Fibulin-5 as a potential therapeutic target in COPD

KEYWORDS:

• COPD
• emphysema
• elastic fiber
• fibrosis
• fibulin-5

1. Fibulin-5 and its relation with disease

Fibulin-5 is an extracellular matrix (ECM) protein containing six calcium-binding epidermal growth factor-like domains, including an integrin-binding tripeptide Arg-Gly-Asp sequence. Fibulin-5 is strongly expressed during vascular development and in elastic fiber-rich tissues, including aorta, lung, uterus, and skin in adults. [1] Studies using fibulin-5 knockout mice have demonstrated that fibulin-5 is present in elastic fibers and is critical for elastogenesis in vivo. These mice survive to adulthood but suffer from severe elastinopathy, resulting in loose skin, vascular abnormalities, and severe emphysema, [2,3] a phenotype very similar to fibulin-5-related cutis laxa syndrome in humans. [4]

In addition to this clear role in elastic fiber-associated pathologies, fibulin-5 has also been associated with chronic obstructive pulmonary disease (COPD) and fibrotic (lung) disease. In a large gene expression analysis of 581 human lung tissue samples, we previously identified fibulin-5 as the most upregulated gene in lung tissue from patients with COPD. [5] COPD is a chronic lung disease characterized by an ongoing, sustained inflammatory response that is associated with various degrees and combinations of chronic bronchitis, small airway fibrosis, and emphysematous lung tissue destruction. Apart from COPD, fibulin-5 protein levels were also reported to be increased in lung tissue of patients with idiopathic pulmonary fibrosis (IPF). [6] In IPF patients, the alveolar lung architecture is destroyed and replaced by fibrotic tissue, probably resulting from an aberrant ongoing repair response. Recently, fibulin-5 was also shown to be increased in hepatic fibrosis. Both fibulin-5 protein and gene expression levels were associated with the severity of fibrosis, with the highest expression in liver biopsies with severe hepatic fibrosis compared to those with mild fibrosis, [7] suggesting that fibulin-5 is intrinsic to this fibrosis.

To explain these observations and elaborate on the therapeutic potential of targeting fibulin-5 in COPD and/or fibrotic diseases, we first discuss the role of fibulin-5 in elastogenesis and (lung) repair as well as its interaction with other proteins involved in these processes.

2. Fibulin-5, elastogenesis and repair

Elastic fiber formation is a complex process in which tropoelastin, the soluble monomer, is deposited on microfibrils and cross-linked into a functional polymer. Next to fibrillin, these microfibrils consist of several other proteins such as microfibril-associated glycoproteins (MAGPs), latent transforming growth factor β (TGF-β) binding proteins (LTBPs), and fibulins. [8] Fibulin-5 co-localizes with elastic fibers in vitro and in vivo [3,5] and binds tropoelastin and fibrillin-1. [1] TGF-β induces fibulin-5 expression in fibroblasts, and in vitro studies have shown that elastic fiber formation is enhanced in fibulin-5-overexpressing cells. [5,9,10] Additionally, the interaction between fibulin-5 and LTBP2 and LTBP4 in elastic fiber assembly is important, and LTBP4 was suggested as a potential therapeutic target for regeneration of elastic fibers in vivo. [11,12] Hirai and colleagues demonstrated that the presence of fibulin-5 protein is necessary for elastic fiber assembly in vitro, and, by comparing skin samples from old and young mice, they identified a cleaved protein form of fibulin-5 that was more prominent in old mice. [13] In subsequent in vitro experiments, they showed that serine proteases can cleave fibulin-5, which results in this cleaved nonfunctional fibulin-5 protein.

These observations are intriguing, especially in light of our findings in COPD, where we also demonstrated cleaved fibulin-5 protein in lung tissue. COPD has been regarded as an aging lung disease with high protease activity, contributing to emphysematous lung tissue destruction. Therefore, we hypothesized that the increased fibulin-5 levels in COPD are the result of an attempt to repair the damaged lung. However, this repair response is ineffective in COPD due to increased proteolytic cleavage of fibulin-5 protein and/or disturbed expression of the other lung elastogenesis proteins. This hypothesis is in line with previous studies showing increased fibulin-5 gene expression after elastase-induced emphysema in mice [14] and increased elastin gene expression in severe COPD. [15]

As mentioned above, fibulin-5 expression is induced by TGF-β, the main growth factor involved in wound repair and fibrosis. In line with this, fibulin-5 was also shown to be involved in TGF-β-driven epithelial–mesenchymal transition, an important process for wound healing, tissue remodeling and repair, and fibrosis. [16] Apart from the hypothesis that fibulin-5 expression is high in COPD as an attempt to repair, and thus theoretically beneficial, it has also been proposed that the increased fibulin-5 expression in COPD is involved in small airway fibrosis, and thus detrimental. [17] This would also be in line with the increased fibulin-5 levels observed in patients with hepatic fibrosis and IPF.

3. Expert opinion: fibulin-5 as a therapeutic target

To aim at fibulin-5 as a therapeutic target in COPD, it is crucial to establish whether increased fibulin-5 expression in COPD is related to emphysema or small airway fibrosis, or both, and thus whether it has beneficial or detrimental effects. We first need to assess the exact location of the increased fibulin-5 expression in COPD lungs, i.e. alveolar tissue versus small airways, and elucidate the relation with emphysema severity and protease activity in the lung. Furthermore, the functional role of fibulin-5 in lung elastogenesis, repair, and fibrosis and its interaction with other elastogenesis proteins should be studied carefully.

If the fibulin-5 increase is related to emphysema and located in the alveolar tissue, but not functional due to increased proteolytic cleavage of fibulin-5, then potential therapeutic approaches could be to restore the levels of functional fibulin-5 in the emphysematous parts of the lung combined with inhibition of the protease activity. If more and/or other proteins involved in lung elastogenesis are deregulated in COPD, these proteins could also be regarded as potential targets to intervene and restore normal lung repair in COPD. We need to realize, however, that elastic fiber formation is a very complex process and that it may be challenging to reestablish this in vivo.

If increased fibulin-5 levels contribute to small airway fibrosis in COPD, ideally, the best therapeutic approach would be to directly target fibulin-5 (or one of its downstream targets) in the small airways. Given the challenge of targeting the small airways, it would be most straightforward to first target fibulin-5 in lung and/or liver fibrosis in general, and, if successful, try to target the small airways in COPD. It would also be of interest to assess whether fibulin-5 (or its cleaved form) can be measured in sputum or serum in order to apply it as biomarker in COPD, similar to what was proposed for hepatic fibrosis. [7]

An interesting example of a fibulin-5 target gene that may explain its role in fibrosis is transglutaminase 2 (TG2). TG2 is the most widely expressed TG and is involved in the cross-linking of ECM proteins such as elastin, collagen, and fibronectin. [18] TG2 expression was shown to increase >30-fold in fibulin-5-overexpressing 3T3 fibroblasts, suggesting that TG2 expression is regulated by fibulin-5. [19] TG2 has been associated with hepatic, renal, cardiac, and pulmonary fibrosis, and increased TG2 expression was demonstrated in lung tissue of IPF patients. Furthermore, TG2 knockout mice are protected against bleomycin-induced lung fibrosis. [18] Regarding the therapeutic potential of TG2, a recent study demonstrated that two small electrophilic compounds inhibit TG2 and its pro-fibrotic activities in human lung fibroblasts. [20] The in vivo therapeutic potential of these compounds in fibrotic lung disease is of great interest, also in relation to fibulin-5 expression.

In conclusion, fibulin-5 is an essential protein for elastic fiber formation, and we previously demonstrated increased levels in COPD lungs. Other groups found increased levels in IPF and hepatic fibrosis. Depending on the role of fibulin-5 in either small airway fibrosis or emphysema in COPD, different potential therapeutic strategies can be envisaged. When involved in small airway fibrosis, fibulin-5, or its target gene TG2, should be directly targeted in the airways. When increased proteolytic cleavage of fibulin-5 is underlying the lack of repair in the emphysematous lung, restoring functional fibulin-5 protein levels in combination with reducing protease activity can be a successful approach to halt further tissue destruction and stimulate lung repair.

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.