Kallistatin induces breast cancer cell apoptosis and autophagy by modulating Wnt signaling and microRNA synthesis

Abstract

Kallistatin is an endogenous protein that regulates differential signaling pathways and biological functions. Our previous studies showed that kallistatin gene therapy inhibited angiogenesis, tumor growth and metastasis in mice, and kallistatin protein suppressed Wnt-mediated growth, migration and invasion by blocking Wnt/β-catenin signaling pathway in breast cancer cells. In this study, we show that kallistatin reduced cell viability, and increased apoptotic cell death and caspase-3 activity in MDA-MB-231 breast cancer cells. Kallistatin also induced cancer cell autophagy, as evidenced by increased LC3B levels and elevated Atg5 and Beclin-1 expression; however, co-administration of Wnt or PPARγ antagonist GW9662 abolished these effects. Moreover, kallistatin via its heparin-binding site antagonized Wnt3a-induced cancer cell proliferation and increased PPARγ expression. Kallistatin inhibited oncogenic miR-21 synthesis associated with reduced Akt phosphorylation and Bcl-2 synthesis, but increased BAX expression. Kallistatin via PKC-ERK activation reduced miR-203 levels, leading to increased expression of suppressor of cytokine signaling 3 (SOCS3), a tumor suppressor. Conversely, kallistatin stimulated expression of the tumorigenic suppressors miR-34a and p53. Kallistatin’s active site is essential for suppressing miR-21 and miR-203, and stimulating miR-34a and SOCS3 expression. This is the first study to demonstrate that kallistatin’s heparin-binding site is essential for inhibiting Wnt-mediated effects, and its active site plays a key role in regulating miR-21, miR-203, miR-34a and SOCS3 synthesis in breast cancer cells. These findings reveal novel mechanisms of kallistatin in inducing apoptosis and autophagy in breast cancer cells, thus inhibiting tumor progression by regulation of Wnt/PPARγ signaling, as well as miR-21, miR-203 and miR-34a synthesis.

Protective Role of Kallistatin in Vascular and Organ Injury

“…36,40 Kallistatin's active site was found to be crucial for stimulating SOCS3 expression in macrophages through activation of a protein kinase C (PKC)-extracellular signal-regulated kinase (ERK) signaling pathway, illustrating a novel mechanism by which kallistatin protects against sepsis-induced organ damage. 32,52 These findings indicate that kallistatin administration significantly enhances survival and protects against organ damage during sepsis.…”

“…41 Likewise, kallistatin's heparin-binding site is essential for suppressing Wnt3a signaling and cancer cell proliferation. 32 Kallistatin antagonizes Wnt3a signaling by forming a complex with the Wnt co-receptor low-density lipoprotein receptor 6 (LRP6), as demonstrated in retinal epithelial and breast cancer cells, 42,43 thereby inhibiting Wnt3a-induced tumor cell proliferation, migration and invasion. 43 Moreover, reduced kallistatin levels are correlated with βII-spectrin (SPTBN1), an adapter protein during Wnt signaling in hepatocellular carcinoma.…”

REST represses miR-124 and miR-203 to regulate distinct oncogenic properties of glioblastoma stem cells

“…In addition, miR-203 is known to act as a tumor suppressor in other cancers. [30][31][32] Interestingly, in two previous publications, miR-203 positively regulated invasion as well as cell proliferation in some glioma cell lines. 28,29 However, our studies indicate that the REST-miR-203 pathway specifically regulates invasion but not cell proliferation or apoptosis in HR-GSCs in vitro and in mouse orthotopic tumor models.…”

Kallistatin in Sepsis: Protective Actions and Potential Therapeutic Applications

“…Kallistatin's active site is crucial for: (1) complex formation with tissue kallikrein and inhibiting tissue kallikrein activity and bioavailability [13,27]; (2) increasing eNOS and SIRT1 expression and activation, leading to elevated NO formation [28]; (3) stimulating SOCS3 expression [48]; and (4) interacting with a tyrosine kinase [28,48]. Kallistatin via its heparin-binding domain interacts with cell surface heparan sulfate proteoglycans, thereby antagonizing the following biological efects: (1) VEGF-mediated angiogenesis and vascular permeability [16,20]; (2) TNF-α-induced NF-κB activation, inlammation, oxidative stress, and apoptosis [20]; (3) HMGB1-induced inlammatory gene expression and oxidative stress [29]; (4) TGF-β-induced endothelial-mesenchymal transition (EndMT), and epithelialmesenchymal transition (EMT) [28]; (5) Wnt-mediated cancer cell proliferation, migration, invasion, and autophagy [42,44]; and (6) EGF-induced cancer cell migration and invasion (unpublished results). Thus, kallistatin, with its multifactorial activities, regulates a wide spectrum of biological processes, such as angiogenesis, inlammation, oxidative stress, apoptosis, ibrosis, and cancer development.…”

“…Circulatory kallistatin levels are also diminished in patients with liver disease, septic syndrome, diabetic retinopathy, severe pneumonia, inlammatory bowel disease, obesity, prostate, and colon cancer [33][34][35][36][37][38][39][40]. Kallistatin administration by gene or protein delivery protects against the pathogenesis of hypertension, heart and kidney damage, arthritis, inluenza virus infection, tumor growth, and metastasis in animal models [15][16][17][18][19][41][42][43][44][45][46]. Conversely, depletion of endogenous kallistatin by neutralizing antibody injection exacerbates cardiovascular and renal injury in hypertensive rats [47].…”