Bronchial premalignant lesions (PMLs) are precursors of lung squamous cell carcinoma, but have variable outcome, and we lack tools to identify and treat PMLs at risk for progression to cancer. Here we report the identification of four molecular subtypes of PMLs with distinct differences in epithelial and immune processes based on RNA-Seq profiling of endobronchial biopsies from high-risk smokers. The Proliferative subtype is enriched with bronchial dysplasia and exhibits up-regulation of metabolic and cell cycle pathways. A Proliferative subtype-associated gene signature identifies subjects with Proliferative PMLs from normal-appearing uninvolved large airway brushings with high specificity. In progressive/persistent Proliferative lesions expression of interferon signaling and antigen processing/presentation pathways decrease and immunofluorescence indicates a depletion of innate and adaptive immune cells compared with regressive lesions. Molecular biomarkers measured in PMLs or the uninvolved airway can enhance histopathological grading and suggest immunoprevention strategies for intercepting the progression of PMLs to lung cancer.

Fibrotic lung diseases are characterized by excess extracellular matrix production, in particular type I collagen. Phenylbutyrate (PB) is a non-toxic pharmacological compound that functions as a weak histone deacetylase inhibitor. In hepatic stellate cells, the synthesis of type I collagen expression is decreased by inhibiting histone acetylation. Our studies examined the regulation of type I collagen by PB in human lung fibroblasts. We found that PB decreases basal and transforming growth factor-beta-stimulated alpha1(I) collagen mRNA and protein levels. Northern blot analyses demonstrated that PB decreases steady-state alpha1(I) collagen mRNA levels by 78% without significantly changing the stability of the mRNA transcript. PB stimulates cAMP production and increases the acetylation of histone H4, but does not affect the activity of two transforming growth factor-beta (TGF-beta)-responsive luciferase reporter constructs. These data suggest that PB regulates type I collagen expression in human lung fibroblasts by mechanisms that include cAMP production and histone acetylation. PB may have therapeutic use in fibrotic lung diseases.

The described micro-CT cryostage provides a novel way to study the three-dimensional lung structure preserved without the effects of fixatives while enabling subsequent studies of the cellular matrix composition and gene expression. This approach will, for the first time, enable researchers to study structural changes of lung tissues that occur with disease and correlate them with changes in gene or protein signatures.