Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal interstitial lung disease. The pathogenesis of interstitial lung diseases, including its most common form, IPF, remains poorly understood. Alveolar epithelial cell (AEC) apoptosis, proliferation, and accumulation of myofibroblasts and extracellular matrix deposition results in progressive loss of lung function in IPF. We found induction of tumor suppressor protein, p53, and apoptosis with suppression of urokinase-type plasminogen activator (uPA) and the uPA receptor in AECs from the lungs of IPF patients, and in mice with bleomycin, cigarette smoke, silica, or sepsis-induced lung injury. Treatment with the caveolin-1 scaffolding domain peptide (CSP) reversed these effects. Consistent with induction of p53, AECs from IPF lungs or mice with diverse types of lung injuries showed increased p53 acetylation and miR-34a expression with reduction in Sirt1. This was significantly reduced after treatment of wild-type mice with CSP, and uPA-deficient mice were unresponsive. Bleomycin failed to induce miR-34a in p53- or plasminogen activator inhibitor-1 (PAI-1)-deficient mice. CSP-mediated inhibition of miR-34a restored Sirt1, suppressed p53 acetylation and apoptosis in injured AECs, and prevented pulmonary fibrosis (PF). AEC-specific suppression of miR-34a inhibited bleomycin-induced p53, PAI-1, and apoptosis and prevented PF, whereas overexpression of precursor-miR-34a increased p53, PAI-1, and apoptosis in AECs of mice unexposed to bleomycin. Our study validates p53-miR-34a feedback as a potential therapeutic target in PF.

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease with a median 5-year survival of ~20%. Current U.S. Food and Drug Administration–approved pharmacotherapies slow progression of IPF, providing hope that even more effective treatments can be developed. Alveolar epithelial progenitor type II cell (AEC) apoptosis and proliferation, and accumulation of activated myofibroblasts or fibrotic lung fibroblasts (fLfs) contribute to the progression of IPF. Full-length caveolin-1 scaffolding domain peptide (CSP; amino acids 82 to 101 of Cav1: DGIWKASFTTFTVTKYWFYR) inhibits AEC apoptosis and fLf activation and expansion and attenuates PF in bleomycin (BLM)–induced lung injury in mice. Like full-length CSP, a seven–amino acid deletion fragment of CSP, CSP7 (FTTFTVT), demonstrated antifibrotic effects in murine models of lung fibrosis. When CSP7 was administered during the fibrotic phase in three preclinical models [single-dose BLM, repeated-dose BLM, and adenovirus expressing constitutively active transforming growth factor–β1 (Ad-TGF-β1)–induced established PF], CSP7 reduced extracellular matrix (ECM) markers characteristic of PF, increased AEC survival, and improved lung function. CSP7 is amenable to both systemic (intraperitoneal) or direct lung delivery in a nebulized or dry powder form. Furthermore, CSP7 treatment of end-stage human IPF lung tissue explants attenuated ECM production and promoted AEC survival. Ames testing for mutagenicity and in vitro human peripheral blood lymphocyte and in vivo mouse micronucleus transformation assays indicated that CSP7 is not carcinogenic. Together, these findings support the further development of CSP7 as an antifibrotic treatment for patients with IPF or other interstitial lung diseases.

Our study adopted a comprehensive exploration of genetic changes using high resolution molecular probes applied to the MCF10 family of cell lines to identify individual genes in a continuum starting from normal breast epithelial cells and progressing through immortalization, transformation and invasive malignancy. Homozygous loss of CDKN2A and CDKN2B genes and gain of MYC were initiating immortalization events. Transformation and progression to malignancy event were marked by gains of IL13, VEGF, HRAS, TRAF2, and BCAS2, IL12A, and MME, respectively.

Objective: To identify the extent and the smallest region of loss for CDKN2B INK4b , CDKN2A ARF,INK4a , and MTAP. Homozygous deletions of human chromosome 9p21 occur frequently in malignant cell lines and are common in squamous cell carcinoma of the head and neck (HNSCC). This complex region encodes the tumor suppressor genes cyclin-dependent kinase 2B (CDKN2B) (p15 INK4b ) and CDKN2A (p14 ARF , p16 INK4a ) and the housekeeping gene methylthioadenosine phosphorylase (MTAP).Design: A targeted probe panel designed to finely map the region of 9p21 loss comprised 3 probes for CDKN2B INK4b , 7 for CDKN2A ARF, INK4a , and 3 for MTAP and was interrogated using the multiplex ligationdependent probe amplification assay (MLPA). The MLPA genomic copy number alterations for CDKN2A were validated using real-time polymerase chain reaction. Subjects: Six HNSCC primary (A) and recurrent or metastatic (B) cell lines were examined: UMSCC-11A/11B, UMSCC-17A/17B, and UMSCC-81A/81B. Results: Cell line UMSCC-11B retained all 9p loci tested in the region. Cell lines UMSCC-17A/B indicated homozygous deletion of CDKN2A ARF, INK4a starting at p16 INK4 exon 1␣ to include exons 2 and 3. Homozygous loss was indicated for CDKN2B INK4b and CDKN2A ARF,INK4a in UMSCC-11A, and UMSCC-81A. Cell line UMSCC-81B indicated retention of all 9p loci except for exon 1␣ (p16 INK4a ). Selective loss of the 3Ј end of MTAP was observed in UMSCC-11A. Genomic alterations by fine-mapping MLPA were validated at the DNA level for CDKN2A.Conclusions: We identified exon 1␣ (p16 INK4a ) as the smallest region of loss in the CDKN2A ARF, INK4a gene. The frequency and precise loss of CDKN2B INK4b , CDKN2A ARF, INK4a , and MTAP in the prognosis of 9p21-deleted HNSCC may provide impetus for use of these targets as therapeutic biomarkers in head and neck cancer.

I. have patents issued for the use of the caveolin-1 spanning domain and its fragments for the treatment of pulmonary fibrosis (869,784,0B2 and 9,630,990); S.I. is an unpaid member of the board of directors and Chief Scientific Officer of Lung Therapeutics, Inc., a University of Texas start-up biotechnology firm that is commercializing caveolin-1 scaffolding domain peptide 7 for the treatment of idiopathic pulmonary fibrosis; S.I. has an equity position in the firm as a founder and investor; S.S. is a consultant for