Faculty Bibliography

INTRODUCTION/UNASSIGNED:Outcomes for NSCLC remain suboptimal. Recent data suggest that cryoablation can generate antitumor immune effects. In this first-in-human phase I clinical trial, we investigated the safety and feasibility of bronchoscopic cryoimmunotherapy (BCI) delivered during standard-of-care bronchoscopy and explored associated systemic immune responses. METHODS/UNASSIGNED:Subjects with known or suspected advanced-stage NSCLC were recruited. BCI was delivered in dose-escalated freeze-thaw cycles to determine maximum dose tolerance. Feasibility assessment was determined with a pre-set goal of achieving successful BCI in more than or equal to 80% of subjects. Safety was assessed by review of BCI-related complications, including grades 2 to 3 bleeding, pneumothorax requiring intervention, and National Cancer Institute Common Terminology Criteria for Adverse Events grade 3 to 5 adverse events. Pre- and post-BCI blood samples were collected to explore changes in the systemic immune profile. RESULTS/UNASSIGNED:Subjects with predominantly clinical TNM stage 3 or 4 adenocarcinoma or squamous cell carcinoma were enrolled. We reached the maximum dose of 30 seconds with 100% feasibility and no BCI-related adverse events. In peripheral blood analysis, we observed a significant decrease in derived neutrophil-to-lymphocyte ratio in the high-dose BCI group in comparison to the low-dose BCI cohort. We also observed increases in inflammatory cytokines-GM-CSF, IFN-γ, IL-1β, IL-17A, and IL-2-and effector memory T cells post-BCI. CONCLUSION/UNASSIGNED:BCI is safe and feasible. In addition, we provide preliminary evidence that at higher dose levels there is a systemic immune response consistent with a cytotoxic profile. Further immune analyses will determine the potential of BCI as an adjunctive therapy in combination with immune checkpoint inhibition in NSCLC treatment.

Variations in the airway microbiome are associated with inflammatory responses in the lung and pulmonary disease outcomes. Regional changes in microbiome composition could have spatial effects on the metabolic environment, contributing to differences in the host response. Here, we profiled the respiratory microbiome (metagenome/metatranscriptome) and metabolome of a patient cohort, uncovering topographical differences in microbial function, which were further delineated using isotope probing in mice. In humans, the functional activity of taxa varied across the respiratory tract and correlated with immunomodulatory metabolites such as glutamic acid/glutamate and methionine. Common oral commensals, such as Prevotella, Streptococcus, and Veillonella, were more functionally active in the lower airways. Inoculating mice with these commensals led to regional increases in several metabolites, notably methionine and tyrosine. Isotope labeling validated the contribution of Prevotella melaninogenica in generating specific metabolites. This functional characterization of microbial communities reveals topographical changes in the lung metabolome and potential impacts on host responses.

The role of dietary fiber in colon cancer prevention remains controversial. We investigated its impact on anti-tumor immunity and the gut microbiota in APCmin/+ mice infected with Enterotoxigenic Bacteroides fragilis. Mice were fed high-fiber, low-fiber, or chow diets, and tumor burden, survival, cytokines, microbiota, and metabolites were analyzed. Contrary to the belief that high fiber inhibits tumor progression, it had no significant impact compared to chow diet. However, the low-fiber diet significantly reduced tumor burden and improved survival. Mechanistically, high fiber increased pro-inflammatory cytokines and CD4+Foxp3+RORγt+IL-17A+ regulatory T cells, while low fiber enhanced anti-inflammatory cytokines and cytotoxic T-cells. High fiber enriched microbial taxa associated with IL-17A+RORγt+ Tregs and altered metabolites, including reduced tryptophan and increased short-chain fatty acids and bile acids. Low fiber produced opposite effects. These findings suggest that dietary fiber's effects on colon cancer depends on microbial infection and immune status, emphasizing the need for personalized dietary interventions in colon cancer management.

Non-small cell lung cancer (NSCLC) remains the most common cause for cancer-related mortality despite advances in treatment. Early detection is crucial for improving patient outcomes, yet current diagnostic and prognostic molecular biomarkers lack the sensitivity and specificity necessary to become clinically useful. Recent studies revealed that the lower airway microbiome play a role in NSCLC and that microbial signatures are associated with NSCLC development, progression, and prognosis, suggesting the potential for microbiome-based biomarkers for early diagnosis and risk stratification. Here we review recent advances in the role of the local and systemic microbiome in early-stage NSCLC. Primarily, several studies have identified specific microbial taxa associated with lung cancer suggesting novel insights into disease pathogenesis and progression. Integration of microbiome data with other 'omics' platforms, such as host transcriptomics and metabolomics, has the potential to enhance our understanding of microbial-host interactions and may provide more comprehensive biomarker signatures. While promising, challenges remain to the development of microbiome-based biomarkers such as those related to differences in samples utilized, sequencing methods, and data analysis. Here, we discuss such challenges as well as future directions for research needed to fulfil the promise of microbiome-based biomarkers for changing early detection and management strategies in NSCLC.

RATIONALE/BACKGROUND:Lower airway enrichment with oral commensals has been previously associated with grade 3 severe primary graft dysfunction (PGD) after lung transplantation (LT). We aimed to determine whether this dysbiotic signature is present across all PGD severity grades, including milder forms, and whether it is associated with a distinct host inflammatory endotype. METHODS:Lower airway samples from 96 LT recipients with varying degrees of PGD were used to evaluate the lung allograft microbiota via 16S rRNA gene sequencing. Bronchoalveolar lavage (BAL) cytokine concentrations and cell differential percentages were compared across PGD grades. In a subset of samples, we evaluated the lower airway host transcriptome using RNA sequencing methods. RESULTS:Differential analyses demonstrated lower airway enrichment with supraglottic-predominant taxa (SPT) in both moderate and severe PGD. Dirichlet Multinomial Mixtures (DMM) modeling identified two distinct microbial clusters. A greater percentage of subjects with moderate-severe PGD were identified within the dysbiotic cluster (C-SPT) than within the no PGD group (48 and 29%, respectively) though this difference did not reach statistical significance (p=0.06). PGD severity associated with increased BAL neutrophil concentration (p=0.03) and correlated with BAL concentrations of MCP-1/CCL2, IP-10/CXCL10, IL-10, and TNF-α (p<0.05). Furthermore, microbial signatures of dysbiosis correlated with neutrophils, MCP-1/CCL-2, IL-10, and TNF-α (p<0.05). C-SPT exhibited differential expression of TNF, SERPINE1 (PAI-1), MPO, and MMP1 genes and upregulation of MAPK pathways, suggesting that dysbiosis regulates host signaling to promote neutrophilic inflammation. CONCLUSIONS:Lower airway dysbiosis within the lung allograft is associated with a neutrophilic inflammatory endotype, an immune profile commonly recognized as the hallmark for PGD pathogenesis. This data highlights a putative role for lower airway microbial dysbiosis in the pathogenesis of this syndrome.

BACKGROUND/UNASSIGNED:Hydroxychloroquine (HCQ) is a frontline treatment for autoimmune diseases, including rheumatoid arthritis, Sjogren's syndrome, and systemic lupus erythematosus (SLE), due to its potent immunomodulatory properties. Efferocytosis, a crucial process for tissue homeostasis by transmitting immune-suppressive signals, is frequently impaired in SLE. We hypothesized HCQ enhances efferocytosis and mediates anti-inflammatory effects. METHODS/UNASSIGNED:. The role of MerTK on HCQ-modulated inflammation was revealed by MerTK inhibitor UNC2025. RESULTS/UNASSIGNED:Long-term HCQ treatment in PIL mice significantly reduced disease activity. HCQ treatment enhanced efferocytosis in RAW264.7 cells, while peritoneal macrophages from HCQ-treated mice showed increased efferocytotic capacity compare to PIL mice. Additionally, HCQ upregulated the expression of the TAM receptor MerTK and Gas6 on macrophages, restoring MerTK levels suppressed by pristane in the spleen of PIL mice. Inhibition of MerTK signaling by UNC2025 mitigated HCQ-mediated enhancements in efferocytosis and reversed the reduction in inflammatory mediators including IL-6 and IFN-α. HCQ-induced anti-inflammatory markers, such as PPARγ, LXR, and IL-10, were also alleviated upon MerTK blockade. CONCLUSION/UNASSIGNED:evidence that HCQ promotes macrophage efferocytosis and anti-inflammatory reprogramming via MerTK/Gas6 signaling, offering insights into potential therapeutic mechanisms in SLE management.

BACKGROUND:Risk of early-stage lung adenocarcinoma (LUAD) recurrence after surgical resection is significant, and post-recurrence median survival is approximately two years. Currently there are no commercially available biomarkers that predict recurrence. Here, we investigated whether microbial and host genomic signatures in the lung can predict recurrence. METHODS:In 91 early-stage (Stage IA/IB) LUAD-patients with extensive follow-up, we used 16s rRNA gene sequencing and host RNA-sequencing to map the microbial and host transcriptomic landscape in tumor and adjacent unaffected lung samples. RESULTS:23 out of 91 subjects had tumor recurrence over 5-year period. In tumor samples, LUAD recurrence was associated with enrichment with Dialister, Prevotella, while in unaffected lung, recurrence was associated with enrichment with Sphyngomonas and Alloiococcus. The strengths of the associations between microbial and host genomic signatures with LUAD recurrence were greater in adjacent unaffected lung samples than in the primary tumor. Among microbial-host features in the unaffected lung samples associated with recurrence, enrichment with Stenotrophomonas geniculata and Chryseobacterium were positively correlated with upregulation of IL-2, IL-3, IL-17, EGFR, HIF-1 signaling pathways among the host transcriptome. In tumor samples, enrichment with Veillonellaceae Dialister, Ruminococcacea, Haemophilus Influenza, and Neisseria were positively correlated with upregulation of IL-1, IL-6, IL17, IFN, and Tryptophan metabolism pathways. CONCLUSIONS:Overall, modeling suggested that a combined microbial/transcriptome approach using unaffected lung samples had the best biomarker performance (AUC=0.83). IMPACT/CONCLUSIONS:This study suggests that LUAD recurrence is associated with distinct pathophysiological mechanisms of microbial-host interactions in the unaffected lung rather than those present in the resected tumor.