Idiopathic pulmonary fibrosis (IPF) is a fatal, chronic and progressive lung disease characterized by excessive inflammatory response, myofibroblast hyperplasia and deposition of extracellular matrix (ECM) [1]. Currently, the prevalence of Idiopathic Pulmonary Fibrosis (IPF) mirrors that of numerous cancers, and unfortunately, the prognosis for survival is bleak. Globally, the estimated incidence of IPF ranges from 1 to 13 cases per 100,000 people, while the prevalence is reported to be between 3 and 45 cases per 100,000 people. Onset typically occurs in individuals above middle age, with a higher prevalence in older men who are heavy smokers (with a history of more than 20 pack-year) [2]. As the world's population continues to age, awareness of the clinical manifestations of IPF grows, and certain risk factors such as air pollution become more prevalent, the global burden of this condition is anticipated to rise. The escalating numbers underscore the urgent need for effective interventions and treatments to address the growing impact of IPF on public health. Numerous studies have elucidated that several risk factors, such as cigarette smoking, environmental pollution, and viral infection, could induce repetitive microinjuries in lung, which lead to subsequent inflammatory response. Persistent inflammatory response contributed to increased cytokine secretion, with transforming growth factor β (TGF-β) playing a primary role in connecting inflammation to fibrogenesis [3], [4]. Through the recruitment of its downstream SMAD3 protein, TGF-β promoted the differentiation of fibroblasts to myofibroblasts, which aggregated abnormally and secreted α-smooth muscle actin (α-SMA) and collagen, resulting in ECM formation and the development of fibroblastic foci [5], [6]. Conversely, myofibroblasts could induce macrophage migration, triggering further inflammatory responses and accelerating the progression of fibrosis [7], [8]. Therefore, considering the significant impact of inflammatory responses on the onset and progression of IPF, inhibiting inflammatory response represents an effective therapeutic strategy for treating IPF.

Studies have reported that anti-inflammatory agents, such as corticosteroids, could alleviate the progression of IPF. Specifically, corticosteroids, especially prednisone, possess the ability to slow down IPF by inhibiting inflammatory responses [9], [10]. In addition to corticosteroids, pirfenidone (PFD, Fig. 1), the first approved anti-IPF agent, also exhibits anti-inflammatory activity. Furthermore, PFD demonstrates anti-fibrotic potency, making it a multi-target agent against IPF by inhibiting inflammatory responses and myofibroblast proliferation [11]. Mechanistic studies shown that PFD suppressed the expression of pro-inflammatory cytokines, like IL-6, IFN-γ and TNF-α, and delayed the progression of IPF through restraining the TGF-β/SMAD3 pathway [12]. However, the efficacy of PFD as an anti-IPF agent was limited, necessitating high dosages to achieve the desired therapeutic effects in clinical use (2403 mg per day) [13]. The rather high clinical dosage causes significant and serious side effect, including gastrointestinal upset and liver toxicity [14], [15]. Therefore, there was an urgent need to enhance the therapeutic efficacy of PFD with the objective of reducing the required dosage.

The insufficient therapeutic potency of PFD stimulated the exploration of modifications to its scaffold [16]. However, few modification strategies of PFD scaffold targeting inflammatory response process, the significant driving factor of IPF, have been investigated. Nonsteroidal anti-inflammatory drugs (NSAIDs) such as salicylic acid, (S)-ibuprofen, naproxen, and flurbiprofen (Fig. 1), were a significant class of anti-inflammatory agents that have been clinical used for decades. Recent studies have also demonstrated that a series of conjugates combining NSAIDs with carbonic anhydrase inhibitors (CAIs) effectively increased the anti-IPF potency by focusing on inflammatory response process [17]. Therefore, design and synthesis of conjugates consisting of PFD and NSAIDs may be a promising modification approach to enhance the anti-IPF potency of PFD by improving the anti-inflammatory activity.

Herein, we focused on the significant role of the inflammatory response in the progression of IPF, aiming to enhance the anti-IPF potency of PFD. To achieve this, we designed and synthesized several PFD-NSAIDs conjugates through a straightforward amide condensation reaction (Fig. 2). Initially, anti-proliferation activities of target compounds were evaluated using NIH3T3 cells as the model, resulting in the identification of the promising compound 5b. Subsequently, the anti-fibrotic effects, anti-inflammatory activities, and the underlying mechanisms of the promising compound 5b were investigated through experiments in vitro & vivo.