Nariaki Kokuho

Pirfenidone inhibits fibrocyte accumulation in the lungs in bleomycin-induced murine pulmonary fibrosis

BackgroundBone marrow-derived fibrocytes reportedly play important roles in the pathogenesis of idiopathic pulmonary fibrosis. Pirfenidone is an anti-fibrotic agent; however, its effects on fibrocytes have not been investigated. The aim of this study was to investigate whether pirfenidone inhibits fibrocyte pool size in the lungs of bleomycin-treated mice.MethodsBleomycin (100 mg/kg) was infused with osmotic pumps into C57BL/6 mice, and pirfenidone (300 mg/kg/day) was orally administered daily for 2 wk. The lungs were removed, and single-cell suspensions were subjected to fluorescence-activated cell sorter (FACS) analysis to detect fibrocytes, which were defined as CD45 and collagen-I double-positive cells. Immunohistochemistry was performed on the lung specimens to quantify fibrocytes. Chemokines in the lung digests were measured with enzyme-linked immunosorbent assay. The effect of pirfenidone on alveolar macrophages was evaluated with bronchoalveolar lavage (BAL). In a therapeutic setting, pirfenidone administration was initiated 10 days after bleomycin treatment. For chemotaxis assay, lung fibrocytes were isolated with immunomagnetic selection (CD45-positive mesenchymal cells) after culture and allowed to migrate toward chemokines in the presence or absence of pirfenidone. Moreover, the effect of pirfenidone on the expression of chemokine receptors on fibrocytes was evaluated.ResultsPirfenidone significantly ameliorated bleomycin-induced pulmonary fibrosis as assessed with quantitative histology and collagen measurement. Fibrocyte pool size in bleomycin-treated mice lungs was attenuated from 26.5% to 13.7% by pirfenidone on FACS analysis. This outcome was also observed in a therapeutic setting. Immunohistochemistry revealed that fibrocytes were significantly decreased by pirfenidone administration compared with those in bleomycin-treated mice (P = 0.0097). Increased chemokine (CC motif) ligand-2 (CCL2) and CCL12 production in bleomycin-treated mouse lungs was significantly attenuated by pirfenidone (P = 0.0003 and P < 0.0001, respectively). Pirfenidone also attenuated macrophage counts stimulated by bleomycin in BAL fluid. Fibrocyte migration toward CCL2 and chemokine (CC motif) receptor-2 expression on fibrocytes was significantly inhibited by pirfenidone in vitro.ConclusionsPirfenidone attenuated the fibrocyte pool size in bleomycin-treated mouse lungs via attenuation of CCL2 and CCL12 production in vivo, and fibrocyte migration was inhibited by pirfenidone in vitro. Fibrocyte inhibition is considered a mechanism of anti-fibrotic action of pirfenidone.

Clinical features, anti-cancer treatments and outcomes of lung cancer patients with combined pulmonary fibrosis and emphysema

BACKGROUND Combined pulmonary fibrosis and emphysema (CPFE) patients may be at significantly increased risk of lung cancer compared with either isolated emphysema or pulmonary fibrosis patients. Acute exacerbation (AE) of interstitial lung disease caused by anticancer treatment is the most common lethal complication in Japanese lung cancer patients. Nevertheless, the clinical significance of CPFE compared with isolated idiopathic interstitial pneumonias (IIPs) in patients with lung cancer is not well understood. METHODS A total of 1536 patients with lung cancer at Nippon Medical School Hospital between March 1998 and October 2011 were retrospectively reviewed. Patients with IIPs were categorized into two groups: (i) CPFE; IIP patients with definite emphysema and (ii) non-CPFE; isolated IIP patients without definite emphysema. The clinical features, anti-cancer treatments and outcomes of the CPFE group were compared with those of the non-CPFE group. RESULTS CPFE and isolated IIPs were identified in 88 (5.7%) and 63 (4.1%) patients respectively, with lung cancer. AE associated with initial treatment occurred in 22 (25.0%) patients in the CPFE group and in 8 (12.7%) patients in the non-CPFE group, irrespective of treatment modality. Median overall survival (OS) of the CPFE group was 23.7 months and that of the non-CPFE group was 20.3 months (P=0.627). Chemotherapy was performed in a total of 83 patients. AE associated with chemotherapy for advanced lung cancer occurred in 6 (13.6%) patients in the CPFE group and 5 (12.8%) patients in the non-CPFE group. Median OS of the CPFE group was 14.9 months and that of the non-CPFE group was 21.6 months (P=0.679). CONCLUSION CPFE was not an independent risk factor for AE and was not an independent prognosis factor in lung cancer patients with IIPs. Therefore, great care must be exercised with CPFE as well as IIP patients when performing anticancer treatment for patients with lung cancer.

Diagnostic Values For Club Cell Secretory Protein (CC16) in Serum of Patients of Combined Pulmonary Fibrosis and Emphysema

Abstract Combined pulmonary fibrosis and emphysema (CPFE) is an under-recognized syndrome for which the diagnostic use of serum biomarkers is an attractive possibility. We hypothesized that CC16 and/or TGF-β1 or combinations with other biomarkers are useful for diagnosing CPFE. Patients with respiratory symptoms and a smoking history, with or without chronic obstructive pulmonary disease, were divided into the following three groups according to findings of high-resolution computed tomography of the chest: controls without either emphysema or fibrosis, patients with emphysema alone, and patients compatible with the diagnosis of CPFE. Serum concentrations of CC16, TGF-β1, SP-D, and KL-6 were measured in patients whose condition was stable for at least 3 months. To investigate changes in biomarkers of lung fibrosis in patients with a life-long smoking history, additional measurements were performed on the patients with idiopathic pulmonary fibrosis (IPF) of smoking history. The mean age of the first three groups was 68.0 years, whereas that of the IPF group was 71.8 years, and the groups contained 36, 115, 27, and 10 individuals, respectively. The serum concentration of CC16 in the four groups was 5.67 ± 0.42, 5.66 ± 0.35, 9.38 ± 1.04 and 22.15 ± 4.64 ng/ml, respectively, indicating that those patients with lung fibrosis had a significantly higher concentration. The combined use of CC16, SP-D, and KL-6 provided supportive diagnosis in conjunction with radiological imaging in diagnosis of CPFE. We conclude that a combination of biomarkers including CC16 could provide useful information to screen and predict the possible diagnosis of CPFE.

Nintedanib modulates surfactant protein-D expression in A549 human lung epithelial cells via the c-Jun N-terminal kinase-activator protein-1 pathway.

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with a high mortality rate. Signalling pathways activated by several tyrosine kinase receptors are known to be involved in lung fibrosis, and this knowledge has led to the development of the triple tyrosine kinase inhibitor nintedanib, an inhibitor of vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR), and fibroblast growth factor receptor (FGFR), for the treatment of IPF. Pulmonary surfactant protein D (SP-D), an important biomarker of IPF, reportedly attenuates bleomycin-induced pulmonary fibrosis in mice. In this study, we investigated whether nintedanib modulates SP-D expression in human lung epithelial (A549) cells using quantitative real-time reverse transcriptase polymerase chain reaction and western blotting. To investigate the mechanisms underlying the effects of nintedanib, we evaluated the phosphorylation of c-Jun N-terminal kinase (JNK) and its downstream target c-Jun. The effect of the JNK inhibitor SP600125 on c-Jun phosphorylation was also tested. Activation of activator protein-1 (AP-1) was examined using an enzyme-linked immunosorbent assay-based test, and cell proliferation assays were performed to estimate the effect of nintedanib on cell proliferation. Furthermore, we treated mice with nintedanib to examine its in vivo effect on SP-D levels in lungs. These experiments showed that nintedanib up-regulated SP-D messenger RNA expression in a dose-dependent manner at concentrations up to 5 μM, with significant SP-D induction observed at concentrations of 3 μM and 5 μM, in comparison with that observed in vehicle controls. Nintedanib stimulated a rapid increase in phosphorylated JNK in A549 cells within 30 min of treatment and stimulated c-Jun phosphorylation, which was inhibited by the JNK inhibitor SP600125. Additionally, nintedanib was found to activate AP-1. A549 cell proliferation was not affected by nintedanib at any of the tested concentrations. Moreover, blocking FGFR, PDGFR, and VEGFR function did not affect nintedanib-induced SP-D expression, suggesting that nintedanib mediates its effects through a mechanism that is distinct from its known role as a tyrosine kinase inhibitor. Nintedanib is also reported to inhibit Src kinase although pre-treatment of cells with a Src kinase inhibitor had no effect on nintedanib-induced SP-D expression. Increased expression of SFTPD mRNA and SP-D protein in the lungs of nintedanib-treated mice was also observed. In this work, we demonstrated that nintedanib up-regulated SP-D expression in A549 cells via the JNK-AP-1 pathway and did not affect cell proliferation. This is the first report describing SP-D induction by nintedanib.

Fatal Pneumonia Associated with Temozolomide Therapy in Patients with Malignant Glioma

This report presents the cases of three patients with fatal pneumonia that was highly suspected to be Pneumocystis pneumonia (PCP) based on serological diagnosis. Their chest radiographs showed bilateral pneumonia and each had presented with severe respiratory failure requiring mechanical ventilation when they arrived at the hospital. Although bronchoscopical sampling could not be performed, their chest computed tomography imaging and a marked elevation of serum KL-6 and β-D-glucan levels were characteristic of Pneumocystis pneumonia. All three were found to have been treated with temozolomide after surgery for malignant glioma. Temozolomide can cause Pneumocystis pneumonia. The three patients did not receive prophylactic medication against Pneumocystis pneumonia during treatment with temozolomide, and their histories suggested that all had delayed seeking treatment. It may be difficult to diagnose Pneumocystis pneumonia because the symptoms are not specific for Pneumocystis pneumonia and they tend to be similar to those of common respiratory infectious diseases. Therefore, patients who receive temozolomide therapy have the potential to develop fatal pneumonia and should be carefully observed. The patients should also be adequately informed about Pneumocystis pneumonia, and prophylaxis against Pneumocystis pneumonia should be considered proactively before treatment with temozolomide is initiated.

Elotuzumab-induced interstitial lung disease: the first case report

Elotuzumab, a humanized immunoglobulin G1 monoclonal antibody targeted against signaling lymphocytic activation molecule F7 (SLAMF7), has recently been used in combination with lenalidomide and dexamethasone for the treatment of patients with relapsed or refractory multiple myeloma. The clinical characteristics of drug-induced interstitial lung disease (ILD) due to elotuzumab have not been clarified. In this report, we describe a patient with refractory multiple myeloma who received elotuzumab in combination with lenalidomide and dexamethasone in whom fatigue, fever and diffuse pulmonary infiltration developed. The patient had a history of long-term therapy with lenalidomide without pulmonary toxicity. Bronchoscopy with bronchoalveolar lavage was negative for infection, and transbronchial lung biopsies showed active alveolitis with lymphocytic infiltration and myxomatous change of the thick alveolar wall. After the discontinuation of elotuzumab and lenalidomide, the patients clinical symptoms gradually improved, and spontaneous remission of the pulmonary infiltration was observed. Based on the chest CT and lung pathology findings, the exclusion of infection and pulmonary edema, and according to the clinical course, we established a diagnosis of drug-induced ILD due to elotuzumab. Clinicians should bear in mind the potential for pulmonary toxicity in patients receiving elotuzumab-containing therapy.

Prognostic Factors in the Acute Exacerbation of Idiopathic Pulmonary Fibrosis: A Retrospective Single-center Study

Objectives Acute exacerbation of idiopathic pulmonary fibrosis (IPF-AE) has been recognized as a fatal pulmonary disorder, but the exact prognostic factors are unknown. The aim of the present study was to analyze the clinical characteristics of patients with IPF-AE and identify the prognostic factors. Methods The medical records of 59 cases of IPF-AE were retrospectively reviewed. Clinical data, laboratory data, radiographic findings, treatment, and time from the onset of symptoms to the initiation of corticosteroid pulse therapy, i.e. symptom duration, and outcome were analyzed. Results The IPF Stage, Gender-Age-Physiology (GAP) Index, symptom duration, and the high-resolution computed tomography (HRCT) score were significantly related to the prognosis in the univariate analysis. In the multivariate analysis, the symptom duration remained a significant prognostic factor (hazard ratio of 1-day increase, 1.11; 95% confidence interval, 1.01-1.15; p=0.0427). The area under the receiver operating characteristics curve of symptom duration was statistically significant for survivors versus non-survivors (area under the curve, 0.73; p=0.012). The survival period was significantly shorter in the late-treatment groups (≥5 days; n=30) than in the early-treatment groups (<5 days; n=29; log-rank test; p<0.0001). Conclusion The time interval between the onset of symptoms and the initiation of corticosteroid pulse therapy may be an independent prognostic factor in patients with IPF-AE.

Granuloma-forming interstitial pneumonia induced by nivolumab: a possible immune-related adverse event of the lung

Nivolumab, a monoclonal antibody targeting the PD-1, has recently been used as a standard treatment for lung cancer, melanoma and renal cell carcinoma. We herein report the case of a patient undergoing treatment for non-small cell lung cancer (NSCLC) who developed interstitial pneumonia which featured nivolumab-induced granuloma formation. An 82-year-old male patient with NSCLC was initially treated with radiation therapy and chemotherapy. Five years later, however, he developed metastatic carcinoma in a hilar lymph node accompanied by ground glass opacity (GGO), suggesting tumor cell invasion. Treatment with nivolumab was initiated. At 21 days after the first dose of nivolumab, he complained of cough and dyspnea. Chest computed tomography scans demonstrated tumor progression and newly formed GGO in the area surrounding the primary tumor. Fibrosing active alveolitis with granuloma formation and organizing pneumonia findings were observed in the pathological examination of a transbronchial lung biopsy (TBLB) specimen. No malignant cells were found in TBLB. A bacteriological analysis of cultures, a PCR, and special staining did not reveal any infections. The patient’s pneumonitis improved after treatment with systemic corticosteroids. Granuloma-forming interstitial pneumonia may be a feature of nivolumab-associated pneumonitis.

Localized pulmonary crystal-storing histiocytosis complicating pulmonary mucosa-associated lymphoid tissue lymphoma presenting with multiple mass lesions

Crystal-storing histiocytosis (CSH) is an uncommon finding in lymphoplasmacytic disorders that presents histiocytes with abnormal intralysosomal accumulations of immunoglobulin light chains as crystals of unknown etiology. A 38-year-old woman with antiphospholipid syndrome had a surgical lung biopsy because of multiple lung mass lesions. In a right middle lobe lesion, lymphoplasmacytic cells had a monocytoid appearance, destructive lymphoepithelial lesions, and positive immunoglobulin heavy chain (IGH) gene rearrangements. A right upper lobe lesion manifested proliferating rounded histiocytes with abundant, deeply eosinophilic cytoplasm and negative IGH gene rearrangements. Electron microscopy and mass spectrometry revealed a case of pulmonary CSH: abnormal proliferation of the immunoglobulin κ chain of a variable region that may be crystallized within plasma cells and histiocytes. We report a rare case of localized pulmonary CSH complicating pulmonary mucosa-associated lymphoid tissue lymphoma with multiple mass lesions. We demonstrate advances in the understanding of the pathogenesis of CSH by various analyses of these lesions.