Antifibrotic Drug Pirfenidone Research Articles

Multiple hour antifibrotic drug release enabled by a thermosensitive quadpolymer.

Injectable drug delivery for uterine fibroid therapy is an ambitious, possibly fertility-preserving concept, that could meet the challenges associated with the structure of these tumors and their location in the uterus. This study was conducted to advance a thermosensitive injectable quadpolymer for effective sustained release of anti-fibrotic drug formulations and to evaluate the feasibility of its use for delivery of the anti-fibrotic drug pirfenidone as a therapy to reduce fibroid cell proliferation. A series of quadpolymers were prepared by free radical polymerization of N-isopropylacrylamide (NIPAM) with different amounts of polylactic acid functionalized hydroxyethyl methacrylate (HEMA-PLA), acrylic acid (AAc), and methacrylate functionalized hyperbranched polyglycerol (HPG-MA) to optimize the sol-gel phase transition temperature and mechanical stiffness. Poly(NIPAM-co-HEMA-PLA-co-AAc-co-HPG-MA) with feed ratio (83-7-1-9), at 17% w/v, readily formed an aqueous solution that could be manipulated by syringe at room temperature. The quadpolymer also rapidly formed a stable gel at physiological body temperature, and partially biodegraded over time as confirmed by several spectroscopic characterization techniques. To evaluate the potential range of utility, quadpolymer 83-7-1-9 was loaded in-vitro with caffeine (a prototype hydrophilic drug) or the hydrophobic drug pirfenidone. Pirfenidone-loaded quadpolymer 83-7-1-9 formulations released 50% of drug loaded in double the time as compared to other reported liposome and nanoparticle injectable pirfenidone formulations. Furthermore, treatment of cultured fibroid cells with pirfenidone-loaded quadpolymer 83-7-1-9 formulations confirmed that activity of pirfenidone was preserved and proliferation of fibroid cells was inhibited. These results support that quadpolymer 83-7-1-9 is a promising candidate to be further developed for localized delivery of drugs for uterine fibroid therapy.

Ternary inulin hydrogel with long-term intestinal retention for simultaneously reversing IBD and its fibrotic complication.

Excessive accumulation of reactive oxygen and nitrogen species (RONS) and dysbiosis of intestinal microbiota are pivotal symptoms for inflammatory bowel disease (IBD) and its associated complications, such as intestinal fibrosis. This research introduces a probiotic inulin hydrogel loaded with polypyrrole (PPy) nanozymes and antifibrotic drug pirfenidone (PFD) (PPy/PFD@Inulin gel) designed for the concurrent amelioration of IBD and its fibrotic complication. Upon oral administration, the inulin gel matrix could extend the gastrointestinal residence time of PPy nanozymes and PFD, facilitating the efficient reduction of pro-inflammatory cytokine levels and enhancement of the intestinal epithelial barrier repair as well as the suppression of intestinal fibrosis through sustained RONS scavenging, modulation of gut microbiota and attenuation of the TGF-β/Smad signaling pathway to inhibit fibroblast proliferation. Notably, the PPy/PFD@Inulin gel demonstrated significant prophylactic and therapeutic efficacy in acute and chronic colitis as well as intestinal fibrosis induced by dextran sodium sulfate (DSS) in mouse models. Thus, the engineered ternary PPy/PFD@Inulin gel offered a pioneered paradigm for simultaneous reversal of IBD and its associated complications, such as intestinal fibrosis, in a single therapeutic regimen.

Injectable Tissue Adhesive Microgel Composite Containing Antifibrotic Drug for Vocal Fold Scarring.

Vocal fold (VF) scarring, a complex problem in laryngology, results from injury and inflammation of the layered architecture of the VFs. The resultant voice hoarseness, for which successful therapeutic options are currently limited, affects the patient's quality of life. A promising strategy to reverse this disorder is the use of antifibrotic drugs. The present study proposes a novel microbead-embedded injectable hydrogel that can sustain the release of the anti-fibrotic drug pirfenidone (PFD) for vocal fold scarring. Microbeads were developed using sodium alginate and gelatin, which were further embedded into a biomimetic and tissue adhesive gellan gum (GG) hydrogel. The microbead-embedded hydrogel exhibited improved injectability, viscoelasticity, tissue adhesiveness, degradability, and swelling compared to the hydrogel without beads. Additionally, the bead-embedded hydrogel could sustain the release of the PFD for a week. In vitro studies showed that the drug-loaded hydrogel could reduce the migration and proliferation of fibroblast cells in a dose-dependent manner. In summary, this study demonstrates the potential of a PFD-loaded injectable hydrogel with enhanced viscoelastic and tissue-adhesive properties for vocal fold scarring applications.

Pirfenidone and Nintedanib in Pulmonary Fibrosis: Lights and Shadows.

Pirfenidone and Nintedanib are specific drugs used against idiopathic pulmonary fibrosis (IPF) that showed efficacy in non-IPF fibrosing interstitial lung diseases (ILD). Both drugs have side effects that affect patients in different ways and have different levels of severity, making treatment even more challenging for patients and clinicians. The present review aims to assess the effectiveness and potential complications of Pirfenidone and Nintedanib treatment regimens across various ILD diseases. A detailed search was performed in relevant articles published between 2018 and 2023 listed in PubMed, UpToDate, Google Scholar, and ResearchGate, supplemented with manual research. The following keywords were searched in the databases in all possible combinations: Nintedanib; Pirfenidone, interstitial lung disease, and idiopathic pulmonary fibrosis. The most widely accepted method for evaluating the progression of ILD is through the decline in forced vital capacity (FVC), as determined by respiratory function tests. Specifically, a decrease in FVC over a 6-12-month period correlates directly with increased mortality rates. Antifibrotic drugs Pirfenidone and Nintedanib have been extensively validated; however, some patients reported several side effects, predominantly gastrointestinal symptoms (such as diarrhea, dyspepsia, and vomiting), as well as photosensitivity and skin rashes, particularly associated with Pirfenidone. In cases where the side effects are extremely severe and are more threatening than the disease itself, the treatment has to be discontinued. However, further research is needed to optimize the use of antifibrotic agents in patients with PF-ILDs, which could slow disease progression and decrease all-cause mortality. Finally, other studies are requested to establish the treatments that can stop ILD progression.

Structural optimization of Moracin M as novel selective phosphodiesterase 4 inhibitors for the treatment of idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and high mortality lung disease. Although the antifibrotic drugs pirfenidone and nintedanib could slow the rate of lung function decline, the usual course of the condition is inexorably to respiratory failure and death. Therefore, new approaches and novel therapeutic drugs for the treatment of IPF are urgently needed. And the selective PDE4 inhibitor has in vivo and in vitro anti-fibrotic effects in IPF models. But the clinical application of most PDE4 inhibitors are limited by their unexpected and severe side effects such as nausea, vomiting, and diarrhea. Herein, structure-based optimizations of the natural product Moracin M resulted in a novel a novel series of 2-arylbenzofurans as potent PDE4 inhibitors. The most potent inhibitor L13 has an IC50 of 36 ± 7 nM with remarkable selectivity across the PDE families and administration of L13·citrate (10.0 mg/kg) exhibited comparable anti-pulmonary fibrosis effects to pirfenidone (300 mg/kg) in a bleomycin-induced IPF mice model, indicate that L13 is a potential lead for the treatment of IPF.

Membrane Fusion Liposomes Deliver Antifibrotic and Chemotherapeutic Drugs Sequentially to Enhance Tumor Treatment Efficacy by Reshaping Tumor Microenvironment.

The intricate tumor microenvironment in triple-negative breast cancer (TNBC) hampers chemotherapy and immunotherapy efficacy due to dense extracellular matrix (ECM) by tumor-associated fibroblasts (TAFs). Nanoparticle-based therapies, especially "all-in-one" nanoparticles, have shown great potential in combined drug delivery strategies to reshape the tumor microenvironment and enhance therapeutic efficiency. However, these "all-in-one" nanoparticles suffer from limitations in targeting different target cells, uncontrollable dosing ratio, and disregarding the impact of delivery schedules. This study prepared cell membrane fusion liposomes (TAFsomes and CCMsomes) to load FDA-approved antifibrotic drug pirfenidone (PFD/TAFsomes) and antitumor drug doxorubicin (DOX/CCMsomes). These liposomes can specifically target TAFs cells and tumor cells, and combined administration can effectively inhibit TAFs activity, reshape the tumor microenvironment (TME), and significantly enhance the tumor chemotherapy efficacy. Combined drug delivery defeats "all-in-one" liposomes (DOX/PFD/Liposomes, DOX/PFD/TAFsomes, and DOX/PFD/CCMsomes) by flexibly adjusting the drug delivery ratio. Moreover, an asynchronous delivery strategy that optimizes the administration schedule not only further improves the therapeutic effect, but also amplifies the effectiveness of α-PD-L1 immunotherapy by modulating the tumor immune microenvironment. This delivery strategy provides a personalized treatment approach with clinical translation potential, providing new ideas for enhancing the therapeutic effect against solid tumors such as TNBC.

Revolutionizing the Treatment of Idiopathic Pulmonary Fibrosis: From Conventional Therapies to Advanced Drug Delivery Systems.

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive interstitial lung disease that has been well-reported in the medical literature. Its incidence has risen, particularly in light of the recent COVID-19 pandemic. Conventionally, IPF is treated with antifibrotic drugs-pirfenidone and nintedanib-along with other drugs for symptomatic treatments, including corticosteroids, immunosuppressants, and bronchodilators based on individual requirements. Several drugs and biologicals such as fluorofenidone, thymoquinone, amikacin, paclitaxel nifuroxazide, STAT3, and siRNA have recently been evaluated for IPF treatment that reduces collagen formation and cell proliferation in the lung. There has been a great deal of research into various treatment options for pulmonary fibrosis using advanced delivery systems such as liposomal-based nanocarriers, chitosan nanoparticles, PLGA nanoparticles, solid lipid nanocarriers, and other nanoformulations such as metal nanoparticles, nanocrystals, cubosomes, magnetic nanospheres, and polymeric micelles. Several clinical trials are also ongoing for advanced IPF treatments. This article elaborates on the pathophysiology of IPF, its risk factors, and different advanced drug delivery systems for treating IPF. Although extensive preclinical data is available for these delivery systems, the clinical performance and scale-up studies would decide their commercial translation.

Efficacy and safety of pirfenidone according to clinical trials

The results of clinical studies on the use of pirfenidone in idiopathic pulmonary fibrosis (IPF) are presented. The purpose of this study was to analyze the effectiveness and safety of pirfenidone in IPF according to clinical studies published in the scientific literature. IPF occupies an important place in the structure of interstitial lung diseases. Currently, IPF is understood as a condition in which there is chronic progressive fibrotic interstitial aseptic pneumonia of unknown etiology, quickly leading to disability and death. The antifibrotic drug pirfenidone has been approved by the EMA and FDA for use in patients with IPF. The clinical efficacy and safety of pirfenidone have been demonstrated in randomized clinical trials. When using pirfenidone, there were 47.9 % fewer patients with an absolute decrease in FVC ≥10 % or who died, and 132.5 % more patients with no decrease in FVC (p <0.001). In addition, the pirfenidone group had significantly better 6-minute walk test scores (p=0.04) and significantly improved progression-free survival (p<0.001). Long-term use of pirfenidone (up to 72 weeks) significantly reduced the rate of decline in FVC, prevented a reduction in distance traveled in the 6-minute walk test, and increased the time until signs of disease progression appeared compared with placebo. Pirfenidone demonstrated a good safety profile; in most cases, adverse effects were mild, disappeared when the drug dose was reduced, and had no adverse long-term consequences. As a result of a generalized analysis of adverse events recorded in the CAPACITY, ASCEND, and RECAP studies, it was found that long-term (maximum duration was 9.9 years) treatment with pirfenidone was accompanied by nausea in 37.6 % of cases, diarrhea in 28.1 %, dyspepsia — in 18.4 %, vomiting — in 15.9 %, and skin rashes — in 25.0 %.

The diagnosis and treatment of idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive fibrosing interstitial lung disease (ILD) of unknown cause. The prognosis of IPF is poor, respiratory failure is the most common cause of mortality. Velcro rales are typical on respiratory system examination. Clubbing is seen in 30-60% of IPF cases. There is no laboratory test specific to IPF. Usual interstitial pneumonia (UIP) pattern is seen in IPF. UIP features in high-resolution computed tomography (HRCT); peripheral subpleural bibasilar reticular opacities, honeycombing, traction bronchiectasis and interseptal thickening. It shows craniocaudal localization. Diagnosis of IPF; It is diagnosed by the combination of HRCT findings and clinical findings. Antifibrotic drugs (Pirfenidone and Nintedanib) slow down the progression of IPF and reduce the number of annual attacks and reduce the frequency of hospitalization.

Capsaicin ameliorate pulmonary fibrosis via antioxidant Nrf-2/ PPAR- γ pathway activation and inflammatory TGF-β1/ NF-κB/COX II pathway inhibition.

Bleomycin is an effective antibiotic with a significant anticancer properties, but its use is limited due to its potential to induce dose-dependent pulmonary fibrosis. Therefore, this study aimed to assess the therapeutic potential of Capsaicin as an additional treatment to enhance patient tolerance to Bleomycin compared to the antifibrotic drug Pirfenidone. Pulmonary fibrosis was induced in rats through by a single intratracheal Bleomycin administration in day zero, followed by either Capsaicin or Pirfenidone treatment for 7days. After the animals were sacrificed, their lungs were dissected and examined using various stains for macroscopic and histopathological evaluation. Additionally, the study assessed various antioxidant, anti-inflammatory, and antifibrotic parameters were assessed. Rats exposed to Bleomycin exhibited visible signs of fibrosis, histopathological alterations, increased collagen deposition, and elevated mucin content. Bleomycin also led to heightened increased inflammatory cells infiltration in the bronchoalveolar lavage, elevated fibrosis biomarkers such as hydroxyproline, alpha-smooth muscle actin (α-SMA) and transforming growth factor-beta (TGF-β1), increased inflammatory markers including tumor necrosis factor-alpha (TNF-α), interlukine-6 (Il-6), interlukine-1β (Il-1β) nuclear factor-kappa B (NF-κB), and Cyclooxygenase-2 (COX-2), and transforming growth factor-beta (TGF-β1),. Furthermore, it reduced the expression of peroxisome proliferator-activated receptor-gamma (PPAR-γ), increased oxidative stress biomarkers like nitric oxide (NO), malondialdehyde (MDA), myeloperoxidase (MPO) and protein carbonyl. Bleomycin also decreased the expression of nuclear factor erythroid 2-related factor 2 (Nrf-2), reduced glutathione (GSH), total antioxidant capacity, and the activities of catalase and superoxide dismutase (SOD). Treating the animals with Capsaicin and Pirfenidone following Bleomycin exposure resulted in improved lung macroscopic and microscopic characteristics, reduced collagen deposition (collagen I and collagen III) and mucin content, decreased inflammatory cell infiltration, lowered levels of hydroxyproline, α-SMA, and TGF-β1, decreased TNF-α, Il-6, Il-1β, NF-κB, and COX-2, increased PPAR-γ and Nrf-2 expression, and improvement improved in all oxidative stress biomarkers. In summary, Capsaicin demonstrates significant antifibrotic activity against Bleomycin-induced lung injury that may be attributed, at least in part, to the antioxidant and anti-inflammatory activities of Capsaicin mediated by upregulation of PPAR-γ and Nrf-2 expression and decreasing. TGF-β1, NF-κB and COX II proteins concentrations.

Effects of pirfenidone on scar size and ventricular remodeling after myocardial infarction: a preclinical study.

An intense fibrotic response after myocardial infarction (MI) may lead to scar expansion and left ventricular (LV) remodeling. We investigated the effects of the antifibrotic drug pirfenidone in this setting. Male Wistar rats were randomized to: sham procedure (n = 13), reperfused MI-induced by ligating the left anterior descending artery (LAD) for 45 min (n = 17), reperfused MI plus standard therapy (aspirin, angiotensin-converting enzyme inhibitor, beta blocker, and mineralocorticoid receptor antagonist) (n = 17), reperfused MI plus pirfenidone alone (n = 17), or reperfused MI plus standard therapy and pirfenidone (n = 17). Rats surviving MI induction underwent cardiac magnetic resonance scans after 72 h and 30 days from MI, and were sacrificed on day 31. Rats completing the whole protocol numbered 11 in the sham group, 9 in the untreated MI group, 8 in the standard treatment group, 9 in the pirfenidone alone group, and 9 in the standard treatment plus pirfenidone group. No significant differences emerged between LV volumes, ejection fraction or mass at 30 days or the differences from 72 h to 30 days. Small, nonsignificant differences between rats on pirfenidone alone vs. those on standard therapy emerged. The total extent of LV fibrosis, quantified as area and percentage of the tissue sample, did not differ significantly between rats on pirfenidone alone vs. those on standard therapy alone. Pirfenidone does not have additional effects on LV remodeling or fibrosis compared with standard therapy, but its effects are similar to standard therapy alone.

Photodermatit developed with pirfenidone treatment in patient diagnosed with IPF

Idiopathic pulmonary fibrosis (IPF) is the most common of the major idiopathic interstitial pneumonia (IIP) of unknown etiology, often presenting in advanced age, limited to the lungs, characterized by histopathologically and/or radiologically usual interstitial pneumonia (UIP) pattern, and progressing with chronic, progressive fibrosis. is the most common form. Since IPF is a disease with irreversible fibrosis, there is no curative treatment except lung transplantation. The aim of the treatments given is to stop the progression of the disease, to prevent exacerbations and to prolong the survival time. The greatest improvement in the treatment of IPF in recent years has been the use of antifibrotic drugs (Pirfenidone and nintedanib) that prevent fibrosis developing in the lung parenchyma. In this case, we aimed to present the side effect of photodermatitis in a patient who applied to our outpatient clinic with complaints of shortness of breath and cough for many years and was diagnosed with IPF and started on pirfenidone treatment.

Increased CTGF expression in alveolar epithelial cells by cyclic mechanical stretch: Its mechanism and the therapeutic effect of pirfenidone

The mechanisms of fibrosis onset and development remain to be elucidated. However, it has been reported that mechanical stretch promotes fibrosis in various organs and cells, and may be involved in the pathogenesis of pulmonary fibrosis. We demonstrated that ventilator-induced lung hyperextension stimulation in mice increased the expression of connective tissue growth factor (CTGF), a profibrotic cytokine, in lung tissue. Increased CTGF expression induced by cyclic mechanical stretch (CMS) was also observed in vitro using A549 human alveolar epithelial cells. Pathway analysis revealed that the induction of CTGF expression by CMS involved MEK phosphorylation. Furthermore, early growth response 1 (Egr-1) was identified as a transcription factor associated with CTGF expression. Finally, the antifibrotic drug pirfenidone significantly reduced CTGF expression, MEK phosphorylation, and Egr-1 levels induced by CMS. Thus, our results demonstrated that profibrotic cytokine CTGF induced by CMS may be a therapeutic target of pirfenidone.

A Protective Effect of Pirfenidone in Lung Fibroblast-Endothelial Cell Network via Inhibition of Rho-Kinase Activity.

Pulmonary fibrosis is a life-threatening disease that has been attributed to several causes. Specifically, vascular injury is thought to be involved in the pathogenesis of fibrosis. The effects of the antifibrotic drug pirfenidone on angiogenesis have not been fully elucidated. This study aimed to investigate the effects of pirfenidone in human lung fibroblast-endothelial cell co-culture network formation and to analyze the underlying molecular mechanisms. Human lung fibroblasts were co-cultured with human umbilical vein endothelial cells to establish a co-culture network cell sheet. The influence of pirfenidone was evaluated for protective effect on the endothelial network in cell sheets stimulated with transforming growth factor β (TGF-β). Results indicated that TGF-β disrupted the network formation. Pirfenidone and Y27632 (Rho-associated coiled-coil containing protein kinase [Rho-kinase or ROCK] inhibitor) protected against the TGF-β-induced endothelial network disruption. TGF-β activated Rho-kinase signaling in cells composing the co-culture cell sheet, whereas pirfenidone and Y27632 inhibited these effects. In conclusion, TGF-β-induced Rho-kinase activation and disrupted endothelial network formation. Pirfenidone suppressed TGF-β-induced Rho-kinase activity in cell sheets, thereby enabling vascular endothelial cells networks to be preserved in the cell sheets. These findings suggest that pirfenidone has potential vascular network-preserving effect via inhibiting Rho-kinase activity in vascular injury, which is a precursor to pulmonary fibrosis.

N-methyl-2-pyridone-5-carboxamide (N-Me-2PY) has potent anti-fibrotic and anti-inflammatory activity in a fibrotic kidney model: is it an old uremic toxin?

Uremic toxins accumulate in renal tissues and cells due to chronic kidney disease (CKD). Abnormalities in nicotinamide adenine dinucleotide (NAD +) metabolism lead to the progression of CKD. NAD + metabolites, such as N-methyl-2-pyridone-5-carboxamide (N-Me-2PY) and N-methyl-4-pyridone-5-carboxamide (N-Me-4PY), have been recognized as uremic toxins. However, no reports have validated whether they are actually harmful to the body. Therefore, we focused on the structural similarity of these metabolites to the anti-fibrotic drug pirfenidone and evaluated their effects on renal fibrosis. Each NAD + metabolite was treated with TGFβ1 to kidney fibroblasts or tubular epithelial cells, and quantitative RT-PCR and Western blot analysis were conducted. N-Me-2PY was orally administered to a ligated murine kidney fibrosis model (UUO) to evaluate its anti-fibrotic and toxic effects on the body. N-Me-2PY, N-Me-4PY, and nicotinamide N-oxide (NNO) inhibited TGFβ1-induced fibrosis and inflammatory gene expression in kidney fibroblasts. N-Me-2PY strongly suppressed the expression of types I and III collagen, αSMA, and IL-6. N-Me-2PY also suppressed TGFβ1-induced type I collagen and IL-6 expression in renal tubular epithelial cells. No toxic effect was observed with N-Me-2PY treatment, while attenuating renal fibrosis and tubular dilation in UUO mice. Suppression of various fibrosis- and inflammation-related genes was also observed. N-Me-2PY did not inhibit TGFβ1-induced Smad3 phosphorylation but inhibited Akt phosphorylation, suggesting that N-Me-2PY exerts anti-fibrotic and anti-inflammatory effects through Akt inhibition, similar to pirfenidone. NAD + metabolites, such as N-Me-2PY, are not uremic toxins but are potential therapeutic agents that have anti-fibrotic effects in CKD.

PO-01-161 PIRFENIDONE MITIGATES THE DEVELOPMENT OF ARRHYTHMOGENIC ATRIAL FIBROSIS DUE TO PREMATURE ATRIAL COMPLEXES IN A SWINE MODEL

Atrial fibrosis is one of the major pathophysiologic mechanisms underlying the progression of atrial arrhythmias. We previously demonstrated in a swine model that frequent premature atrial complexes (PACs) promote atrial fibrillation (AF) via atrial fibrosis and slow conduction. Whether such fibrosis can be reversed or prevented is unknown. To evaluate the effect of (1) PAC eradication and (2) the antifibrotic drug pirfenidone (PFD) on atrial fibrosis in a swine model of frequent, dyssynchronous PACs. Thirty-five swine were exposed to 50% paced PACs from the lateral left atrium (LA) via the coronary sinus for 16 weeks in 4 groups: (1) controls without PACs (CTRL, n=10); (2) PAC group (PAC; n=10); 3) PACs followed by additional 6 weeks of PAC cessation (Rec-PAC, n=10); and (4) PACs treated with oral PFD (801mg 2 times per day) (PFD-PAC; n=5). Detailed EP study and echocardiography were performed at baseline and prior to sacrifice followed by blinded histological quantification of LA fibrosis. In the PAC group, peak atrial strain declined significantly at 16 weeks compared to CTRL. Compared to the PAC group, both the Rec-PAC and PFD-PAC groups had a less significant decline in peak atrial strain (terminal - baseline: PAC -17.7±3.3% vs. Rec-PAC -8.3±3.2% vs. PFD-PAC -11.5±3.0% vs. CTRL -0.7±4.2%; p<0.001). Conduction velocity (CV) was significantly decreased in PAC and Rec-PAC, but this decrease was attenuated in the PFD-PAC group (PAC CV 1.1±0.2m/s vs. Rec-PAC 1.2±0.1 m/s vs. PFD-PAC 1.3±0.1m/s vs. CTRL 1.5±0.2m/s; p<0.001). Posterior wall LA fibrosis was significantly increased in the PAC group, with no significant change after 6 weeks of recovery without PACs. However, the PFD-PAC group showed significantly lower fibrosis than either the PAC or Rec-PAC groups (Figure, Panel A). The PFD-PAC group also had less AF inducibility compared to the PAC and Rec-PAC groups (Panel B). In a swine model, frequent PACs produce atrial fibrosis and arrhythmogenic AF substrate. This fibrotic substrate did not regress after 6 weeks without PACs. However, treatment with PFD significantly attenuated the development of atrial fibrosis and AF inducibility. These findings suggest: (1) early management may be important for preventing ectopy-induced adverse atrial remodeling, and (2) pharmacological therapy targeted at the fibrotic substrate may play an important, novel role in the prevention of atrial remodeling and the substrate for developing AF.

PO-455617-17 PIRFENIDONE MITIGATES THE DEVELOPMENT OF ARRHYTHMOGENIC ATRIAL FIBROSIS DUE TO PREMATURE ATRIAL COMPLEXES IN A SWINE MODEL

Atrial fibrosis is one of the major pathophysiologic mechanisms underlying the progression of atrial arrhythmias. We previously demonstrated in a swine model that frequent premature atrial complexes (PACs) promote atrial fibrillation (AF) via atrial fibrosis and slow conduction. Whether such fibrosis can be reversed or prevented is unknown. To evaluate the effect of (1) PAC eradication and (2) the antifibrotic drug pirfenidone (PFD) on atrial fibrosis in a swine model of frequent, dyssynchronous PACs. Thirty-five swine were exposed to 50% paced PACs from the lateral left atrium (LA) via the coronary sinus for 16 weeks in 4 groups: (1) controls without PACs (CTRL, n=10); (2) PAC group (PAC; n=10); 3) PACs followed by additional 6 weeks of PAC cessation (Rec-PAC, n=10); and (4) PACs treated with oral PFD (801mg 2 times per day) (PFD-PAC; n=5). Detailed EP study and echocardiography were performed at baseline and prior to sacrifice followed by blinded histological quantification of LA fibrosis. In the PAC group, peak atrial strain declined significantly at 16 weeks compared to CTRL. Compared to the PAC group, both the Rec-PAC and PFD-PAC groups had a less significant decline in peak atrial strain (terminal - baseline: PAC -17.7±3.3% vs. Rec-PAC -8.3±3.2% vs. PFD-PAC -11.5±3.0% vs. CTRL -0.7±4.2%; p<0.001). Conduction velocity (CV) was significantly decreased in PAC and Rec-PAC, but this decrease was attenuated in the PFD-PAC group (PAC CV 1.1±0.2m/s vs. Rec-PAC 1.2±0.1 m/s vs. PFD-PAC 1.3±0.1m/s vs. CTRL 1.5±0.2m/s; p<0.001). Posterior wall LA fibrosis was significantly increased in the PAC group, with no significant change after 6 weeks of recovery without PACs. However, the PFD-PAC group showed significantly lower fibrosis than either the PAC or Rec-PAC groups (Figure, Panel A). The PFD-PAC group also had less AF inducibility compared to the PAC and Rec-PAC groups (Panel B). In a swine model, frequent PACs produce atrial fibrosis and arrhythmogenic AF substrate. This fibrotic substrate did not regress after 6 weeks without PACs. However, treatment with PFD significantly attenuated the development of atrial fibrosis and AF inducibility. These findings suggest: (1) early management may be important for preventing ectopy-induced adverse atrial remodeling, and (2) pharmacological therapy targeted at the fibrotic substrate may play an important, novel role in the prevention of atrial remodeling and the substrate for developing AF.

Triple-negative breast cancer cells invade adipocyte/preadipocyte-encapsulating geometrically inverted mammary organoids.

This paper describes the manufacture of geometrically inverted mammary organoids encapsulating primary mammary preadipocytes and adipocytes. Material manipulation in an array of 192 hanging drops induces cells to self-assemble into inside-out organoids where an adipose tissue core is enveloped by a cell-produced basement membrane, indicated by laminin V staining and then a continuous layer of mammary epithelial cells. This inverted tissue structure enables investigation of multiple mammary cancer subtypes, with a significantly higher extent of invasion by triple-negative MDA-MB-231 breast cancer cells compared to MCF7 cells. By seeding cancer cells into co-culture around pre-formed organoids with encapsulated preadipocytes/adipocytes, invasion through the epithelium, then into the adipose core is observable through acquisition of confocal image stacks of whole mount specimens. Furthermore, in regions of the connective tissue core where invasion occurs, there is an accumulation of collagen in the microenvironment. Suggesting that this collagen may be conducive to increased invasiveness, the anti-fibrotic drug pirfenidone shows efficacy in this model by slowing invasion. Comparison of adipose tissue derived from three different donors shows method consistency as well as the potential to evaluate donor cell-based biological variability. Insight box Geometrically inverted mammary organoids encapsulating primary preadipocytes/adipocytes (P/As) are bioengineered using a minimal amount of Matrigel scaffolding. Use of this eversion-free method is key to production of adipose mammary organoids (AMOs) where not only the epithelial polarity but also the entire self-organizing arrangement, including adipose position, is inside-out. While an epithelial-only structure can analyze cancer cell invasion, P/As are required for invasion-associated collagen deposition and efficacy of pirfenidone to counteract collagen deposition and associated invasion. The methods described strike a balance between repeatability and preservation of biological variability: AMOs form consistently across multiple adipose cell donors while revealing cancer cell invasion differences.

Pirfenidone affects human cardiac fibroblast proliferation and cell cycle activity in 2D cultures and engineered connective tissues

The anti-fibrotic drug pirfenidone (PFD) is currently in clinical testing for the treatment of heart failure with preserved ejection fraction; however, its effects on human cardiac cells have not been fully investigated. Therefore, we aimed to characterize the impact of PFD on human cardiac fibroblasts (CF) in 2D culture as well as in 3D-engineered connective tissues (ECT). We analyzed proliferation by automated cell counting and changes in signaling by immunoblotting. We generated ECT with different geometries to modify the cellular phenotype and investigated the effects of PFD on cell number and viability as well as on cell cycle activity. We further studied its effect on ECT compaction, contraction, stiffening, and strain resistance by ECT imaging, pole deflection analysis, and ultimate tensile testing. Our data demonstrate that PFD inhibits human CF proliferation in a concentration-dependent manner with an IC50 of 0.43 mg/ml and its anti-mitogenic effect was further corroborated by an inhibition of MEK1/2, ERK1/2, and riboprotein S6 (rpS6) phosphorylation. In ECT, a lower cell cycle activity was found in PFD-treated ECT and fewer cells resided in these ECT after 5 days of culture compared to the control. Moreover, ECT compaction as well as ECT contraction was impaired. Consequently, biomechanical analyses demonstrated that PFD reduced the stiffness of ECT. Taken together, our data demonstrate that the anti-fibrotic action of PFD on human CF is based on its anti-mitogenic effect in 2D cultures and ECT.

Regulation of TGF-β1-Smad-1-7 signaling to Inhibit Epithelial Mesenchymal Transition by Repurposed Anti-Fibrotic Drug Pirfenidone can Attenuate Lung Cancer Progression

Regulation of TGF-β1-Smad-1-7 signaling to Inhibit Epithelial Mesenchymal Transition by Repurposed Anti-Fibrotic Drug Pirfenidone can Attenuate Lung Cancer Progression