Bleomycin in the setting of lung fibrosis induction: From biological mechanisms to counteractions

Abstract

Bleomycin (BLM) is a drug used to treat different types of neoplasms. BLM's most severe adverse effect is lung toxicity, which induces remodeling of lung architecture and loss of pulmonary function, rapidly leading to death. While its clinical role as an anticancer agent is limited, its use in experimental settings is widespread since BLM is one of the most widely used drugs for inducing lung fibrosis in animals, due to its ability to provoke a histologic lung pattern similar to that described in patients undergoing chemotherapy. This pattern is characterized by patchy parenchymal inflammation, epithelial cell injury with reactive hyperplasia, epithelial–mesenchymal transition, activation and differentiation of fibroblasts to myofibroblasts, basement membrane and alveolar epithelium injuries. Several studies have demonstrated that BLM damage is mediated by DNA strand scission producing single- or double-strand breaks that lead to increased production of free radicals. Up to now, the mechanisms involved in the development of pulmonary fibrosis have not been fully understood; several studies have analyzed various potential biological molecular factors, such as transforming growth factor beta 1, tumor necrosis factor alpha, components of the extracellular matrix, chaperones, interleukins and chemokines. The aim of this paper is to review the specific characteristics of BLM-induced lung fibrosis in different animal models and to summarize modalities and timing of in vivo drug administration. Understanding the mechanisms of BLM-induced lung fibrosis and of commonly used therapies for counteracting fibrosis provides an opportunity for translating potential molecular targets from animal models to the clinical arena.

Introduction

Bleomycin (BLM) is a chemotherapeutic agent used to treat several neoplastic diseases such as lymphomas, head and neck squamous cell carcinomas, testicular carcinoma, ovarian cancer and malignant pleural effusions. This drug was discovered in 1962 and described for the first time by Umezawa et al. in 1966 [1]; its structure was revised in 1978 and it was then confirmed by total synthesis in 1982 [2]. BLM is a water-soluble glycopeptidic antibiotic of approximately 1500 Da and it is part of a specific group of glycopeptide-derived natural products isolated from the bacterium Streptomyces verticillus [3], [4], [5], [6], [7]. This group includes over 200 closely related compounds, while the antineoplastic drug is a mixture of 11 molecules differing only in their N-terminal part. The best-known product used for this purpose is commercially available and comprises bleomycin A₂ (55–70%) and B₂ (25–32%); it does not induce myelosuppression and/or cardiotoxicity and for these reasons it can be administered either alone or in combination with Vinblastine and cis-diammine-dichloroplatinum(II) [6], [8]. Minor side effects of BLM treatment include nausea, vomiting, fever and occasional allergic-type reactions [8]. The most severe adverse effect is lung toxicity (Bleomycin Lung Toxicity – BLT), which may occur in up to 46% of the patients under treatment: for this reason the clinical use of BLM as anti-cancer agent in humans is limited [4], [5]. BLT occurrence is greater in patients older than 70 years and in those who have pre-existing lung disease or renal failure [9], [10], [11]. Symptoms and signs of BLT include cough, dyspnea, tachypnea, cyanosis, bibasal rales, pleural friction, intercostal retraction, reduced exercise tolerance and episodes of fever. These clinical signs are not specific for an underlying fibrotic process, which can be diagnosed on the basis of chest radiographs and high-resolution computed tomography (HRCT) scans. In patients undergoing BLM treatment, chest X-ray may show decreased lung volume and interstitial damage near the lung bases [12], [13]. Generally, HRCT chest scans show fibrotic changes in both lungs as reticular opacities (often irregularly distributed at basal, sub-pleural and peripheral lung level) and the typical honeycombing condition, features that characterize interstitial lung diseases (ILDs) as well as the subgroup of the idiopathic interstitial pneumonias (IIPs) [14]. Besides diseases with known etiology, IIPs also include disorders with unknown etiology such as idiopathic pulmonary fibrosis (IPF). IPF shows a histological pattern similar to that of usual interstitial pneumonia (UIP) and for this reason it is commonly referred to as IPF/UIP. According to ATS/ERS/JRS/ALAT 2011 guidelines [12], [13], when HRCT is not able to identify a clear fibrotic event, it is necessary to perform a surgical lung biopsy for a detailed histological examination. Histological and electron microscopy observations allow identification of the key features of UIP, such as a typical honeycombing aspect, the formation of activated fibroblast foci, the increased production and deposition of collagen from the extracellular matrix as well as interstitial fibrosis and scarring. The hypothetical progression of pathological events caused by BLT at the pulmonary level initially concerns endothelial and interstitial capillary edema, pneumocyte type II proliferation and surfactant overproduction, pneumocyte type II necrosis and surfactant release, surfactant phagocytosis and mediator release by alveolar macrophages. Subsequently, fibroblast proliferation and trans-differentiation to myofibroblasts (or activated fibroblasts) can occur [10], [11].

Conversely, BLT has been exploited in order to gain insight into the mechanisms of development and progression of pulmonary fibrosis. At present BLM administration is the most important and widely used method for inducing lung fibrosis in animal models. BLM is able to mediate DNA strand scission in presence of iron and oxygen, producing single- or double-strand breaks with consequent higher reactive oxygen species (ROS) and reactive nitrogen species (RNS) production [5], thus explaining the antineoplastic activity of the drug [3], [4], [15], [16]. Differences in the extent and distribution of BLM-induced lung fibrosis are dose-dependent. A single BLM dose causes sub-chronic changes, while repeated administrations lead to long-lasting lesions. In contrast to intravenous or intraperitoneal injections, which lead to sub-pleural scarring, intra-tracheal instillation of BLM determines bronchiolocentric accentuated fibrotic changes, acute interstitial and intra-alveolar inflammation, macrophage activation and up-regulation of tumor necrosis factor alpha (TNF-α), granulocyte-macrophage colony-stimulating factor (GM-CSF) and some interleukins. Following the acute inflammatory event, other cytokines such as transforming growth factor beta (TGF-β) and connective tissue growth factor (CTGF) are up-regulated during the repair and fibrotic stage. The aim of this paper is to review the existing literature on the specific characteristics of BLM-induced lung fibrosis in different animal models as well as to summarize modalities and timing of in vivo drug administration. Better comprehension of BLM-induced lung fibrosis could help us understand the biological mechanisms underlying lung fibrosis development. This approach, along with the knowledge of commonly used treatments to counteract fibrosis, may suggest new therapies, allowing the translation from animal models to the clinical arena.