Injected mRNA vaccines have been proven effective and safe in the SARS-CoV-2 pandemic. Using the machinery of the cell, mRNA vaccines translate into an antigen, which triggers an adaptive immune re...

Chronic obstructive pulmonary disease (COPD) is the third leading cause of death worldwide, with a significant impact on low‐ and middle‐income countries, making it a critical challenge for global health equity and sustainable development goals. Extracellular vesicles (EVs) are emerging as a promising treatment for COPD, but conventional storage at −80 °C limits their global accessibility. This study explores alternative storage methods to enhance EV stability and accessibility, particularly in low‐resource settings. EVs from lung fibroblasts are subjected to freezing, freeze drying, and spray drying with inulin or mannitol. The biophysical properties are evaluated by their capacity to support lung organoid formation. Inulin proves effective in stabilizing EVs, maintaining functionality after freezing at −20 °C for 12 weeks. Freeze drying with inulin preserves EV stability at 20 °C and 43% relative humidity. In contrast, EVs with mannitol exhibit aggregation and reduce bio‐functionality. Additionally, spray drying EVs with inulin and leucine produces a dry powder suitable for inhalation, maintaining biophysical properties and functionality for 12 weeks. The powder demonstrates efficient lung deposition using the Cyclops inhaler. These findings suggest inulin as a stabilizer for EVs, eliminating the need for ultra‐low temperature storage and improving the practicality of EV‐based inhalable therapies.

Chronic obstructive pulmonary disease (COPD) is one of the most common lung diseases worldwide, characterized by an accelerated loss of lung function. A key problem underlying COPD is increased tissue destruction in combination with defective lung tissue repair. As current therapies do not modify the progression of the disease, new therapies aimed at restoring lung tissue repair in COPD need to be developed. In an attempt to address this major unmet need, there has been a surge in both preclinical and clinical studies, aiming to identify key mechanisms underpinning defective lung repair and the ability to inhibit or even reverse this defect. This includes small molecules such as retinoids, as well as advanced therapy medicinal products such as cell therapies or therapies with cell-derived products such as extracellular vesicles, or secreted proteins. The results of these endeavors have been variable with failures as well as successful proof-of-concepts. In this review, we provide an overview of the current state of the field, including modes of action of the therapeutics that are or have been considered for lung regeneration, including a discussion on the reasons for failure where relevant. In addition, we discuss hurdles in the clinical development of regenerative therapeutics for COPD including clinical outcomes, route of administration and formulation as these are pivotal considerations moving forward. SIGNIFICANCE STATEMENT: Chronic obstructive pulmonary disease is characterized by progressive alveolar destruction and defective epithelial regeneration. Targetable mechanisms, including cellular senescence, altered mesenchymal-epithelial signaling, and chronic inflammation, impair progenitor function and niche integrity. Therapeutic strategies that restore epithelial repair, including small molecules, biologics, and cell-based approaches, represent a promising path toward disease modification and long-term lung function restoration.

Background: Pulmonary administration offers a promising route for certain biologics, particularly when treating respiratory diseases. Spray drying is widely employed to produce inhalable powders with the biologics incorporated in a stabilizing amorphous sugar. Hydrophobic amino acids such as leucine are frequently added to improve dispersibility. Objectives: While the aerodynamic benefits of leucine are well established, its influence on irreversible moisture-induced dry powder particle aggregation and protein stability during storage remains less evaluated. Methods: In this work, inulin-based powders with and without 4 wt-% leucine were spray dried and stored at 43%, 58%, 69%, and 75% relative humidity (RH) at 20 ± 2 °C. Results: Immediately after drying, both formulations displayed comparable physicochemical characteristics. However, during storage of inulin-only formulations, dry powder particles showed viscous flow and formed big irreversible aggregates after storage at an RH of 58% and above, whereas leucine-containing powders remained intact across all tested conditions up to 20 days. Protein stability was assessed using lactate dehydrogenase (LDH) and β-galactosidase (β-gal) as model proteins. At 43% RH, the Tg remained above the storage temperature, and both LDH and β-gal retained their enzymatic activity for up to 20 days. At 75% RH, however, the Tg dropped to below storage temperature, resulting in a loss of stability for LDH, consistent with its reliance on vitrification. In contrast, β-gal maintained its activity at 75% RH, indicative of stabilization through water replacement. Conclusions: Overall, these results demonstrate that leucine enhances the physical stability of inulin powders by preventing irreversible aggregation under humid conditions. However, this effect does not extend to protecting proteins reliant on vitrification. These findings highlight the potential of inulin and leucine to reduce the need for stringent storage conditions of biologics.

Background: Self-amplifying mRNA (sa-mRNA) represents a promising platform for vaccines and gene therapies, offering sustained protein expression at low doses through self-replication. For vaccines targeting respiratory pathogens, pulmonary delivery of sa-mRNA lipid nanoparticles (LNPs) is particularly advantageous, enabling direct delivery to the infection site and induction of mucosal immunity. Objective: In this study, we evaluated the stability of sa-mRNA–LNPs under refrigerated and frozen conditions and developed a dry powder formulation suitable for inhalation, produced by freeze-drying followed by cryomilling with leucine. Methods: sa-mRNA–LNPs formulated in HEPES buffer with 20% (w/v) sucrose were stored for up to 8 weeks as liquid or freeze-dried samples at various temperatures (−80 °C, −20 °C, 4 °C, and 20 °C). Biological stability was assessed by transfection efficiency in HeLa cells, while physical stability was characterized by encapsulation efficiency, zeta potential, particle size, and polydispersity index. Results: Liquid formulations remained stable for at least 8 weeks at −80 °C and −20 °C but rapidly lost stability at 4 °C and 20 °C. Freeze-drying effectively preserved sa-mRNA–LNP functionality and structural integrity for up to 8 weeks at 4 °C, with only minor structural changes. Subsequent cryomilling in the presence of 4 wt-% leucine produced a respirable dry powder while retaining approximately 60% of the original sa-mRNA–LNP functionality. Although cryomilling induced some structural alterations, the remaining functional fraction remained stable during storage. The resulting powders displayed favorable aerosol performance for deep lung delivery, as demonstrated by cascade impaction (MMAD = 4.13 ± 0.26 µm). Conclusions: In conclusion, freeze-drying effectively preserved sa-mRNA–LNP integrity at 4 °C, whereas cryomilling with leucine produced a respirable dry powder suitable for pulmonary delivery, providing a foundation for globally accessible, needle-free sa-mRNA vaccines against respiratory diseases.

Chronic obstructive pulmonary disease (COPD) is characterized by irreversible airflow limitation and progressive lung function decline, with a critical need for innovative treatments that can repair damaged lung tissue. The active fragment of osteoglycin (OGN) has previously shown significant regenerative potential in activating growth of alveolar epithelial cells, making it a promising candidate for therapeutic development. In this study, we developed an inhalable dry powder formulation of the OGN fragment using spray drying, incorporating inulin or mannitol as excipients and leucine as a dispersion enhancer. The formulations were assessed for their ability to support lung organoid formation and differentiation in vitro. After spray drying with inulin or mannitol, the OGN fragment maintained its ability to induce organoid formation, but only the formulation containing inulin supported the differentiation towards alveolar-type organoids (surfactant protein C positive). After 28 days of storage at high temperatures (60 °C) inulin was found to provide superior stability compared to mannitol. The dry powder formulation with OGN fragment and inulin demonstrated favorable aerodynamic properties, meeting the requirements for deep lung deposition when dispersed via a dry powder inhaler. Additionally, we showed that adding 4 % leucine or inulin sweeper particles significantly improved the dispersion behavior of the formulation and reduced dry powder inhaler retention. This study highlights the potential of an inhalable OGN fragment formulation as a novel therapeutic approach for lung tissue repair in COPD. The findings support further development and clinical evaluation of this formulation, particularly with inulin as the preferred stabilizing excipient.