Pulmonary fibrosis is an irreversible condition where the tissue (interstitium) surrounding the small air sacs (alveoli) in the lungs become thickened and scarred, leading to impaired gas transfer, loss of lung function and in many cases, death. Pulmonary fibrosis may be caused by known insults, such as asbestos, silica, radiation exposure, pathogens such as SARS-CoV-2 (COVID) and some autoimmune diseases but can also develop in the absence of any known stimulus, such as in idiopathic pulmonary fibrosis (IPF), which is a type of idiopathic interstitial pneumonia. IPF has no known cause and patients have a very poor prognosis with limited treatment options and a life expectancy of fewer than five years post-diagnosis (Fernandez Perez et al., 2010; Ley & Collard, 2013; Nathan et al., 2011). The incidence of IPF is increasing and likely to be 20–30 per 100,000 in the USA and Europe and as high as 45 per 100,000 in Asia (Maher et al., 2021). The pathogenesis of IPF remains poorly understood, but the most accepted hypothesis is that recurrent or ongoing injury to alveolar epithelial cells leads to impaired re-epithelialisation, release of proinflammatory mediators and accumulation of immune and profibrotic cells in the lung that deposit large amounts of extracellular matrix (ECM), predominantly collagen, within the lung interstitium (Betensley, Sharif, & Karamichos, 2016). The existence of familial forms of IPF demonstrates that genetic influences are also important in regulating profibrotic events, which can modulate the immunological response to injury and collagen metabolism in the lung (Sgalla et al., 2018). IPF is currently treated with the antifibrotic drugs pirfenidone or nintedanib (Mackintosh et al., 2023), however clinical outcomes are variable, with reduced lung function decline and improved, although limited, survival benefit in some but not all patients (Finnerty, Ponnuswamy, Dutta, Abdelaziz, & Kamil, 2021).

Although early studies focussed on identifying cytokine and growth factor signalling pathways and their role in fibroblast accumulation, differentiation and collagen production, there is now a growing appreciation for a role of both the innate (always present) and adaptive (induced) immune response in the initiation and progression of pulmonary fibrosis. The discovery of a new repertoire of innate lymphocytes that can facilitate activation of T and B cells has uncovered new insights into disease pathogenesis in conditions such as IPF. There is also evidence of a breakdown in self-tolerance to lung autoantigens derived from damaged or dying epithelial cells in some patients, which may drive immune pathology (Moua, Maldonado, Decker, Daniels, & Ryu, 2014).

The innate immune response is composed of both physical barriers and a variety of different cell types that maintain the integrity of mucosal surfaces and tissue homeostasis within the lung, gut and skin. The innate immune response is directed by various cell types including cells of the myeloid lineage such as macrophages and dendritic cells and cells of the innate lymphoid lineage (e.g. ILC1, 2 and 3) (Vivier et al., 2018; Wynn & Vannella, 2016).

Fibroblasts, epithelial cells, endothelial cells and mesothelial cells also contribute to the innate defense process as these cells express pattern recognition receptors (PRR) that allow them to sense pathogens or signal to neighbouring cells in response to cell damage, and secrete cytokines that influence immune responses (Lafferty, Qureshi, & Schnare, 2010; Mutsaers, Pixley, Prele, & Hoyne, 2020). Mucosal epithelium lining the lung, gut and reproductive tract produce mucin and surfactant proteins providing a protective surface and barrier, preventing the access of microbial pathogens to the underlying submucosa (Johnston, Goldblatt, Evans, Tuvim, & Dickey, 2021). Altered epithelial barrier function has been associated with the induction and propagation of disease. Excessive mucus production and reduced mucociliary clearance contributes to transient respiratory infections and to the pathogenesis of numerous respiratory diseases (Roy et al., 2014). For example, a gain-of-function MUC5B promoter variant, expressed in the bronchoalveolar epithelium of IPF patients (Schwartz, 2018), is strongly associated with reduced mucociliary movement and the development of IPF (Allen et al., 2017; Hancock et al., 2018). Ongoing disease processes leads to further tissue destruction and if this is not resolved, the cytokine networks that regulate innate and adaptive immune cell populations drive a chronic inflammatory response that further disrupt tissue integrity leading to pathology.

Macrophages play a critical role in distinguishing foreign antigens to defend the host from potential pathogens that breach mucosal surfaces. They can elicit rapid immune responses through sensing of pathogens via PRRs, which enable the cell to link intracellular signalling responses to changes in cellular functions. That leads to the secretion of various effector cytokines that can influence the growth and differentiation of neighbouring cells and help coordinate responses by the adaptive immune response (Zigmond et al., 2012). Recent single cell RNAseq studies have identified profibrotic macrophage populations in the lungs of bleomycin (Blm)-treated mice and in patients with hypersensitivity pneumonitis (Wang et al., 2022; Wang et al., 2022). These cells, identified in patients with fibrotic hypersensitivity pneumonitis on the basis of high levels of phospholipase A2 group VII (PLA2G7), are associated with increased chemokine (CC-motif) ligand (CCL)4 levels and enrichment of a gene expression signature associated with the inflammatory response (Wang, Zhang, et al., 2022). Subsequent animal studies have demonstrated that PLA2G7high macrophages exert their pro-fibrotic effects through the induction of myofibroblast differentiation and that inhibition of PLA2G7 reduces Blm-induced fibrosis in vivo (Wang, Jiang, et al., 2022).

Type 1 and Type 2 immune responses reflect a distinct pattern of cytokine release in response to pathogen exposure or following tissue damage (Shinkai, Mohrs, & Locksley, 2002; Stetson et al., 2004) (Fig. 1). Type I responses are generally proinflammatory, marked by the release of cytokines including interferon gamma (IFNγ), tumour necrosis factor alpha (TNFα) and interleukin (IL)-1β, which induces the differentiation of macrophages into a proinflammatory M1 phenotype. M1 macrophages have an increased phagocytic activity and can process and present foreign antigens via major histocompatibility complex (MHC) molecules that are recognised by the antigen receptor on T cells within the adaptive immune system (Zigmond et al., 2012). TNFα and IL-1β are also known profibrotic cytokines that can drive fibrosis across multiple tissues including lung, liver and gut (Murray et al., 2014). The proinflammatory effects of TNFα and IL-1β can drive parenchymal damage in the lung and can induce epithelial-to-mesenchymal transition and myofibroblast differentiation that is dependent upon transforming growth factor beta (TGF-β) (Fan et al., 2001).

Type 2 cytokines, IL-4 and IL-13, which signal through IL-4 receptor alpha, direct the differentiation of macrophages into an M2 phenotype, which is crucial for tissue repair. M2 macrophages display a specific cellular phenotype; mannose receptor (CD206), chitinase-3–like protein-3 (also known as Ym1), resistin-like molecule-α (also known as FIZZ-1), major histocompatibility complex class II (MHCII) antigens and the enzyme arginase-1 (Van Dyken & Locksley, 2013; Wang et al., 2020; Ye et al., 2020). IL-4 and IL-13 levels are elevated in bronchial alveolar lavage (BAL) fluid in IPF patients compared to controls and these cytokines can direct the differentiation of M2 cells (Hancock, Armstrong, Gama, & Millar, 1998; Park et al., 2009). Both IL-4/IL-13 can promote cell proliferation and differentiation of fibroblasts, macrophages and myofibroblasts (Jakubzick et al., 2003; Rafii, Juarez, Albertson, & Chan, 2013). The differentiation of profibrotic fibroblasts is directed by the expression of the PU.1 transcription factor which is enhanced by TGF-β1 signalling and antagonised by TNFα signalling within the tissue microenvironment (Wohlfahrt et al., 2019). M2 cells, through secretion of IL-10, can also direct the differentiation of CD4+ Th cells into the T regulatory (Treg) cell lineage that inhibits inflammatory immune responses to promote tissue homeostasis and tissue repair (Murray & Wynn, 2011). Further investigation of human Tregs in blood, synovial tissue, lung and colon can express the thymic stromal lymphopoietin (TSLP) receptor, ST-2 (Lam et al., 2019). However, ST-2+ human Tregs appear to display an innate-like function in tissue repair that is independent of T cell receptor signalling as they rely instead on ST2-mediated signalling to induce amphiregulin to promote tissue repair which is similar to Treg cells in mice that mediate tissue repair and which rely on IL-33 signalling (Burzyn et al., 2013; Kuswanto et al., 2016).

Furthermore, tissue macrophages sense IL-4 and IL-13 in the presence of apoptotic cells to promote tissue repair (Bosurgi et al., 2017). IPF is considered to involve damage and subsequent apoptosis of epithelial cells which disrupt the mucosal epithelial barrier. Macrophages recognise apoptotic cells by sensing phosphatidylserine (Fadok et al., 1992; Savill, Henson, & Haslett, 1989) via signalling of the phosphatidylserine-dependent receptor tyrosine kinases Axl and Mer, which are expressed by both bone marrow-derived monocytes (Zagorska, Traves, Lew, Dransfield, & Lemke, 2014) and tissue-resident macrophages (Gautier et al., 2012; Zagorska et al., 2014).

Despite the detection of IL-4 and IL-13 in IPF lung tissue, several recent clinical trials that have targeted the IL-4/IL-13 pathway with either monospecific or bi-specific therapeutic monoclonal antibodies have failed to demonstrate any clinical benefit to IPF patients in reducing the decline in lung function or to modify disease progression in IPF (Raghu et al., 2018). Further, a recent clinical trial by Maher and colleagues confirmed that anti-IL-13 given in the presence or absence of the known antifibrotic drug pirfenidone, had no beneficial effect on patient outcomes (Maher et al., 2021). The lack of success observed by targeting IL-4/IL-13 in IPF may suggest that these cytokines are not the primary drivers of lung fibrosis in IPF. These type 2 cytokines may work in association with other humoral factors to drive tissue fibrosis and it highlights the necessity to understand the interplay of various cytokines, chemokines and growth factors in driving tissue fibrosis.

Fibroblast and immune cell crosstalk has emerged as a critical process in promoting an efficient wound healing response. Wound healing is regulated via overlapping immune responses associated with migration and proliferation of various cell types, the deposition of ECM and tissue remodelling (Hara & Tallquist, 2023). Inflammatory monocytes and tissue resident macrophages are key regulators of tissue fibrosis (Wynn & Ramalingam, 2012). Results from the ASCEND, CAPACITY and INSPIRE clinical trials demonstrated that IPF patients with an elevated monocyte count (≥0.95 × 109 cells/L) experienced IPF progression, defined by an absolute decline in predicted forced vital capacity (FVC), decline in 6-min walk distance or death (Kreuter et al., 2021). These findings were consistent with a recent meta-analysis showing that an elevated blood monocyte count was associated with increased risk of mortality and disease progression in ILD (Min, Grant-Orser, & Johannson, 2023). These findings suggest that blood monocyte counts may be a promising biomarker to identify IPF patients that are at risk of disease progression and mortality.

Tissue resident macrophages can be a source of chemokines that attract T cells and fibroblasts to establish a fibrotic niche (Gibbons et al., 2011). Blood-derived monocytes can respond to cytokines released from the tissue microenvironment, such as TGF-β1 and IL-4/IL-13, and differentiate into alternatively activated M2 macrophages which can produce TGF-β1 and IL-4 (Van Dyken & Locksley, 2013). TGF-β1 can direct the differentiation of fibroblasts into collagen producing myofibroblasts that lay down ECM associated with tissue scarring (Ueno et al., 2011). There has been much discussion around fibroblast heterogeneity and the role it plays in disease pathogenesis. Transcriptomic analysis has recently revealed an increasing number of fibroblast sub-phenotypes. The general consensus is that myofibroblasts are responsible for excessive ECM deposition in the lungs of IPF patients with recent lineage tracing results demonstrating that platelet-derived growth factor receptor-β+ cells are the predominate source of myofibroblasts in the mouse lung (Chandran et al., 2021). In addition to myofibroblasts, Liu et al., using sc-RNAseq, showed that other fibroblast subtypes, including lipofibroblasts and Ebf1+ fibroblasts, contribute to the excessive matrix protein secretion in lung fibrosis (Liu et al., 2023). While the specific effects of Th1 and Th2 cytokines on each of these specific sub-populations is unknown and require further investigation, cytokines released from damaged epithelial cells or inflammatory cells facilitate the crosstalk between macrophages and myofibroblasts (von Moltke, Ji, Liang, & Locksley, 2016).

ILCs play a critical role in tissue homeostasis in the gut and lung. There are three major ILC subsets that have been characterised; ILC1, 2 and 3 (Vivier et al., 2018), that are derived from the common lymphoid progenitor but differ from conventional lymphocytes in that they lack expression of antigen specific receptors and are activated in response to the release of alarmin-like signals (Moro et al., 2010; Neill et al., 2010). ILCs are usually tissue resident cells with ILC2 and ILC3 enriched at mucosal barrier surfaces in the gut, lung and skin (Chen, Hardman, Yadava, & Ogg, 2020). ILCs have emerged as key players in regulating immune responses to tissue damage. ILCs are responsive to cytokines released in response to pathogen detection but can also respond to alarmins released in response to tissue damage. They can interact with a range of immune and non-immune cell lineages within barrier tissues and can migrate to tissues where they establish a niche. IL-25, a type 2 cytokine, can direct the differentiation of ILC2 cells (Moro et al., 2010; Neill et al., 2010; Saenz et al., 2010). In turn, ILC2 cells can direct CD4+ Th2 cell differentiation via secretion of IL-4 and IL-13 and interactions with OX40/OX40L (also called CD134) signalling and this crosstalk sustains their mutual survival, proliferation and cytokine production within tissue niches (Drake, Iijima, & Kita, 2014; Halim et al., 2014; Mirchandani et al., 2014). In addition, IL-13 produced by ILC2 cells can induce dendritic cells to produce the Th2 chemokine CCL17 which has been implicated with tissue fibrosis (Halim et al., 2016).

MHCII+ ILC2 and ILC3 subsets have been identified in the lung, gut and skin (Mirchandani et al., 2014; Oliphant et al., 2014). The MHCII+ ILC3 subset can process and present antigen via MHCII and can display suppressive or immune activating responses which appear to be tissue dependent. Further studies are required to understand the dichotomy of responses by MHCII+ ILCs (von Burg et al., 2014). Skin-derived ILC2 cells can express CD1d, and this is thought to endow these cells with the ability to present lipid antigens (Hardman et al., 2017). Three recent studies have identified the retinoic acid-related organ receptor gamma t (RORγt)+ antigen presenting cells that play a critical role in establishing mucosal tolerance to commensal gut bacteria (Akagbosu et al., 2022; Kedmi et al., 2022; Lyu et al., 2022). Kedmi et al. identified a novel antigen presenting cell population in the gut that expresses RORγt+ that were distinct from the classical dendritic cells of the myeloid lineage (Kedmi et al., 2022). These RORγt+ ILC3 cells localise in regions occupied by Treg cells and inhibit expansion of Th17 cells. Moreover, the ILC3s appear to shift the balance in the gut mucosa to favour of antigen specific RORγt+ Tregs over Th17 cells and this is essential to establish immune tolerance to the gut microbiota (Lyu et al., 2022). Further work is required to understand if the ILC3 cells have equivalent roles to induce Tregs outside of gut tissue.

The function of ILC3 cells extends beyond just the regulation of mucosal tolerance. There is evidence that ILC3s also play important roles in a number of autoimmune diseases including systemic lupus erythematosus, systemic sclerosis (SSc), inflammatory bowel disease, multiple sclerosis, psoriatic arthritis and ankylosing spondylitis that involves IL-17 and IL-22 signalling (Ardain, Porterfield, Kloverpris, & Leslie, 2019). ILC3s have also been implicated in a range of lung diseases including asthma, chronic obstructive pulmonary disease and pulmonary fibrosis (Ardain et al., 2019). Epithelial cell-derived cytokines IL-33, IL-25 and TSLP, produced in response to epithelial cell damage or detection of pathogens by PRRs, are able to induce IL-4/IL-13 production in macrophages (Roan, Obata-Ninomiya, & Ziegler, 2019). IL-33 and TSLP can also activate ILC2 cells which rapidly produce IL-5 and IL-13 in response to these cytokines (Duerr et al., 2016; Han et al., 2012; Stier et al., 2016). IL-25, IL-33 and TSLP and their corresponding receptors; IL-17BR, ST2L and TSLPR are all up-regulated in IPF patients and in mouse models of Blm-induced lung fibrosis and it has been suggested that the IL-25/IL-33/TSLP axis may be the master regulator of abnormal epithelial–mesenchymal interaction (Xu, Dai, & Zhang, 2022). Clearly more research needs to be done on the role of these molecules in the pathogenesis of lung fibrosis, but they are promising targets for the treatment of fibrotic lung disease, in particular IPF.

The original discovery of the dichotomy of different Th cell subsets referred to as Th1 and Th2 cells by Mosman and Coffmann, represented an important paradigm that underpinned cellular immunology since the late 1980s (Mosmann, Cherwinski, Bond, Giedlin, & Coffman, 1986). The discovery of Th1 and Th2 cells was followed by the identification of CD4+ Tregs that express the transcription factor Foxp3 and suppress immune responses to self-antigens and mediate mucosal tolerance to microbial antigens (Hori, Nomura, & Sakaguchi, 2003; Khattri, Cox, Yasayko, & Ramsdell, 2017). The identification of CD4+ Th17 cells was another important milestone as these cells produced IL-17 in response to activation in the presence of TGF-β1 and IL-6 (O'Garra, Stockinger, & Veldhoen, 2008; Veldhoen, Hocking, Atkins, Locksley, & Stockinger, 2006). This led to a better understanding of the cytokine transcriptional works that were responsible for the differentiation of Th cell subsets due to the lineage specific expression of unique transcription factors that could direct the differentiation and plasticity of CD4+ Th cell differentiation in vivo (Ivanov et al., 2006; Lighvani et al., 2001; Mullen et al., 2001; Ricard et al., 2019; Szabo et al., 2000; Zheng & Flavell, 1997) (Fig. 1). TGF-β1 was originally identified as an inhibitory cytokine that blocked T cell proliferation (Kulkarni et al., 1993; Shull et al., 1992). However, it was later revealed that TGF-β1 also has a central role in the differentiation of CD4+ Th cells into either the Treg or Th17 cell lineage. Studies by Zhou and colleagues revealed that TGF-β1 can synergise with IL-6 and IL-21 to induce expression of RORγt expression which directs the differentiation of CD4+ cells to the Th17 lineage (Veldhoen et al., 2006; Zhou et al., 2008). In contrast, high concentrations of TGF-β1 alone directs the differentiation of CD4+ cells into Treg lineage by inducing expression of Foxp3 that antagonises RORγt expression to inhibit differentiation of CD4+Th17 cells (Zhou et al., 2008).

CD4+ Th17 cells are associated with tissue inflammation through the release of the proinflammatory cytokine IL-17 A, but these cells are also an important regulator of tissue fibrosis. Using mouse models of tissue fibrosis, it was demonstrated that IL-1β and IL-23 are important upstream modulators of Th17-induced profibrotic responses together with TGF-β1 (Gasse et al., 2011; Wilson et al., 2010). IL-17 appears to promote tissue fibrosis indirectly by inducing tissue damage. Celada and colleagues identified a subset of CD4+ programmed cell death protein 1 (PD-1)+ Th cells in patients with IPF and sarcoidosis that actively produced IL-17 A and TGF-β (Celada et al., 2018). Co-culture of CD4+ PD-1+ cells with human fibroblasts in vitro increased collagen synthesis which was abrogated by anti-PD-1 treatment of T cells. An equivalent cell population of CD4+ PD-1+ cells were amplified in response to Blm-induced pulmonary fibrosis in mice (Celada et al., 2018). Blockade of PD-1 signalling subsequently reduced Blm-induced tissue fibrosis. Collectively these results highlight that Th17 cells exhibit a pro-fibrotic response in a range of tissue settings and therefore targeting PD-1+ CD4+ T cells could be a feasible immune therapy to block tissue fibrosis.

IL-13 and TGF-β1 are both pro-fibrotic cytokines that are produced by Th2 cells and Treg cells respectively, and both cytokines have been implicated in lung fibrosis associated with IPF, chronic asthma and a range of other fibrotic diseases (Wynn & Ramalingam, 2012). TGF-β1 can induce expression of procollagen 1 and III and promote collagen deposition by myofibroblasts (Verrecchia, Chu, & Mauviel, 2001). IL-13 activity is regulated by IL13Rα signalling, and secretion of the IL-13Rα2 decoy protein expressed on myofibroblasts can bind to IL-13 in the extracellular space to neutralise its activity (Chiaramonte et al., 2003; Ramalingam et al., 2008). Genetic depletion of IL-13Rα2 in mice exacerbates IL-13 driven tissue fibrosis, but they are resistant to IL-1β and IL-17-induced inflammation (Mentink-Kane et al., 2011; Wilson et al., 2011). These studies revealed that regulation of IL-13 signalling in vivo via the IL-13 decoy protein helps to regulates the balance between Th17-induced inflammation and Th2 driven tissue fibrosis.

Th1 cells secrete IFNγ which antagonises the production of type 2 cytokines such as IL-4 and IL-13 by Th2 cells (Mosmann et al., 1986). By reducing the release of T cell-derived IL-4, IFNγ can inhibit the differentiation of M2 macrophages which prevents SMAD3 interaction with TGF-β receptor thus abrogating TGF-β signalling in macrophages (Ulloa, Doody, & Massague, 1999). IFNγ can also decrease TGF-β signalling in myofibroblasts via increased expression of SMAD7, which in turn can inhibit the expression of collagen synthesis and fibroblast proliferation (Gurujeyalakshmi & Giri, 1995). IL-12, which is also a potent inducer of IFNγ production by CD4+ Th1 cells, can also protect mice from pulmonary fibrosis (Keane, Belperio, Burdick, & Strieter, 2001; Wynn et al., 1995). Despite the promising pre-clinical findings in animal models of tissue fibrosis, the same results were never achieved in clinical trials, with IFNγ being unable to reduce tissue fibrosis in humans (King Jr., Albera, Bradford, Costabel, Hormel, Lancaster, Noble, Sahn, Szwarcberg, Thomeer, Valeyre, du Bois, and Group IS, 2009).

Follicular B helper T cells (Tfh) are located within B cell follicles in secondary lymphoid tissues and play a crucial role in providing help to antigen specific B cells within the germinal centre (Vinuesa et al., 2005; Vinuesa, Linterman, Yu, & MacLennan, 2016). Tfh cells are characterised by the expression of PD-1, ICOS, Bcl6 and the CXC chemokine receptor type 5 (CXCR5) that binds to its ligand, chemokine (C-X-C motif) ligand (CXCL)13, which directs the migration of Th cells into B cell follicles where they eventually take up residence in the germinal centre (Yu et al., 2007). Elevated Tfh cell numbers can lead to autoimmune diseases such as systemic lupus erythematosus with autoantibody production (Li et al., 2012; Linterman et al., 2009). Asai and colleagues identified that Tfh cell numbers were elevated in the peripheral blood in IPF patients compared with healthy controls (Asai et al., 2019) and Tfh cells were also upregulated in patients with SSc (Taylor et al., 2018).

The different Th cell subsets help guide B cells to produce different classes of antigen-specific antibodies which influence the effector response to an antigenic stimulus. During an immune response, B cells can undergo clonal expansion and affinity maturation of their antibody during the germinal centre response (Young & Brink, 2021). B cells that bind antigen are selected by Tfh cells in the follicle to enter the germinal centre to undergo proliferation and mutation to increase the affinity of their receptors (Krautler et al., 2017). Two distinct thymic-derived lineages of CD4+Tregs were discovered in 2011; one was found to co-express CD4+ Foxp 3 and a T cell-associated transcription factor (Tbet) and could suppress Th1 cells (Koch et al., 2009), and the second co-expressed CD4, Foxp3 and Bcl6 and localised to lymphoid follicles where they inhibited B cells (Chung et al., 2011; Linterman et al., 2011). The Tfr cells modulate germinal centre responses in the follicle by secreting neuritin that acts to inhibit Tfh cells, which in turn inhibits plasma B cell differentiation (Botta et al., 2017; Sage et al., 2016) and autoantibody development against tissue-specific antigens (Gonzalez-Figueroa et al., 2021).

In IPF, T cells, B cells, macrophages and dendritic cells accumulate within specific foci in the lung tissue adjacent to areas of active tissue fibrosis (Hoyne, Elliott, Mutsaers, & Prele, 2017; Prele et al., 2022). This raises the possibility of immune cell cross talk between antigen specific lymphocytes and fibroblasts, but exactly how this operates is poorly understood. Ali et al., showed that B cells could induce fibroblast migration and activation (Ali et al., 2021). However, the role of B cells in lung fibrosis remains controversial.

The aggregates of mature B cells within fibrotic foci are a feature of IPF lung (Hoyne et al., 2017; Prele et al., 2022; Todd et al., 2013). This may reflect trafficking of B cells rather than local proliferation, as Xue et al. reported that B cells located within fibrotic foci are negative for the proliferation marker Ki67 (Xue et al., 2013) but CXCL13, a mediator of B cell chemotaxis, is upregulated in IPF tissue and serum (Vuga et al., 2014) and IPF serum has high levels of circulating B cell-activating factor (BAFF) (O'Dwyer et al., 2017; Vuga et al., 2014). Similarly, DePianto and colleagues demonstrated that high serum levels of CXCL13 were positively correlated with dyspnea score and negatively correlated with diffusing capacity for carbon monoxide (DePianto et al., 2015). Vuga et al., also showed that CXCL13 is a prognostic marker for IPF, suggesting an important role for B cells in progression and hence severity of the disease (Vuga et al., 2014). The reason for B cell migration and accumulation in IPF and their role in fibrosis is unclear. However, their presence has been associated with IPF progression (Marchal-Somme et al., 2006; Todd et al., 2013).

Approximately a third of IPF patients overexpress the human leukocyte antigen DRB*1501 allele, encoding MHCII, which is the protein responsible for antigen presentation (Xue et al., 2011). This suggests that these IPF patients may have an altered capacity for antigen presentation to CD4+ Th cells. The increase in human leukocyte antigen DRB*1501 has also been associated with impaired gas exchange in the lung (Xue et al., 2011) suggesting a role for the interaction between immune cells and antigens in IPF, however, whether these are microbial, environmental, or self-antigens, is undefined.

There appears to be a higher level of plasmablast differentiation in IPF patients compared to healthy controls (Hamada et al., 2015; Heukels et al., 2019; Xue et al., 2013). Recent proteome analysis of lung material obtained from end-stage ILD patients demonstrated a population of marginal zone plasma cells ([MZ]B1+CD38+CD138+CD27+CD20−) prevalent in these samples (Schiller et al., 2017), supporting a role for autoantibody-producing mature B cells and plasma cells in the pathogenesis of lung fibrosis. There have been several studies demonstrating the presence of autoantibodies in IPF but they have been predominantly observational.

Serological autoantibody status has been recommended as a clinical tool to reclassify patients to myositis-associated (MA)-ILD, connective tissue disease-ILD or interstitial pneumonia with autoimmune features, depending on the autoantibody profile (Shao et al., 2021; Stevenson et al., 2019). The advantage of this is that a combination of a serological test as well as high-resolution computed tomography can be used to make a more informed decision regarding appropriate therapies, as autoantibody-positive IPF patients may benefit from immunomodulatory therapies, such as administration of glucocorticoids (Ghang et al., 2019), although this area has not been further explored. The presence of autoantibodies in IPF patients have been identified by several groups that include antigens expressed by epithelial cells (e.g. periplakin [PPL], vimentin, HSP70) and systemic autoantigens such as DNA, RNA or nuclear proteins and rheumatoid factor (Hoyne et al., 2017; Koether et al., 2023). In autoimmune disease, the levels of autoantibodies correlate with disease progression (Wallace et al., 1994) and high levels of autoantibodies have been associated with acute exacerbations in IPF patients, characterised by a rapid deterioration in lung function that is medically untreatable and often results in death within days (Donahoe et al., 2015). Boustini and colleagues recently demonstrated that there is an increase in immunoglobulin (Ig)G1 and total IgA autoantibodies within the BAL fluid of IPF patients, indicating increased B cell activity (Boustani et al., 2022). Interestingly, they identified that IPF patients had increased BAL titres of autoantibodies against collagen V and vitronectin, but there was no correlation between the levels of specific autoantibodies and IgG1 or IgA autoantibody levels.

Koether and colleagues recently showed that IPF patients with autoantibodies to the intermediate filament protein PPL had a significantly worse outcome in terms of progression free survival (Koether et al., 2023). Using the Blm mouse model of pulmonary fibrosis, they demonstrated that mice treated with Blm and a cocktail of anti-PPL antibodies (to mimic PPL autoantibodies in IPF patients), showed increased fibronectin and collagen mRNA expression above Blm and control immunoglobulin treated mice at day 14. However, the level of collagen production was not significantly different between groups, suggesting the need for a more sustained antibody-induced fibrosis stimuli before differences could be seen at the protein level. Mice treated with anti-PPL antibodies on their own without Blm did not induce lung pathology but again, 14 days may not be long enough to demonstrate significant lung changes.

In 2014, a phase II clinical trial investigated the use of rituximab (Rtx) to treat patients with severe acute exacerbations of IPF (AE-IPF) (Donahoe et al., 2015). Rtx is an anti-CD20 antibody (CD20 is a B cell antigen) traditionally used to treat certain autoimmune diseases such as rheumatoid arthritis (Edwards et al., 2004) and B cell cancers like non-Hodgkin's lymphoma (Hauptrock & Hess, 2008). Nine patients (82%) showed improved pulmonary gas exchange compared to historical controls suggesting a novel anti-B cell therapy approach for AE-IPF (Donahoe et al., 2015).

Kulkarni and colleagues performed a similar study where a small cohort of AE-IPF patients were treated with Rtx in combination with plasma exchange and intravenous Ig (Kulkarni et al., 2021). Sixty three percent of patients showed a decrease in supplemental oxygen requirements and the one-year survival rate was greatest in the subgroup of patients that had an immediate beneficial response to the treatment regimen. Interestingly, when assessing autoantibody titers, the authors demonstrated that clear clinical improvements were associated with high anti-epithelial autoantibody levels. This indicates that Rtx, plasma exchange and intravenous Ig therapy may only be appropriate for a sub-group of AE-IPF patients. Whether genetic or other factors also contributed to improved outcome in this sub-group is unclear.

To date, Rtx has only been trialed in patients with AE-IPF. Given that the mechanisms regulating AE-IPF are unlikely to be the same as IPF and concerns that Rtx treatment itself may induce lung fibrosis (Tej, Undrajavarapu, Kadian, Khurana, & Goyal, 2022), no clinical trials have examined the effect of Rtx on stable IPF patients.

Lung fibrosis or ILD is a common feature of autoimmune-related connective tissue disease, where high levels of autoantibodies are a key feature. In conditions including SSc, rheumatoid arthritis, Sjögren's syndrome and idiopathic inflammatory myopathies, high levels of circulating autoantibodies are associated with ILD (Kinder et al., 2007; Vij, Noth, & Strek, 2011). Rtx has been demonstrated to improve lung function in these diseases and improve high-resolution computed tomography score in Sjögren's syndrome and idiopathic inflammatory myopathies (Chen et al., 2016; Fasano, Gordon, Hajji, Loyo, & Isenberg, 2017).

Several B cell blocking studies have shown a role for B cells in mouse models of fibrosis. Genetic depletion of B cells in a CD19 knockout mouse (CD19−/−) protected from Blm-induced dermal fibrosis and lung fibrosis (Saito et al., 2002). Furthermore, blocking T cell-mediated B cell activation with an anti-CD40L antibody reduced hapten immune pulmonary interstitial fibrosis in these mice (Zhang-Hoover, Sutton, & Stein-Streilein, 2001). Further studies showed that treatment with an anti-CD20 or anti-BAFF antibody reduced Blm-induced lung fibrosis (Hasegawa et al., 2006; Matsushita et al., 2018). Our laboratory and others have previously shown the importance of STAT3 in the pathogenesis of IPF (Prele, Yao, O'Donoghue, Mutsaers, & Knight, 2012). We subsequently demonstrated that transgenic mice with increased expression of STAT3 and deficient in both total mature T cells and B cells (RAG1−/−) or mature B cells (μMT−/−) are protected from Blm-induced lung fibrosis (O'Donoghue et al., 2012) although we were unable to demonstrate any reduction in Blm-induced fibrosis following anti-CD20 treatment (Prele et al., 2022). This is likely because anti-CD20 treatment doesn't deplete plasmablasts or plasma cells as they don't express CD20 on their surface. When we depleted plasmablasts and plasma cells from the blood and lungs of Blm treated mice using the proteasome inhibitor bortezomib, we showed a significant inhibition in lung fibrosis (Prele et al., 2022). This observation, although only in the Blm mouse model, also questions the use of Rtx, which does not deplete long-lived antibody-producing plasmablasts and plasma cells, for IPF treatment.

There are many different immunomodulatory treatments for non-IPF ILDs with inflammatory components, such as alveolitis that progresses upon antigen presenting cell-induced CD4+ T cell activation, release of proinflammatory cytokines and collections of immune cells in the alveoli and interstitium (van den Bosch, Luppi, Ferrara, & Mura, 2022). This leads to chronic inflammation, granuloma formation, fibroblast recruitment, ECM deposition and fibrosis. Interfering with the acute inflammatory component of the disease process can halt disease progression. Therefore, it is crucial that a correct diagnosis is made so that patients receive appropriate treatments. Immunosuppression is still the main treatment for many of the ILDs including corticosteroids, mycophenolate mofetil, azathioprine, methotrexate, cyclophosphamide and Rtx. Corticosteroids are usually first line therapy in ILDs as they are both anti-inflammatory and immunosuppressive, blocking the action of inflammatory mediators and inhibiting leukocyte movement (Liu et al., 2013). However, they are associated with significant side effects and are usually avoided in the long term. Mycophenolate mofetil is a steroid-sparing immunosuppressant used to treat ILDs. Mycophenolate mofetil inhibits inosine monophosphate dehydrogenase which has an anti-proliferative/cytostatic effect on lymphocytes (Allison, 2005). Azathioprine acts as an immunosuppressant by inhibiting purine synthesis to block T and B cell proliferation (Maltzman & Koretzky, 2003). Methotrexate is anti-inflammatory and immunosuppressive and is used to treat non-IPF ILDs with an inflammatory component. It acts as an anti-folate to interfere with purine and pyrimidine synthesis and prevent lymphocyte proliferation. It has also been shown to inhibit T cell activation (Wessels, Huizinga, & Guchelaar, 2008). Cyclophosphamide is a potent alkylating immunosuppressant used to treat cancers, autoimmune diseases and prevent graft versus host disease in transplant patients. However, it is highly toxic with significant side effects (Teles, Medeiros-Souza, Lima, Araujo, & Lima, 2017). As previously discussed, Rtx is a chimeric monoclonal antibody that targets CD20 expressed on normal and malignant B cells but spares plasma cells and hematopoietic stem cells as they do not express CD20 on their surface. Rtx is traditionally used to treat certain autoimmune diseases and cancers such as non-Hodgkin's lymphoma but more recently has also been used to treat fibrosing ILDs and AE-IPF (Edwards et al., 2004; Hauptrock & Hess, 2008; Kulkarni et al., 2021). The use of these agents in treating inflammatory ILDs has recently been reviewed by van den Bosch and colleagues (van den Bosch et al., 2022) and won't be further discussed here.

Current therapies for IPF; pirfenidone and nintedanib, are considered to function by blocking TGF-β activity and growth factor signalling respectively (Richeldi et al., 2011; Ruwanpura, Thomas, & Bardin, 2020). However, their exact mode of action, particularly for pirfenidone, remains unclear, and some studies have suggested they may have immunosuppressive effects. Pirfenidone reduces CD4+ T cell proliferation and pro-inflammatory cytokine production (Visner et al., 2009) and nintedanib, whilst not altering the total number of CD4+ T cells, has been shown to reduce T cell activation, proinflammatory cytokine release and the number of Th1- and Th2-like cells in vitro. Interestingly, nintedanib treatment increased the number of Th17-like cells but did not increase IL-17 A production (Ubieta, Thomas, & Wollin, 2021). However, the potential role of immunotherapies in IPF is controversial and novel approaches will be discussed in more detail.

The use of immunomodulatory medications for the treatment of IPF has received little support until recently, based mainly on historical data and more recently, concerns identified in some patients receiving immune checkpoint inhibitor (ICI) therapies for cancer. Medications such as prednisone and azathioprine were once a commonly accepted therapy for IPF (ATS, 2000) as many patients demonstrated a significant benefit from their use. However, subsequent studies showed that when used in patients with the new criteria that is used to define IPF (Raghu, Remy-Jardin, Myers, Richeldi, Ryerson, Lederer, Behr, Cottin, Danoff, Morell, Flaherty, Wells, Martinez, Azuma, Bice, Bouros, Brown, Collard, Duggal, Galvin, Inoue, Jenkins, Johkoh, Kazerooni, Kitaichi, Knight, Mansour, Nicholson, SNJ, Buendia-Roldan, Selman, Travis, Walsh, Wilson, and American Thoracic Society ERSJRS and Latin American Thoracic S, 2018), these drugs caused significant harm, suggesting that previous patients that benefitted from these therapies may not have had what we now define as IPF. These findings also fuelled a debate over the role of inflammation and the immune system in the pathogenesis of IPF and until recently it was generally accepted that inflammation and immune dysregulation did not have a significantly impact on disease development. Furthermore, the use of ICIs for cancer therapy has shown that a proportion (up to 10% depending on the disease and type of ICI used) of patients develop pneumonitis (Banavasi et al., 2023; Su et al., 2019). In most cases these patients can be treated with steroids and recover but ICI-mediated ILD has resulted in death of a small proportion of patients (Puzanov, Diab, Abdallah, Bingham III, Brogdon, Dadu, Hamad, Kim, Lacouture, NR, Lenihan, Onofrei, Shannon, Sharma, Silk, Skondra, Suarez-Almazor, Wang, Wiley, Kaufman, and Ernstoff MS and Society for Immunotherapy of Cancer Toxicity Management Working G, 2017). However, the complication of pneumonitis from an ICI is more likely to be fatal in a patient with IPF. Which patients will develop ICI-mediated complications such as pneumonitis is unclear and is the focus of ongoing biomarker studies (Chennamadhavuni, Abushahin, Jin, Presley, & Manne, 2022; Guberina, Wirsdorfer, Stuschke, & Jendrossek, 2023).

Despite these concerns, recent studies have once again highlighted a role for inflammation and the immune system in the pathogenesis of IPF, and logically, a role for immunoregulatory therapies. Importantly, with the advent of in-depth molecular diagnostics, it is becoming evident that the pathogenesis of IPF is unlikely to follow the same disease trajectory for all patients. Predisposing factors including host genetics, environmental pollutants, infection, aberrant repair, and dysregulated immune responses may all play important yet varied roles. Therefore each factor may require a different diagnostic and/or therapeutic approach. While broad immunosuppressive approaches are largely viewed to have failed in IPF, this may be because they target a multitude of immune mechanisms and cells and therefore targeting more specific pathways and cell types in appropriate patients may be a more suitable approach. The challenge is to identify each subgroup of patients so that the best treatment can be given to stop disease progression and ideally reverse the disease process.

There are currently a number of clinical trials in IPF using anti-inflammatory approaches. Treprostinil is a stable prostacyclin analogue used for the treatment of pulmonary arterial hypertension. It functions by promoting vasodilation of pulmonary and systemic arteries vascular beds and inhibiting platelet aggregation (El-Kersh & Jalil, 2023). Treprostinil has also been shown experimentally in vitro and in animal models to have antifibrotic properties by inhibiting profibrotic fibroblast activity and the synthesis and deposition of collagen and fibronectin (Kolb et al., 2022). It was recently demonstrated in the phase 3 INCREASE trial that inhaled treprostinil improved FVC in patients with ILD and associated pulmonary hypertension, in particular those patients with IPF (Nathan et al., 2021). Several trials are currently underway using inhaled treprostinil in patients with IPF with or without associated pulmonary hypertension (NCT04708782, NCT05255991, NCT04905693).

The PRECISION trial (NCT04300920) examines the efficacy of treating patients with the antioxidant N-acetylcysteine (NAC) who have the toll-interacting protein (TOLLIP) rs3750920 TT genotype. TOLLIP is an inhibitory adaptor protein that acts downstream of the toll-like receptors; key mediators of the innate and adaptive immune response (Li, Goobie, Gregory, Kass, & Zhang, 2021). Single-nucleotide polymorphisms within TOLLIP have been linked to alterations in the lung immune response and are associated with IPF susceptibility and survival (Fingerlin et al., 2013). The TOLLIP rs3750920 TT genotype results in increased expression of TOLLIP in monocytes, which may influence toll-like receptor signalling and host response to immunomodulatory therapies (Shah et al., 2012). The IPFnet PANTHER trial previously examined the effect of NAC alone or combined with prednisone and azathioprine versus placebo on lung function in IPF patients (Idiopathic Pulmonary Fibrosis Clinical Research N, Raghu, Anstrom, King Jr., Lasky, and Martinez, 2012). The combined therapy increased mortality so was terminated, but there was no difference seen in NAC alone compared with placebo. Despite this, post hoc analysis demonstrated that a subpopulation of IPF patients with the TOLLIP rs3750920 TT genotype (~ 25% of the cohort) benefited from NAC treatment and has led to the design of the PRECISION trial (Podolanczuk, Kim, Cooper, Lasky, Murray, Oldham, Raghu, Flaherty, Spino, Noth, Martinez, and Team PS, 2022) .

Setanaxib (GKT137831), an inhibitor of nicotinamide adenine dinucleotide phosphate oxidase (NOX) isoforms, has been used in many studies to examine the effect of blocking NOX on cell function (Elbatreek, Mucke, & Schmidt, 2021). In clinical trial NCT03865927, ambulatory patients with IPF are being treated with Setanaxib to determine if blocking NOX1 and NOX4, known to be important in generating reactive oxygen species and thought to be involved in IPF pathogenesis (Hecker et al., 2009), protect cells and tissues in the lungs against fibrosis. The study will assess changes in the oxidative stress biomarker o,o’-dityrosine, as well as changes in patient's FVC and collagen fragment levels in plasma.

There are a growing number of monoclonal antibody treatments being trialed, targeting specific molecules thought to be important in IPF pathogenesis. While some of these treatments target molecules that specifically affect immune function; such as tralokinumab, a human IgG4 antibody that binds and neutralises IL-13 (Parker et al., 2018), others inhibit the function of molecules important in cell proliferation, differentiation and collagen production. Connective tissue growth factor (CTGF), also known as cellular communication network 2, is a TGF-β-target gene and a member of the cellular communication network family of secreted proteins that play a crucial role in many different biological processes, particularly in the regulation of connective tissue formation and tissue repair (Holbourn, Acharya, & Perbal, 2008). CTGF is produced by various cell types and interacts with other proteins, growth factors and ECM proteins to regulate cell adhesion, migration, proliferation and differentiation, influencing tissue remodelling. CTGF is important in fibrosis by promoting the differentiation of fibroblasts into myofibroblasts and production of ECM components such as collagen (Effendi & Nagano, 2022). Previous studies demonstrated high levels of CTGF in the lungs and BAL of IPF patients and upregulation in the lungs of mice following Blm treatment (Allen, Knight, Bloor, & Spiteri, 1999; Pan et al., 2001). Fibrosis was significantly attenuated in Blm treated mice following treatment with an anti-CTGF antibody, highlighting the role of CTGF in the pathogenesis of lung fibrosis. Subsequent human trials demonstrated that Pamrevlumab, a humanised monoclonal antibody that targets CTGF, was safe in patients, reducing fibrosis and improving FVC over time (Raghu et al., 2016), which was confirmed in the subsequent phase 2 PRAISE trial (Richeldi et al., 2020). These trials have led to the phase 3 Zephyrus I (NCT03955146) and Zephyrus II (NCT04419558) trials, examining the safety and efficacy of intravenous infusions of pamrevlumab in IPF patients over 48 weeks. In the Zephyrus II trial, there is also an optional open-label extension phase. Although not classically immunotherapy, use of neutralising antibodies to block mediators such as growth factors is likely to be an important approach to treat diseases such as IPF.

ICIs, particularly those targeting the PD-1/programmed death ligand-1 (PD-L1) axis, have revolutionised cancer therapy and recent studies have suggested ICIs may be a novel approach for IPF. An increasing number of studies have demonstrated increased expression of PD-1 and/or PD-L1 on different cell types in IPF lung (Ahmadvand et al., 2022; Celada et al., 2018; Jiang et al., 2022) and inhibition of these molecules using specific PD-1 and PD-L1 monoclonal antibodies have demonstrated reduced lung fibrosis in animal models of fibrosis (Celada et al., 2018; Geng et al., 2019; Ni et al., 2018). Several studies have suggested different mechanisms for PD-1/PD-L1-induced lung fibrosis. Celada and colleagues identified a population of high PD-1 expressing CD4+ T cells and showed that these were predominantly Th17 cells (Celada et al., 2018); important in host defense and shown to promote fibrosis in the lung (Simonian et al., 2009). Co-culture of PD-1+ Th17 cells with human lung fibroblasts induced collagen production through STAT3 and TGF-β-mediated pathways, which was reduced by blocking PD-1 in T cells (Celada et al., 2018). Furthermore, Wang et al., demonstrated that PD-1 inhibited the differentiation of CD4+ T cells into Treg cells. Co-culture of CD4+ T cells and myofibroblasts (lung fibroblasts treated with TGF-β), induced collagen-1 production and inhibited the proliferation of CD4+ T cells. Blocking PD-1 reversed the inhibition of Treg cell differentiation and decreased collagen-1 production (Wang, Bai, Ma, & Li, 2021).

Geng and colleagues demonstrated increased PD-L1 expression on invasive lung fibroblasts and using a humanised mouse model, showed that PD-L1-induced fibroblast invasion and mediated fibrosis through p53 and focal adhesion kinase (FAK) pathways (Geng et al., 2019). Guo et al., demonstrated that PD-L1 was also required for fibroblast to myofibroblast differentiation and that this occurred through both Smad3-dependent and independent pathways to promote TGF-β-mediated fibrosis (Guo et al., 2022). Xia and colleagues previously identified a population of cells they termed pathologic mesenchymal progenitor cells (MPCs) that they suggested are the cells of origin for IPF fibroblasts (Xia et al., 2014). In a recent study they showed that IL-8 promoted both senescence and upregulation of PD-L1 in MPCs which were intermingled with natural killer cells in fibroblastic foci, suggesting that senescent MPCs elude immune cell surveillance. Disruption of PD-1/PD-L1 interaction promoted natural killer cell killing of IPF MPCs and arrested IPF MPC-mediated experimental lung fibrosis (Yang et al., 2023).

More research is required to elucidate the role(s) of PD-1 and PD-L1 in IPF as well as the potential roles of other checkpoint molecules in IPF pathogenesis. However, preclinical models have suggested a therapeutic benefit to using ICIs, although as previously mentioned, the concern that using ICIs can induce pneumonitis and other inflammatory/fibrotic complications including autoreactive humoral responses in some patients (Banavasi et al., 2023; Chen et al., 2022; Michot et al., 2016; Su et al., 2019) potentially leading to death, particularly in IPF patients, clearly needs thorough investigation. Despite this, Atezolizumab, a humanised monoclonal anti-PD-L1 antibody, used to treat several cancers including non-small cell lung cancer and metastatic breast cancer, is currently in a Phase 1 safety and efficacy trial for IPF. This is a small trial of only 24 patients and will be assessed over 24 weeks. This is an exciting study that if proven to be safe, may pave the way for many more similar studies using ICIs as IPF therapies. The challenge will be identifying those patients that are likely to have their pathologies exacerbated by ICI treatment.

There are also currently two trials (NCT03584802 – EXCHANGE-IPF and NCT03286556 - STRIVE-IPF) examining the therapeutic benefit of combining therapeutic plasma exchange with Rtx and intravenous Ig on AE-IPF. Therapeutic plasma exchange, also known as plasmapheresis or plasma exchange therapy, involves the removal of a patient's plasma and replacing it with a substitute solution or donor plasma. Plasma exchange is often used in autoimmune disorders to remove autoantibodies and immune complexes from the plasma to alleviate symptoms and modulate the immune response (Zanatta, Cozzi, Marson, & Cozzi, 2019). As already highlighted, there is increasing evidence of B cell abnormalities and autoantibodies present in IPF and AE-IPF and these are associated with disease severity. Therefore, by removing plasma autoantibodies and depleting B cells with Rtx, these trials aim to ameliorate B cell and autoantibody-mediated pulmonary injury. In a previous trial, 9 of 11 critically ill AE-IPF patients demonstrated some improvement in lung function after a combination of therapeutic plasma exchange with Rtx and intravenous Igs (Donahoe et al., 2015). Similar findings were shown in a comparative but larger study of 24 patients (Kulkarni et al., 2021). Autoantibody reduction therapy was associated with a reduction of supplemental oxygen requirement and/or an improved ability to ambulate in all 10 AE-IPF patients that survived for at least one year (Kulkarni et al., 2021). However, it is possible that the positive responses seen were due solely to Rtx treatment. Rtx has previously been shown to improve lung function in AE-IPF although the mechanism isn't clear, as Rtx is unlikely to affect autoantibody production as antibody producing plasmablasts and plasma cells don't express CD20. Furthermore, the assumption is that reducing blood autoantibodies will reduce pulmonary injury. However, this approach may not affect autoantibodies produced by resident plasma cells in the lung.

There are many molecules being investigated as therapeutic targets to treat IPF and other fibrotic lung conditions but many of these are still at the pre-clinical stage. Several therapeutic approaches used to target malignant B cells and plasma cells are being investigated as novel treatments for IPF. The proteasome inhibitor bortezomib depletes malignant plasma cells in multiple myeloma (MM) patients (Montefusco, Mussetti, Salas, Martinelli, & Cerchione, 2020) although it may have off target effects (Wang, 2011). Prele and colleagues demonstrated a reduction in Blm-induced lung fibrosis in mice following bortezomib treatment whereas depletion of CD20+ cells had no effect, suggesting antibody producing plasmablasts and plasma cells should be targeted to inhibit the progression of pulmonary fibrosis rather than all B cells (Prele et al., 2022). Differentiation of B cells into antibody producing plasmablasts and plasma cells leads to changes in the expression of cell surface markers, which have been used as targets for monoclonal antibody and chimeric antigen receptor (CAR)-T cell therapies (e.g. CD20, CD38, CD138, B cell maturation antigen [BCMA] and SlamF7) (Bou Zerdan et al., 2022; Geis et al., 2021). Bispecific antibodies (bsAb) have also been used clinically to deplete malignant plasma cells in MM (Killock, 2021). A bsAb is a single antibody designed with two different antigen binding domains; one binds a surface protein specific for a target cell, in this case malignant plasma cells, and the other binds CD3 to activate killer T cells. The bsAb acts as a bridge to link the cells together to allow the T cell to kill the target cell. For example, Teclistamab targets both BCMA expressed on plasma cells and CD3 on T cells, triggering T cells to attack plasma cells (Fig. 2). Teclistamab has recently been approved for the treatment of MM (Killock, 2021).

CAR-T cell therapy has been successful in treating blood cancers, in particular B cell malignancies (De Marco, Monzo, & Ojala, 2023) but have also been studied as therapies for non-malignant pathologies such as autoimmune and infectious diseases, cardiac fibrosis and cellular senescence (Mazzi, Hajdu, Ribeiro, & Bonamino, 2021). CAR-T cells are genetically engineered T cells that express receptors that both target a specific antigen and activate the T cell on the one receptor. Clinically successful CAR-T cells have targeted CD19 (present on all B cells) but CAR-T cells have also been developed to target BCMA (to treat MM (Sammartano et al., 2023) (Fig. 2). CAR-T cells have also been used to target fibroblast activation protein (FAP) which is a membrane protease that is highly expressed by activated fibroblasts including myofibroblasts (Hettiarachchi et al., 2020) and cancer associated fibroblasts (Park et al., 1999). Adoptive transfer of T cells that express a CAR against FAP resulted in a significant reduction in cardiac fibrosis and restoration of function after injury in mice (Aghajanian et al., 2019). Several studies have also shown anti-tumour activity of FAP-CAR-T cells in preclinical models of cancers. Targeting FAP with CAR-T cells in cancer is one approach being investigated to overcome the challenges of the tumour microenvironment such as restrictions to T cell extravasation, immune exclusion and immunosuppression, by depleting ECM producing cancer associated fibroblasts (Bughda, Dimou, D'Souza, & Klampatsa, 2021; Xiao et al., 2023).

CAR-T cells have also been used as a senolytic agent by targeting senescent cells (Merkt, Zhou, Han, & Lagares, 2021). Cellular senescence is a state in which cells lose their ability to proliferate. Although a normal part of the aging process, cellular senescence can also contribute to the pathogenesis of certain diseases including IPF by releasing various pro-inflammatory cytokines, chemokines, matrix metalloproteinases and ECM molecules. This senescence-associated secretory phenotype can perpetuate chronic inflammation, recruit immune cells, and promote fibroblast activation and fibrosis (Kellogg, Kellogg Jr., Musi, & Nambiar, 2021; Waters et al., 2018). Under normal circumstances senescent cells are cleared by the immune system, but this process is impaired in IPF. Urokinase-type plasminogen activator receptor is a cell-surface protein that is induced during senescence. Using urokinase-type plasminogen activator receptor-specific CAR-T cells, Amor and colleagues demonstrated the therapeutic potential of CAR-T cells to remove senescent cells in animal models as they extended the survival of mice with lung adenocarcinoma and restored tissue homeostasis in mice with experimentally induced liver fibrosis (Amor et al., 2020).

Vaccines are also being developed to reduce fibrosis by targeting myofibroblasts. Sobecki and colleagues identified overexpression of the metalloproteinase disintegrin and metalloproteinase domain-containing protein 12 (ADAM12) and transcription factor Gli1 in fibrotic foci of Blm treated mice which appeared to co-localise with myofibroblasts. They then immunised mice with either ADAM12 or Gli1 prior