Casein Kinase 1δ/ε Inhibitor, PF670462 Attenuates the Fibrogenic Effects of Transforming Growth Factor-β in Pulmonary Fibrosis

Abstract

Transforming growth factor-beta (TGF-β) is a major mediator of fibrotic diseases, including idiopathic pulmonary fibrosis (IPF). However, therapeutic global inhibition of TGF-β is limited by unwanted immunosuppression and mitral valve defects. We performed an extensive literature search to uncover a little-known connection between TGF-β signaling and casein kinase (CK) activity. We have examined the abundance of CK1 delta and epsilon (CK1δ/ε) in lung tissue from IPF patients and non-diseased controls, and investigated whether inhibition of CK1δ/ε with PF670462 inhibits pulmonary fibrosis. CK1δ/ε levels in lung tissue from IPF patients and non-diseased controls were assessed by immunohistochemistry. Anti-fibrotic effects of the CK1δ/ε inhibitor PF670462 were assessed in pre-clinical models, including acute and chronic bleomycin mouse models and in vitro experiments on spheroids made from primary human lung fibroblast cells from IPF and control donors, and human A549 alveolar-like adenocarcinoma-derived epithelial cells. Increased expression of CK1δ and ε in IPF lungs compared to non-diseased controls was accompanied by increased levels of the product, phospho-period 2. In vitro, PF670462 prevented TGF-β-induced epithelial-mesenchymal transition. The stiffness of IPF-derived spheroids was reduced by PF670462 and TGF-β-induced fibrogenic gene expression was inhibited. The CK1δ/ε inhibitor PF670462 administered systemically or locally by inhalation prevented both acute and chronic bleomycin-induced pulmonary fibrosis in mice. PF670462 administered in a ‘therapeutic’ regimen (day 7 onward) prevented bleomycin-induced lung collagen accumulation. Elevated expression and activity of CK1 δ and ε in IPF and anti-fibrogenic effects of the dual CK1δ/ε inhibitor, PF670462, support CK1δ/ε as novel therapeutic targets for IPF.

TGF‐β: Master regulator of inflammation and fibrosis

“…Previous commentaries on TGF‐β have highlighted contextualization as a means to make sense of its myriad actions, some of which are even mutually antagonistic. This contextualization includes the target tissue and cell types, soluble and mechanical microenvironments, concurrent exposure to other growth factors, inflammogens and modulators and bidirectional circadian influences, the latter likely having a relationship relevant to the effectiveness of the CLOCK (Circadian Locomotor Output Cycles Kaput)‐modulating casein kinase 1δ/ε inhibitor PF670462 in fibrogenesis …”

“…In contrast, blockade of downstream signalling that is specific to pathways influencing glucocorticoid activity may yield a more acceptable safety/efficacy profile for chronic inhibition in respiratory disease. As the casein kinase 1δ/ε inhibitor, PF670462, also shows glucocorticoid‐enhancing activity and overcomes TGF‐β signalling of resistance, targeting this pathway offers a potentially safe TGF‐β modulator with activities that include suppression of fibrogenesis and allergic inflammation …”

“…This contextualization includes the target tissue and cell types, soluble and mechanical microenvironments, concurrent exposure to other growth factors, inflammogens and modulators and bidirectional circadian influences, 1 the latter likely having a relationship relevant to the effectiveness of the CLOCK (Circadian Locomotor Output Cycles Kaput)-modulating casein kinase 1δ/ε inhibitor PF670462 in fibrogenesis. 2 The contribution of TGF-β to chronic respiratory disease is extensively evidenced. 3 In cystic fibrosis, TGF-β gene polymorphism, which results in increased production, is recognized as a modifier of variable disease phenotypes.…”

Oncogenic and Circadian Effects of Small Molecules Directly and Indirectly Targeting the Core Circadian Clock

“…Although different mechanisms for regulating cell proliferation and migration via CKIε/δ activity may exist, we suspect that inhibition of migration via PF-670462 was induced through the TGFβ pathway: it has been shown that this molecule can prevent TGFβinduced epithelial-mesenchymal transitions that result in enhanced motility. 59 Circadian period length may not be correlated with cell migration. Many cellular functions, including cell cycle, proliferation, and migration, are controlled by circadian rhythms, but little is known regarding the association between period length and the rates of these processes.…”

A-Kinase Anchoring Proteins Diminish TGF-β1 / Cigarette Smoke-Induced Epithelial-to-Mesenchymal Transition

“…Based on gene and protein expression of EMT markers, 3 ng/ml TGF-β1 treatment for 24 hours was used in current study. Cells were pretreated for 30 minutes before stimulation with TGF-β1 for 24 hours with st-Ht31 (50 μM) to disrupt AKAP-PKA interaction [21] or with the casein kinase 1δ/ε inhibitor PF-670462 (1 and 10 μM) [22] to confirm that TGF-β1-induced EMT could be reversed in BEAS-2B cells. The β2-agonist fenoterol (0.001-10 μM), the phosphodiesterase (PDE)4 inhibitor rolipram (1 or 10 μM), the PDE3 inhibitor cilostamide (10 μM) and adenylyl cyclase agonist forskolin (10 μM) were added 30 minutes before 24 hours TGF-β1 stimulation.…”