Molecular Assessment of Single-Piece Mucosal Biopsies is Highly Reproducible Compared to Transbronchial Biopsies in Lung Transplantation

Purpose Histologic assessment for rejection on transbronchial biopsies (TBB) has poor reproducibility. Mucosal biopsies are safer than TBBs but their histologic assessment has not been systematically studied or standardized. We developed molecular platforms to assess TBBs and mucosal biopsies from the third airway bifurcation (3BMB). We examined the reproducibility of scores assigned by molecular assessment of both biopsy types in lung transplant patients from 9 international centers. Methods We trained machine learning models (archetypal analysis, principal component analysis) on microarray data from 457 TBBs and 243 3BMBs. We processed 14 pairs of TBB pieces (28 total) taken during the same procedure and 13 pairs of 3BMBs (26 total). The models assigned each biopsy sets of archetype scores (denoted by “R”) and principal component scores (denoted by “PC”) describing injury and rejection phenotypes. We studied within-score agreement between biopsy pairs using intraclass correlations (ICC). Results Most scores had excellent reproducibility in 3BMB pairs compared to TBBs (Table 1). In 3BMBs the R1 (normal), R2 (rejection), R3 (late injury), PC1 (inflammation/rejection), and PC2 (late injury) scores were highly reproducible, but R4 and PC3 (both recent injury) were not. In TBBs the R2 (rejection) and PC1 (inflammation/rejection) scores were moderately reproducible; other scores had poor reproducibility. Conclusion Molecular assessment of paired, single piece 3BMBs achieves excellent reproducibility in its key scores. TBB scores were less reproducible likely because of tissue heterogeneity and greater anatomic distance between samples. Measurements could be stabilized by combining ≥2 pieces per microarray. Importantly, molecular assessment with 3BMBs or TBBs requires fewer pieces (2-3) than TBB histology, which requires up to ten. These data support continued study of the diagnostic and prognostic utility of 3BMBs in lung transplantation.