Elsevier

Journal of Controlled Release

Volume 213, 10 September 2015, Pages 112-119
Journal of Controlled Release

Pulmonary administration of phosphoinositide 3-kinase inhibitor is a curative treatment for chronic obstructive pulmonary disease by alveolar regeneration

https://doi.org/10.1016/j.jconrel.2015.07.004Get rights and content

Abstract

Chronic obstructive pulmonary disease (COPD) is an intractable pulmonary disease, causing widespread and irreversible alveoli collapse. The discovery of a low-molecular-weight compound that induces regeneration of pulmonary alveoli is of utmost urgency to cure intractable pulmonary diseases such as COPD. However, a practically useful compound for regenerating pulmonary alveoli is yet to be reported. Previously, we have elucidated that Akt phosphorylation is involved in a differentiation-inducing molecular mechanism of human alveolar epithelial stem cells, which play a role in regenerating pulmonary alveoli. In the present study, we directed our attention to phosphoinositide 3-kinase (PI3K)-Akt signaling and examined whether PI3K inhibitors display the pulmonary alveolus regeneration. Three PI3K inhibitors with different PI3K subtype specificities (Wortmannin, AS605240, PIK-75 hydrochloride) were tested for the differentiation-inducing effect on human alveolar epithelial stem cells, and Wortmannin demonstrated the most potent differentiation-inducing activity. We evaluated Akt phosphorylation in pulmonary tissues of an elastase-induced murine COPD model and found that Akt phosphorylation in the pulmonary tissue was enhanced in the murine COPD model compared with normal mice. Then, the alveolus-repairing effect of pulmonary administration of Wortmannin to murine COPD model was evaluated using X-ray CT analysis and hematoxylin–eosin staining. As a result, alveolar damages were repaired in the Wortmannin-administered group to a similar level of normal mice. Furthermore, pulmonary administration of Wortmannin induced a significant recovery of the respiratory function, compared to the control group. These results indicate that Wortmannin is capable of inducing differentiation of human alveolar epithelial stem cells and represents a promising drug candidate for curative treatment of pulmonary alveolar destruction in COPD.

Introduction

Emphysema and chronic bronchitis that cause respiratory failure due to destruction of alveolar structures are collectively referred to as chronic obstructive pulmonary disease (COPD), which is currently the fourth leading cause of death worldwide [1]. The number of patients with COPD reportedly exceeds 200 million in the world including the potential ones on the basis of epidemiological surveys by WHO (Global Alliance Against Chronic Respiratory Diseases (GARD)). However, no therapeutic drug is available for curative treatment of COPD, and the development of a therapeutic agent to repair alveolar destruction is of particular urgency.

The targets of human lung alveolar remodeling are resident stem and progenitor cells in the lung that function in tissue repair and homeostasis. The adult lung consists of the following four major biologically distinct components: the trachea, bronchi, bronchioles, and alveoli. Each component is biologically distinct and has its own stem and progenitor population [2], [3], [4], [5]. Alveoli are terminal structures of distal airways specialized for gas exchange. The gaseous alveolar surface is lined by alveolar type I cells (AT-I) and alveolar type II cells (AT-II) [6]. Recent studies have characterized resident alveolar stem cells in human [7]. However, potent regenerative compounds have not been identified for human alveolar stem cells.

We identified a differentiation-inducing agent for human alveolar epithelial stem cells responsible for the regeneration of pulmonary alveoli, and further studied its differentiation-inducing molecular mechanism. Based on this study, for first time, we reported that suppression of the phosphorylation of Ser/Thr kinase Akt leads to differentiation of human alveolar epithelial stem cells into AT-I and AT-II, which constitute pulmonary alveoli [8]. In this study, we focus on phosphoinositide 3-kinase (PI3K)-Akt signaling pathway. PI3K is categorized into class I, class II, and class III according to its primary structure and substrate specificity, with class I being involved in cellular survival and differentiation [9]. Class I PI3K is further divided into four subunits, α and β subunits involved in cellular survival, δ and γ subunits reported to be involved in inflammation [10], [11]. While PI3K-δ and PI3K-γ have been reported to suppress inflammation in COPD [12], [13], their differentiation-inducing effect on alveolar epithelial stem cells has yet to be elucidated.

Thus, in this study, we focused on phosphoinositide 3-kinase (PI3K), which phosphorylates Akt, and examined whether PI3K inhibitors induced the differentiation of human alveolar epithelial stem cells. We further examined the effectiveness of the PI3K inhibitors on the repair of pulmonary alveoli and improvement of respiratory function in a murine COPD model.

Pulmonary administration is a drug delivery system (DDS) that is superior to other methods of drug administration in delivering a drug to the lung. Previously, the efficacy of PI3K inhibitors has been studied primarily in oral administration, and no study has been reported on their pharmacological effects, including the representative antitumor effect, through pulmonary administration. This study is the first to report the effect of PI3K inhibitors on the lung when they are delivered by pulmonary administration. We demonstrated the alveolus-repairing effect of PI3K inhibitors in mice using a pulmonary administration method of breath-actuated inhalation, and found the possibility of an additional indication of PI3K inhibitors for COPD.

Section snippets

Animals and cells

Male ICR mice were purchased from Sankyo Labo Service Corporation (Tokyo, Japan). Animals were housed in a temperature-controlled (24 ± 1 °C) facility maintained on a light (12 h):dark (12 h) cycle with standard food available ad libitum. All animal procedures followed the guidelines established by the Animal Care and Use Committee of the Tokyo University of Science.

Six-week-old male mice were anesthetized with isoflurane, and a solution of porcine pancreatic elastase (Elastin Products Company,

Results

First, we tested three PI3K inhibitors (AS605240, PIK-75 hydrochloride, Wortmannin) to evaluate their differentiation-inducing effect on human alveolar epithelial stem cells. AS605240 and PIK-75 hydrochloride are inhibitors highly specific to PI3K-γ and PI3K-α [15], [16], respectively, whereas Wortmannin is a PI3K inhibitor with low substrate specificity. The differentiation-inducing effects of the three PI3K inhibitors on human alveolar epithelial stem cells were evaluated by counting cells

Discussion

To the best of our knowledge, our results are the first to demonstrate a high potential of PI3K inhibitors as a novel class of therapeutic agents for regenerating pulmonary alveoli by inducing the differentiation of human alveolar epithelial stem cells.

The results from evaluation of the distribution and pulmonary pharmacological effect of the drugs administered via pulmonary, oral, and intraperitoneal routes demonstrated the effectiveness of pulmonary administration as a means for local drug

Conclusions

PI3K inhibitors were found to induce the differentiation of human alveolar epithelial stem cells, and Wortmannin, which was the most potent differentiation inducer, was shown to indeed repair pulmonary alveoli and recover respiratory function in murine COPD model. Wortmannin was novel curative treatment of pulmonary alveolar destruction in COPD.

Acknowledgments

Human alveolar epithelial stem cells were provided by Dr. Hiroshi Kubo (Tohoku University, Sendai, Japan). This study was partially supported by the Grant-in-Aid for Research Activity Start-up [Michiko Horiguchi, 24890257], and the Grant-in-Aid for Young Scientists (B) [Michiko Horiguchi, 25860029] from the Japan Society for the Promotion of Science.

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