Beata Kosmider

Enhanced P53 and BAX gene expression and apoptosis in A549 cells by cis-Pt(II) complex of 3-aminoflavone in comparison with cis-DDP.

Lung cancer remains one of the most common causes of cancer-related death worldwide. Approximately 80% is histologically non-small cell lung carcinoma (NSCLC) and in about 70% of patients it is an unresectable type. Clinical studies indicated that application of platinum derivatives caused good results and combinations of platinum with other agents could improve median survivals. In view of the central problem of sufficient efficiency of drugs in chemotherapy, efforts have focused on the development of alternative platinum-based analogues that can be more effective in cancer treatment. cis-bis(3-aminoflavone)dichloroplatinum(II) (cis-Pt(II) complex of 3-aminoflavone) represents a novel class of platinum-based potential antitumour agents. In order to evaluate the degree of apoptosis, acridine orange/ethidium bromide and Hoechst 33258/propidum iodide double staining as well as RT-PCR (P53 and BAX expression evaluation) were used in lung cancer cell line A549 after treatment with this compound in comparison with cis-diamminedichloroplatinum(II) (cis-DDP). Apoptotic cells at early and late stages and also necrotic ones were observed after usage of cis-Pt(II) complex of 3-aminoflavone and the percentage of these cells outnumbered the values obtained after cis-DDP application. The former compound induced a higher percentage of P53 and BAX expression in A549 cells in comparison with the latter one. Results indicate the beneficial properties of cis-Pt(II) complex of 3-aminoflavone as a potential antitumor drug.

DJ-1 Modulates Nuclear Erythroid 2–Related Factor-2–Mediated Protection in Human Primary Alveolar Type II Cells in Smokers

Cigarette smoke (CS) is a main source of oxidative stress and a key risk factor for emphysema, which consists of alveolar wall destruction. Alveolar type (AT) II cells are in the gas exchange regions of the lung. We isolated primary ATII cells from deidentified organ donors whose lungs were not suitable for transplantation. We analyzed the cell injury obtained from nonsmokers, moderate smokers, and heavy smokers. DJ-1 protects cells from oxidative stress and induces nuclear erythroid 2-related factor-2 (Nrf2) expression, which activates the antioxidant defense system. In ATII cells isolated from moderate smokers, we found DJ-1 expression by RT-PCR, and Nrf2 and heme oxygenase (HO)-1 translocation by Western blotting and immunocytofluorescence. In ATII cells isolated from heavy smokers, we detected Nrf2 and HO-1 cytoplasmic localization. Moreover, we found high oxidative stress, as detected by 4-hydroxynonenal (4-HNE) (immunoblotting), inflammation by IL-8 and IL-6 levels by ELISA, and apoptosis by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay in ATII cells obtained from heavy smokers. Furthermore, we detected early DJ-1 and late Nrf2 expression after ATII cell treatment with CS extract. We also overexpressed DJ-1 by adenovirus construct and found that this restored Nrf2 and HO-1 expression and induced nuclear translocation in heavy smokers. Moreover, DJ-1 overexpression also decreased ATII cell apoptosis caused by CS extract in vitro. Our results indicate that DJ-1 activates the Nrf2-mediated antioxidant defense system. Furthermore, DJ-1 overexpression can restore the impaired Nrf2 pathway, leading to ATII cell protection in heavy smokers. This suggests a potential therapeutic strategy for targeting DJ-1 in CS-related lung diseases.

Lipidomic characterization and localization of phospholipids in the human lung

Lipids play a central role in lung physiology and pathology; however, a comprehensive lipidomic characterization of human pulmonary cells relevant to disease has not been performed. The cells involved in lung host defense, including alveolar macrophages (AMs), bronchial epithelial cells (BECs), and alveolar type II cells (ATIIs), were isolated from human subjects and lipidomic analysis by LC-MS and LC-MS/MS was performed. Additionally, pieces of lung tissue from the same donors were analyzed by MALDI imaging MS in order to determine lipid localization in the tissue. The unique distribution of phospholipids in ATIIs, BECs, and AMs from human subjects was accomplished by subjecting the large number of identified phospholipid molecular species to univariant statistical analysis. Specific MALDI images were generated based on the univariant statistical analysis data to reveal the location of specific cell types within the human lung slice. While the complex composition and function of the lipidome in various disease states is currently poorly understood, this method could be useful for the characterization of lipid alterations in pulmonary disease and may aid in a better understanding of disease pathogenesis.

Abstract 1327: CD147 contributes to extracellular matrix remodeling and degradation

Chronic obstructive pulmonary disease (COPD) is an incurable, progressive lung disease characterized by shortness and difficulty in breathing. Patients with COPD are at higher risk of developing lung cancer and often are associated with poor outcome of lung cancer diagnosis and treatment. Extracellular matrix (ECM) remodeling and degradation is one of the common hallmarks between cancer and COPD. It contributes to degradation of air sacs (emphysema) or small airways (obstructive bronchiolitis). It is also involved in cancer initiation and progression to assist tumor expansion and cancer cell migration. In this study, we analyzed lung tissue and primary alveolar type II (ATII) cells isolated from patients with emphysema and control organ donors. ATII cells secrete and store pulmonary surfactant and restore the epithelium after damage. We performed RNA sequencing in ATII cells to identify dysregulated proteins, which are involved in ECM degradation in emphysema. To further study the mechanism involved in the ECM degradation in this disease, we analyzed the expression of selected proteins and we focused on the role of extracellular matrix metalloproteinase inducer (CD147). CD147 is a cell membrane protein involved in inducing the production of extracellular matrix metalloproteinases (MMPs). First, we found that cells isolated from patients with emphysema have higher expression of CD147 in ATII cells in comparison with controls as detected by western blotting and RT-PCR. This may explain high ECM degradation in this disease. Second, we observed higher MMPs levels in plasma obtained from individuals with this disease. Third, we found significantly higher expression of cathepsin B in ATII cells isolated from emphysema in comparison with controls. Cathepsin B is an enzyme that facilitates a direct degradation of ECM proteins and activate other proteases capable of degrading ECM. Fourth, our data indicate that CD147 interacts with cathepsin B, which may lead to the activation of MMPs and emphysema progression. In summary, our results show for the first time the important role of CD147 in emphysema development. Moreover, we found that cathepsin B may contribute to CD147-mediated disease progression. Further studies will elucidate its role in lung cancer development in patients with emphysema. Note: This abstract was not presented at the meeting. Citation Format: Tiffany Tan, Beata Kosmider, Nathaniel Marchetti, Sudhir Bolla, Chenna Mandapati, Gerard Criner, Karim Bahmed. CD147 contributes to extracellular matrix remodeling and degradation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1327. doi:10.1158/1538-7445.AM2017-1327

Structure-activity relationship and apoptosis induction in A549 cells by the potential anticancer compound cis-bis(3-aminoflavone)dichloroplatinum(II)

cis-DDP (cis-diamminedichloroplatinum(II), CAS 15663-27-1) is widely used in chemotherapy of many types of cancer. However, besides effectiveness, it gives many side effects which limit its clinical application. Therefore, nowadays studies are focused on searching for novel analogs of cis-DDP, at least equally effective in chemotherapy but less toxic. One of them might be cis-BAFDP (cis-bis(3-aminoflavone)dichloroplatinum(II)) with one of the hydrogen atoms of the amino group of cis-DDP replaced by a flavone ring. 3-Aminoflavone (AF) which posseses the desired NH2 group has been used as non-leaving ligand. The complex has been obtained in the reaction of AF and K2PtCl4. There are no data concerning evaluation of structural studies of cis-BAFDP, the beneficial anticancer properties of which were proved in vitro and in vivo. Therefore it was worthwhile to undertake a confirmation of the chemical structure of this compound by applying various spectroscopic techniques especially because of its potential pharmacological application. With this aim in mind this compound was characterized by: IR, 1H NMR, 195Pt NMR, UV absorption and fluorescence spectroscopy. Moreover, stronger apoptosis induction by cis-BAFDP than cis-DDP in the human nonsmall cancer cell line A549 was observed using Hoechst 33258/propidium iodide double staining.