 billion in the US in 2017 only. Early detection of cancer would not only reduce cancer mortality rates but also dramatically reduce healthcare costs given that the 17 million new cancer cases in 2018 are estimated to grow 27.5 million new cases by 2040. Analytical devices based upon paper substrates could provide effective, rapid, and extremely low cost alternatives for early cancer detection compared to existing testing methods. However, low concentrations of biomarkers in body fluids as well as the possible association of any given biomarker with multiple diseases remain as one of the greatest challenges to widespread adoption of these paper-based devices. However, recent advances have opened the possibility of detecting multiple biomarkers within the same device, which could be predictive of a patient's condition with unprecedented cost-effectiveness. Accordingly, this review highlights the recent advancements in paper-based analytical devices with a multiplexing focus. The primary areas of interest include lateral flow assay and microfluidic paper-based assay formats, signal amplification approaches to enhance the sensitivity for a specific cancer type, along with current challenges and future outlook for the detection of multiple cancer biomarkers.

Simultaneous Pharmacologic Inhibition of Yes-Associated Protein 1 and Glutaminase 1 via Inhaled Poly(Lactic-co-Glycolic) Acid-Encapsulated Microparticles Improves Pulmonary Hypertension.

Background Pulmonary hypertension (PH) is a deadly disease characterized by vascular stiffness and altered cellular metabolism. Current treatments focus on vasodilation and not other root causes of pathogenesis. Previously, it was demonstrated that glutamine metabolism, as catalyzed by GLS1 (glutaminase 1) activity, is mechanoactivated by matrix stiffening and the transcriptional coactivators YAP1 (yes-associated protein 1) and transcriptional coactivator with PDZ-binding motif (TAZ), resulting in pulmonary vascular proliferation and PH. Pharmacologic inhibition of YAP1 (by verteporfin) or glutaminase (by CB-839) improved PH in vivo. However, systemic delivery of these agents, particularly YAP1 inhibitors, may have adverse chronic effects. Furthermore, simultaneous use of pharmacologic blockers may offer additive or synergistic benefits. Therefore, a strategy that delivers these drugs in combination to local lung tissue, thus avoiding systemic toxicity and driving more robust improvement, was investigated. Methods and Results We used poly(lactic-co-glycolic) acid polymer-based microparticles for delivery of verteporfin and CB-839 simultaneously to the lungs of rats suffering from monocrotaline-induced PH. Microparticles released these drugs in a sustained fashion and delivered their payload in the lungs for 7 days. When given orotracheally to the rats weekly for 3 weeks, microparticles carrying this drug combination improved hemodynamic (right ventricular systolic pressure and right ventricle/left ventricle+septum mass ratio), histologic (vascular remodeling), and molecular markers (vascular proliferation and stiffening) of PH. Importantly, only the combination of drug delivery, but neither verteporfin nor CB-839 alone, displayed significant improvement across all indexes of PH. Conclusions Simultaneous, lung-specific, and controlled release of drugs targeting YAP1 and GLS1 improved PH in rats, addressing unmet needs for the treatment of this deadly disease.

Local Sustained Delivery of Anti-IL-17A Antibodies Limits Inflammatory Bone Loss in Murine Experimental Periodontitis.

Periodontal disease (PD) is a chronic destructive inflammatory disease of the tooth-supporting structures that leads to tooth loss at its advanced stages. Although the disease is initiated by a complex organization of oral microorganisms in the form of a plaque biofilm, it is the uncontrolled immune response to periodontal pathogens that fuels periodontal tissue destruction. IL-17A has been identified as a key cytokine in the pathogenesis of PD. Despite its well documented role in host defense against invading pathogens at oral barrier sites, IL-17A-mediated signaling can also lead to a detrimental inflammatory response, causing periodontal bone destruction. In this study, we developed a local sustained delivery system that restrains IL-17A hyperactivity in periodontal tissues by incorporating neutralizing anti-IL-17A Abs in poly(lactic-coglycolic) acid microparticles (MP). This formulation allowed for controlled release of anti-IL-17A in the periodontium of mice with ligature-induced PD. Local delivery of anti-IL-17A MP after murine PD induction inhibited alveolar bone loss and osteoclastic activity. The anti-IL-17A MP formulation also decreased expression of IL-6, an IL-17A target gene known to induce bone resorption in periodontal tissues. This study demonstrates proof of concept that local and sustained release of IL-17A Abs constitutes a promising therapeutic strategy for PD and may be applicable to other osteolytic bone diseases mediated by IL-17A-driven inflammation.

Biologics and their delivery systems: Trends in myocardial infarction.

Cardiovascular disease is the leading cause of death around the world, in which myocardial infarction (MI) is a precipitating event. However, current therapies do not adequately address the multiple dysregulated systems following MI. Consequently, recent studies have developed novel biologic delivery systems to more effectively address these maladies. This review utilizes a scientometric summary of the recent literature to identify trends among biologic delivery systems designed to treat MI. Emphasis is placed on sustained or targeted release of biologics (e.g. growth factors, nucleic acids, stem cells, chemokines) from common delivery systems (e.g. microparticles, nanocarriers, injectable hydrogels, implantable patches). We also evaluate biologic delivery system trends in the entire regenerative medicine field to identify emerging approaches that may translate to the treatment of MI. Future developments include immune system targeting through soluble factor or chemokine delivery, and the development of advanced delivery systems that facilitate the synergistic delivery of biologics.

Auto-antigen and Immunomodulatory Agent-Based Approaches for Antigen-Specific Tolerance in NOD Mice.

PURPOSE OF REVIEW: Type 1 diabetes (T1D) can be managed by insulin replacement, but it is still associated with an increased risk of microvascular/cardiovascular complications. There is considerable interest in antigen-specific approaches for treating T1D due to their potential for a favorable risk-benefit ratio relative to non-specific immune-based treatments. Here we review recent antigen-specific tolerance approaches using auto-antigen and/or immunomodulatory agents in NOD mice and provide insight into seemingly contradictory findings. RECENT FINDINGS: Although delivery of auto-antigen alone can prevent T1D in NOD mice, this approach may be prone to inconsistent results and has not demonstrated an ability to reverse established T1D. Conversely, several approaches that promote presentation of auto-antigen in a tolerogenic context through cell/tissue targeting, delivery system properties, or the delivery of immunomodulatory agents have had success in reversing recent-onset T1D in NOD mice. While initial auto-antigen based approaches were unable to substantially influence T1D progression clinically, recent antigen-specific approaches have promising potential.

Biorelevant and screening dissolution methods for minocycline hydrochloride microspheres intended for periodontal administration.

Currently, there is no compendial-level method to assess dissolution of particulate systems administered in the periodontal pocket. This work seeks to develop dissolution methods for extended release poly(lactic-co-glycolic acid) (PLGA) microspheres applied in the periodontal pocket. Arestin®, PLGA microspheres containing minocycline hydrochloride (MIN), is indicated for reduction of pocket depth in adult periodontitis. Utilizing Arestin® as a model product, two dissolution methods were developed: a dialysis set-up using USP apparatus 4 and a novel apparatus fabricated to simulate in vivo environment of the periodontal pocket. In the biorelevant method, the microspheres were dispersed in 250 µL of simulated gingival crevicular fluid (sGCF) which was enclosed in a custom-made dialysis enclosure. sGCF was continuously delivered to the device at a biorelevant flow rate and was collected daily for drug content analysis using UPLC. Both methods could discriminate release characteristics of a panel of MIN-loaded PLGA microspheres that differed in composition and process conditions. A mechanistic model was developed, which satisfactorily explained the release profiles observed using both dissolution methods. The developed methods may have the potential to be used as routine quality control tools to ensure batch-to-batch consistency and to support evaluation of bioequivalence for periodontal microspheres.

Cranberry extract-based formulations for preventing bacterial biofilms.

Generating formulations for the delivery of a mixture of natural compounds extracted from natural sources is a challenge because of unknown active and inactive ingredients and possible interactions between them. As one example, natural cranberry extracts have been proposed for the prevention of biofilm formation on dental pellicle or teeth. However, such extracts may contain phenolic acids, flavonol glycosides along with other constituents like coumaroyl iridoid glycosides, flavonoids, alpha-linolenic acid, n-6 (or n-3) fatty acids, and crude fiber. Due to the presence of a variety of compounds, determining which molecules (and how many molecules) are essential for preventing biofilm growth is nontrivial to ascertain. Therefore, a formulation that could contain natural, unrefined, cranberry extract (with all its constituent compounds) at high loading would be ideal. Accordingly, we have generated several candidate formulations including poly(lactic-co-glycolic) acid (PLGA)-based microencapsulation of cranberry extract (CE15) as well as formulations including stearic acid along with polyvinylpyrrolidone (PVP) or Ethyl lauroyl arginate (LAE) complexed with cranberry extracts (CE15). We found that stearic acid in combination with PVP or LAE as excipients led to higher loading of the active and inactive compounds in CE15 as compared with a PLGA microencapsulation and also sustained release of CE15 in a tunable manner. Using this method, we have been able to generate two successful formulations (one preventative based, one treatment based) that effectively inhibit biofilm growth when incubated with saliva. In addition to cranberry extract, this technique could also be a promising candidate for other natural extracts to form controlled release systems.Graphical abstract.

Injectable thermoresponsive hydrogels as drug delivery system for the treatment of central nervous system disorders: A review.

The central nervous system (CNS), consisting of the brain, spinal cord, and retina, superintends to the acquisition, integration and processing of peripheral information to properly coordinate the activities of the whole body. Neurodegenerative and neurodevelopmental disorders, trauma, stroke, and brain tumors can dramatically affect CNS functions resulting in serious and life-long disabilities. Globally, the societal and economic burden associated with CNS disorders continues to grow with the ageing of the population thus demanding for more effective and definitive treatments. Despite the variety of clinically available therapeutic molecules, medical interventions on CNS disorders are mostly limited to treat symptoms rather than halting or reversing disease progression. This is attributed to the complexity of the underlying disease mechanisms as well as to the unique biological microenvironment. Given its central importance, multiple barriers, including the blood brain barrier and the blood cerebrospinal fluid barrier, protect the CNS from external agents. This limits the access of drug molecules to the CNS thus contributing to the modest therapeutic successes. Loco-regional therapies based on the deposition of thermoresponsive hydrogels loaded with therapeutic agents and cells are receiving much attention as an alternative and potentially more effective approach to manage CNS disorders. In this work, the current understanding and challenges in the design of thermoresponsive hydrogels for CNS therapy are reviewed. First, the biological barriers that hinder mass and drug transport to the CNS are described, highlighting the distinct features of each barrier. Then, the realization, characterization and biomedical application of natural and synthetic thermoresponsive hydrogels are critically presented. Advantages and limitations of each design and application are discussed with the objective of identifying general rules that could enhance the effective translation of thermoresponsive hydrogel-based therapies for the treatment of CNS disorders.

A thermoresponsive hydrogel system for long-acting corticosteroid delivery into the paranasal sinuses.

Delivering localized treatment to the paranasal sinuses for diseases such as chronic rhinosinusitis (CRS) is particularly challenging because of the small natural openings leading from the sinuses that can be further obstructed by presence of inflammation. As such, oral steroids, topical nasal sprays or irrigation, and surgery can be utilized to treat persistent sinonasal inflammation, but there exists a need for post-operative options for long-term steroid delivery to prevent disease recurrence. In the present study, a Thermogel, Extended-release Microsphere-based-delivery to the Paranasal Sinuses (TEMPS) is developed with the corticosteroid mometasone furoate. Specifically, the bioactive steroid is released for 4 weeks from poly(lactic-co-glycolic acid) (PLGA) microspheres embedded in a poly(N-isopropylacrylamide) (p-NIPAAm)-based hydrogel. The temperature-responsive system undergoes a reversible sol-gel transition at 34-35 °C such that it can be applied as a liquid at ambient temperature, conforming to the sinonasal epithelium as it gels. In a rabbit model of CRS, TEMPS was maintained in rabbit sinuses and effectively reduced sinonasal inflammation as characterized by micro-computed tomography and histopathology analysis. Ultimately, the combination of controlled release microspheres with a thermoresponsive hydrogel provides flexibility for encapsulating therapeutics in a reversible and conforming system for localized delivery to the sinuses.