By Gilang Cahya 
In preclinical models, the inhalation of a mixture of living Lactobacilli bacteria attenuated pulmonary inflammation and improved lung function and structure for the chronic lung diseases bronchopulmonary dysplasia and chronic obstructive pulmonary disease. This groundbreaking research, published in the esteemed journal Nature Communications, offers a novel therapeutic avenue for debilitating respiratory conditions affecting both vulnerable infants and adults. The study, spearheaded by a team of researchers at the University of Alabama at Birmingham (UAB), elucidated the intricate mechanism by which this live biotherapeutic product—a carefully engineered powder mixture of living Lactobacilli bacteria—effectively reduces neutrophilic inflammation and a wide spectrum of inflammatory markers in conditions like bronchopulmonary dysplasia (BPD) and chronic obstructive pulmonary disease (COPD).
Charitharth Vivek Lal, M.D., a neonatologist at UAB, and Amit Gaggar, M.D., Ph.D., a pulmonologist at UAB, co-led this significant research. Their findings, as Dr. Lal articulated, "provide a paradigm for the progression of structural lung disease." This is largely due to the identification of Lactobacilli as critical regulators of lung protease activity. This activity is intrinsically linked to the destructive processes driven by matrikine generation, extracellular matrix turnover, and chronic neutrophilic inflammation, all of which contribute to the damage of lung air sacs.
The foundation for investigating the potential protective role of Lactobacilli in the lungs and their application in treating chronic lung disease was laid in 2016. At that time, Dr. Lal and his UAB colleagues observed a striking correlation: infants suffering from severe bronchopulmonary dysplasia exhibited diminished populations of Lactobacilli, alongside an increased abundance of proteobacteria and elevated concentrations of proteobacterial endotoxin in their airways. This initial observation ignited the pursuit of understanding how these microbial imbalances contribute to lung disease. The latest study builds upon this crucial insight by providing a concrete mechanism of action for Lactobacilli-based therapy, demonstrating its capacity to mitigate downstream disease development. Furthermore, the research rigorously assessed the safety and effectiveness of this live biotherapeutic treatment in both a mouse pup model specifically engineered to mimic BPD and three distinct mouse models of COPD.
Understanding the Diseases: BPD and COPD
To fully appreciate the significance of these findings, it is essential to understand the scope and impact of the diseases targeted. Bronchopulmonary dysplasia (BPD) is a serious chronic lung disease that primarily affects premature infants. It typically develops as a consequence of damage inflicted by the high oxygen concentrations or mechanical ventilation often required to support their fragile respiratory systems during the critical neonatal period. These interventions, while life-saving, can lead to long-term lung injury and impaired development.
Chronic Obstructive Pulmonary Disease (COPD), on the other hand, is a progressive and irreversible lung disease that predominantly affects older individuals, with smoking being the most significant risk factor. COPD encompasses conditions such as emphysema and chronic bronchitis, leading to airflow limitation and breathing difficulties. The impact of COPD is substantial, claiming approximately 130,000 American lives annually and affecting an estimated 3 million more individuals worldwide. The persistent inflammation and structural damage characteristic of both BPD and COPD underscore the urgent need for innovative therapeutic strategies.
Unraveling the Mechanism: Lactobacilli and Lung Proteases
The UAB researchers’ hypothesis centered on a key molecular player: acetylated proline-glycine-proline (Ac-PGP). This peptide, derived from the extracellular matrix, was previously observed at heightened levels in premature infants diagnosed with BPD. The study confirmed this observation in their BPD mouse models, providing a crucial link between extracellular matrix degradation and lung disease.
Through meticulous gain- and loss-of-function studies, the researchers definitively established the impact of Ac-PGP on lung health. They found that intranasal instillation of Ac-PGP in the mouse models led to a significant increase in neutrophilic inflammation and a corresponding degradation of lung tissue. Conversely, when an inhibitor of Ac-PGP was administered concurrently with Ac-PGP, markers of neutrophilic inflammation were substantially reduced, and the structural integrity of the lungs showed marked improvement. This demonstrated Ac-PGP’s direct role in driving inflammation and tissue damage.
The next critical step was to identify how Lactobacilli could counteract this detrimental process. The research team discovered that a proprietary blend of Lactobacilli, specifically L. plantarum, L. acidophilus, and L. rhamnosus, demonstrated remarkable synergy in reducing the activity of a key inflammatory proteinase known as MMP-9. MMP-9 plays a pivotal role in the breakdown of the extracellular matrix, and its overactivity is implicated in the release of Ac-PGP. Notably, the study also revealed that the supernatant derived from Lactobacilli growth medium exhibited a comparable reduction in MMP-9 levels to that of live Lactobacilli bacteria, suggesting that soluble factors produced by the bacteria were also at play.
A particularly significant finding emerged from the investigation into the components of the Lactobacilli growth medium. Researchers pinpointed L(+) lactic acid, a byproduct of Lactobacilli metabolism, as a potent anti-inflammatory molecule. In vitro experiments confirmed that L(+) lactic acid effectively reduced MMP-9 activity. This discovery provided a crucial insight into the therapeutic mechanism: live Lactobacilli residing in the lungs could facilitate a sustained and controlled release of L(+) lactic acid, thereby exerting ongoing anti-inflammatory effects in a well-tolerated manner.
Technological Innovation and Therapeutic Efficacy
A major technological hurdle addressed by the researchers was the development of an effective delivery method for the live bacteria to the lungs. They achieved a significant breakthrough by engineering inhaled Lactobacilli powder through particle engineering. This process ensured that the particles were sufficiently small to reach deep within the lung’s delicate structures while crucially preserving the viability of the bacteria. This advanced formulation, the live biotherapeutic product, was then rigorously tested in the established BPD and COPD mouse models.
In the COPD mouse models, the Lactobacilli blend demonstrated remarkable efficacy. It successfully reduced inflammation within the lung microenvironment, regardless of whether treatment was administered concurrently with the injury or post-injury. This indicated broad anti-inflammatory capabilities, a decrease in several pro-inflammatory markers, and a notable elevation of the anti-inflammatory marker immunoglobulin A (IgA).
Perhaps most compelling was the direct comparison of the live biotherapeutic product’s performance against a widely used pharmaceutical. The Lactobacilli blend was found to reduce MMP-9 and other pro-inflammatory cytokines as effectively as, and in some cases even better than, fluticasone furoate. Fluticasone furoate is a United States Food and Drug Administration-approved inhaled corticosteroid commonly incorporated into combination therapies for COPD. This finding suggests a potential for Lactobacilli-based therapies to offer comparable or superior efficacy to existing treatments, potentially with a different safety profile.
Safety and Future Prospects
Crucially, the study included comprehensive safety and biodistribution assessments in one of the COPD mouse models. These evaluations confirmed that the inhalation of the bacterial powder did not induce adverse reactions or exacerbate disease progression. Furthermore, the Lactobacilli did not translocate to distant tissues or accumulate in unintended areas of the lungs, indicating a favorable safety profile for this inhaled biotherapeutic approach.
The implications of these preclinical findings are profound. Dr. Lal expressed optimism about the future, stating, "Inhaled live biotherapeutic products show promise in addressing common pathways of disease progression that in the future can be targeted at a variety of lung diseases." He emphasized that while the preclinical animal data is highly suggestive, the safety of this potential drug in humans will be rigorously tested in a forthcoming clinical trial. "Human adult safety data in COPD will help de-risk the pathway to approval for use of the drug in bronchopulmonary disease infants," he added, highlighting the strategic approach to advancing this therapy.
A Collaborative Endeavor and Path to Commercialization
The research was a testament to extensive collaboration, with Teodora Nicola and Nancy Wenger from the UAB Department of Pediatrics, Division of Neonatology, serving as co-first authors of the study, titled "A Lactobacilli-based inhaled live biotherapeutic product attenuates pulmonary neutrophilic inflammation."
The broader research team included Xin Xu, Camilla Margaroli, Kristopher Genschmer, and J. Edwin Blalock from the UAB Department of Medicine, Division of Pulmonary, Allergy and Critical Care Medicine. Additionally, Michael Evans, Luhua Qiao, Gabriel Rezonzew, Youfeng Yang, Tamas Jilling, Kent Willis, and Namasivayam Ambalavanan from the UAB Department of Pediatrics, Division of Neonatology, made significant contributions.
The research received vital support from multiple grants awarded by the National Heart, Lung, and Blood Institute of the National Institutes of Health, including grants HL141652, HL135710, HL166433, HL156275, and HL164156.
Further underscoring the translational nature of this work, a portion of the research is protected by a patent titled "Inhaled respiratory probiotics for lung diseases of infancy, childhood and adulthood" (U.S. 11,141,443 B2). This patent is held by the University of Alabama at Birmingham Research Foundation, an entity within the Bill L. Harbert Institute for Innovation and Entrepreneurship, with Dr. Lal, Dr. Gaggar, and Dr. Ambalavanan listed as inventors. The proprietary product has been successfully commercialized through Alveolus Bio, Inc., a UAB startup based in Birmingham, Alabama, and Boston, Massachusetts, signaling a clear pathway from laboratory discovery to potential clinical application.
At UAB, the departments of Pediatrics and Medicine are integral to the Marnix E. Heersink School of Medicine. Dr. Lal holds a leadership role as the director of Clinical Innovation at the Marnix E. Heersink Institute for Biomedical Innovation. He also serves as an associate professor in the Division of Neonatology. Dr. Gaggar is a distinguished professor in the Division of Pulmonary, Allergy and Critical Care Medicine. Dr. Lal is also recognized as the founder of Alveolus Bio, Inc., and Resbiotic Nutrition, Inc., further demonstrating his commitment to pioneering new therapeutic solutions.
This comprehensive study represents a significant leap forward in the quest for effective treatments for chronic lung diseases. By harnessing the power of beneficial bacteria and a novel delivery system, the research offers a beacon of hope for improved lung health in vulnerable populations and beyond. The transition from preclinical promise to human clinical trials will be a critical next step in validating these exciting findings and potentially revolutionizing the management of these pervasive respiratory conditions.
