Potential novel biomarkers for chronic lung allograft dysfunction and azithromycin responsive allograft dysfunction

Chronic lung allograft dysfunction (CLAD) and its obstructive form, the obliterative bronchiolitis (OB), are the main long-term complications related to high mortality rate postlung transplantation. CLAD treatment lacks a significant success in survival. Here, we investigated a new strategy through inhibition of the proinflammatory mitogen- and stress-activated kinase 1 (MSK1) kinase. MSK1 expression was assessed in a mouse OB model after heterotopic tracheal allotransplantation. Pharmacological inhibition of MSK1 (H89, fasudil, PHA767491) was evaluated in the murine model and in a translational model using human lung primary fibroblasts in proinflammatory conditions. MSK1 expression was graded over time in biopsies from a cohort of CLAD patients. MSK1 mRNA progressively increased during OB (6.4-fold at D21 posttransplantation). Inhibition of MSK1 allowed to counteract the damage to the epithelium (56% restoration for H89), and abolished the recruitment of MHCII+ (94%) and T cells (100%) at the early inflammatory phase of OB. In addition, it markedly decreased the late fibroproliferative obstruction in allografts (48%). MSK1 inhibitors decreased production of IL-6 (whose transcription is under the control of MSK1) released from human lung fibroblasts (96%). Finally, we confirmed occurrence of a 2.9-fold increased MSK1 mRNA expression in lung biopsies in patients at 6 months before CLAD diagnosis as compared to recipients with stable lung function. These findings suggest the overall interest of the MSK1 kinase either as a marker or as a potential therapeutic target in lung dysfunction posttransplantation.

Chronic Lung Allograft Dysfunction (CLAD) manifests as Bronchiolitis Obliterans Syndrome (BOS) and the recently described Restrictive Allograft Syndrome (RAS). CLAD is unpredictable and irreversible. Thus predictive biomarkers of CLAD are needed to provide an early and personalized intervention. Our objective is to establish a predictive blood transcriptomic signature of CLAD. We hypothesized that gene and miRNA expression profiling of peripheral blood could uncover the early alterations of CLAD before the degradation of lung function. We selected 88 lung transplant recipients from the COLT (Cohort in Lung Transplantation) cohort. Patients were unequivocally phenotyped by an adjudication committee at 3 years post transplantation as stable (n=49), BOS (n=29) or RAS (n=10). Blood transcriptome (mRNA and microRNA) was investigated longitudinally at 6 months and 1 year post-transplantation, <i>i.e</i>. before the onset of CLAD. Integrated analysis of miRNA and mRNA expression was performed. Preliminary results show more than 500 differentially expressed genes (DEG) between Stable and BOS groups. Hierarchical clustering of DEG discriminates between stable and BOS patients with an accuracy approaching 80%. Gene ontology was conducted to categorize the function of the DEG. Analysis of KEGG pathways revealed several enrichment-related pathways including haematopoietic cell lineage or B cell receptor signaling pathway. As a conclusion, our work supports blood transcriptome analysis to predict and to explore the physiopathology of CLAD. Several biomarkers and pathways were identified. Investigations are ongoing to define the final gene-set predictor. Systems prediction of CLAD: http://www.sysclad.eu/

Cohort characteristics and chronic lung allograft dysfunction (CLAD) adjudication in a FP-7 funded project, systems prediction of CLAD (SysCLAD)

Patients with Bronchiolitis Obliterans Syndrome (BOS) Have Later Onset of Acute Rejection Than BOS-Free Patients

Imaging Mass Cytometry for Detailed Cellular and Spatial Characterization of Chronic Lung Allograft Dysfunction (CLAD)

Intragraft IgG Levels and Donor-Specific anti-HLA Antibodies in Different Phenotypes of Chronic Lung Allograft Dysfunction

Chronic lung allograft dysfunction (CLAD) is the major long-term cause of morbidity and mortality after lung transplantation. Both bronchiolitis obliterans syndrome and restrictive lung allograft syndrome, two main types of CLAD, lead to fibrosis in either the small airways or alveoli and pleura. Pathological pathways in CLAD and other types of fibrosis, for example idiopathic pulmonary fibrosis, are assumed to overlap and therefore fibrosis biomarkers could aid in the early detection of CLAD. These biomarkers could help to differentiate between different phenotypes of CLAD and could, in comparison to biomarkers of inflammation, possibly distinguish an infectious event from CLAD when a decline in lung function is present. This review gives an overview of known CLAD specific biomarkers, describes new promising fibrosis biomarkers currently investigated in other types of fibrosis, and discusses the possible use of these fibrosis biomarkers for CLAD.

Prediction of chronic lung allograft dysfunction: a systems medicine challenge

Restrictive chronic lung allograft dysfunction (CLAD) demonstrates poor outcomes after lung transplantation. However, the impact of radiological patterns on survival within a restrictive CLAD under the new International Society for Heart and Lung Transplantation (ISHLT) criteria remains unclear. We analyzed retrospectively 241 bilateral lung transplant recipients between 2005 and 2021. CLAD was diagnosed and classified per the 2019 ISHLT criteria. Restrictive phenotype included restrictive allograft syndrome (RAS) and mixed phenotype. In these cases, RAS-like opacities (RLOs) were evaluated both qualitatively and semiquantitatively on computed tomography at CLAD diagnosis. RLOs were classified into upper-predominant and diffuse/lower-predominant distribution groups. Overall survival after CLAD diagnosis was assessed using Kaplan-Meier method with log-rank test and Cox proportional hazards models. Eighty-three patients were diagnosed with CLAD after transplantation. Twenty-one (25.3%) had restrictive phenotype, which showed shorter survival compared to bronchiolitis obliterans syndrome (median survival: 19.8 vs 68.1months; hazard ratio [HR], 4.53; 95% confidence interval [CI], 1.96-10.49; p<0.001). Within the restrictive phenotype, the upper-predominant group demonstrated longer survival than the diffuse/lower-predominant group (median survival: 61.1 vs 15.5months; p=0.008). The diffuse/lower-predominant group had shorter survival compared to any other CLAD phenotype (HR, 8.45; 95% CI, 3.40-21.04; p<0.001). The extent of RLOs within each distribution pattern was not significantly associated with survival. In restrictive phenotype CLAD, RLO distribution patterns determined survival outcomes, with diffuse/lower-predominant showing the poorest prognosis, while the extent of RLOs within each pattern did not correlate with prognosis.

<b>Aims:</b> Long-term survival after lung transplantation (LT) is hampered by the occurrence of chronic lung allograft dysfunction (CLAD), manifesting as bronchiolitis obliterans syndrome (BOS) or restrictive allograft syndrome (RAS). CLAD is triggered by several factors, e.g. recurrent infections. As immunoglobulin (Ig) A is crucial to ensure mucosal immunity and limit airway microbial load, we explored whether IgA and its epithelial receptor, the polymeric Ig receptor (pIgR) are impaired in BOS. <b>Methods:</b> Bronchoalveolar lavages (BAL, n=120) from LT recipients included in the Cohort for Lung Transplantation were collected at pre-defined timepoints prior to the diagnosis of functional stability (BOS-free, n=30) or BOS (pre-BOS, n=30), and assessed for secretory (S)-IgA. Bronchiolar epithelium pIgR expression was quantified in transbronchial biopsies from BOS-free (n=20), pre-BOS (n=19) and BOS LT recipients (n=12), as well as in end-stage BOS explants (n=15). <b>Results:</b> S-IgA levels were reduced in BAL from pre-BOS LT recipients versus BOS-free (16.1 vs 33.4 µg/ml, p&lt;0.01). pIgR bronchiolar expression was reduced in transbronchial biopsies from BOS (p&lt;0.05 vs BOS-free and pre-BOS), with further decrease in end-stage BOS explants (p&lt;0.0001 vs BOS-free and pre-BOS). <b>Conclusions:</b> BAL S-IgA and pIgR decreased levels suggest that the pIgR/IgA system is impaired in BOS. This could play a pathogenic role by increasing susceptibility to local infections.

Background.Chronic rejection, defined as chronic lung allograft dysfunction (CLAD), is the major factor limiting long-term survival after lung transplantation (LTx). A specific subgroup of CLAD is restrictive allograft syndrome (RAS). CLAD’s pathogenesis is largely unknown, but previous findings suggest that it is associated with increased fibrosis in the transplanted lung. Cartilage oligomeric matrix protein (COMP) has been associated with multiple fibrotic conditions. The current study aimed to explore the relation between COMP serum levels and development of CLAD, and RAS in particular, in a retrospective cohort of LTx patients.Methods.This study included retrospective data from patients who underwent LTx during 2009–2011. Blood samples and spirometry data were obtained at follow-up visits 1, 3, 6, 9, and 12 mo after transplantation. Serum samples were analyzed for COMP. CLAD and RAS were defined according to the 2019 International Society for Heart and Lung Transplantation consensus document.Results.Data from 38 patients (19 men and women, respectively) were collected. Twenty-three patients (60.5%) developed CLAD, of whom 6 (26.1 %) fulfilled the criteria for RAS. Patients who developed RAS had higher mean COMP levels between 1 and 3 mo after LTx than those who did not develop RAS (10.9 [3.9-17.5] U/L vs 7.4 [3.9-10.8] U/L, P = 0.008). RAS was also associated with shorter survival. We found no association between COMP levels and CLAD of other types than RAS.Conclusions.Serum level of COMP early after LTx seems to be associated with RAS development and might serve as a biomarker suitable for clinical use in the LTx setting.

The emerging role of exosomal cargo in allograft rejection after lung transplantation

Chronic lung allograft dysfunction (CLAD) limits long-term survival after lung transplantation. Of the two subtypes, restrictive allograft syndrome (RAS) is characterized by a larger lung volume decrease and worse prognosis than bronchiolitis obliterans syndrome (BOS). We used computed tomography (CT) volumetry to classify CLAD subtypes and determined their clinical impact. Adult primary lung transplants performed 2003-2015 (n=167) were retrospectively evaluated for CLAD and subclassified with CT volumetry. Lung volume decrease of<15% from baseline resulted in BOSCT-vol and ≥15% resulted in RASCT-vol diagnosis. Clinical impact of CLAD subtypes was defined, and the prognostic value of different lung function, radiological, and lung volume parameters present at the time of CLAD diagnosis were compared. CLAD affected 43% of patients and was classified with CT volumetry as BOSCT-vol in 89% and RASCT-vol in 11%. Median graft survival estimate in RASCT-vol was significantly decreased compared to BOSCT-vol (1.6vs. 9.7 years, P=.038). At CLAD onset, RASCT-vol diagnosis (P=.05), increased lung density (P=.007), and more severe FEV1 (P=.004) decline from baseline, increased graft loss risk in multivariate analysis. CT volumetry serves to identify lung transplant patients with a poor clinical outcome but should be validated in prospective trials.

What we expected all along: Bronchiolitis obliterans syndrome is not specific for bronchiolitis obliterans in pediatric lung transplantation!

Chronic rejection of lung allografts has two major subtypes, bronchiolitis obliterans syndrome (BOS) and restrictive allograft syndrome (RAS), which present radiologically either as air trapping with small airways disease or with persistent pleuroparenchymal opacities. Parametric response mapping (PRM), a computed tomography (CT) methodology, has been demonstrated as an objective readout of BOS and RAS and bears prognostic importance, but has yet to be correlated to biological measures. Using a topological technique, we evaluate the distribution and arrangement of PRM-derived classifications of pulmonary abnormalities from lung transplant recipients undergoing redo-transplantation for end-stage BOS (N = 6) or RAS (N = 6). Topological metrics were determined from each PRM classification and compared to structural and biological markers determined from microCT and histopathology of lung core samples. Whole-lung measurements of PRM-defined functional small airways disease (fSAD), which serves as a readout of BOS, were significantly elevated in BOS versus RAS patients (p = 0.01). At the core-level, PRM-defined parenchymal disease, a potential readout of RAS, was found to correlate to neutrophil and collagen I levels (p < 0.05). We demonstrate the relationship of structural and biological markers to the CT-based distribution and arrangement of PRM-derived readouts of BOS and RAS.