Design and synthesis of novel thiazole-derivatives as potent ALK5 inhibitors.

TGF-ß is an immunosuppressive cytokine and plays a key role in progression of cancer by inducing immunosuppression in tumor microenvironment. Therefore, inhibition of TGF-ß signaling pathway may provide a potential therapeutic intervention in treating cancers. Herein, we report the discovery of a series of novel thiazole derivatives as potent inhibitors of ALK5, a serine-threonine kinase which is responsible for TGF-ß signal transduction. Compound 29b was identified as a potent inhibitor of ALK5 with an IC50 value of 3.7 nM with an excellent kinase selectivity.

Liebig's law of the minimum in the TGF-ß/SMAD pathway.

Cells use signaling pathways to sense and respond to their environments. The transforming growth factor-ß (TGF-ß) pathway produces context-specific responses. Here, we combined modeling and experimental analysis to study the dependence of the output of the TGF-ß pathway on the abundance of signaling molecules in the pathway. We showed that the TGF-ß pathway processes the variation of TGF-ß receptor abundance using Liebig's law of the minimum, meaning that the output-modifying factor is the signaling protein that is most limited, to determine signaling responses across cell types and in single cells. We found that the abundance of either the type I (TGFBR1) or type II (TGFBR2) TGF-ß receptor determined the responses of cancer cell lines, such that the receptor with relatively low abundance dictates the response. Furthermore, nuclear SMAD2 signaling correlated with the abundance of TGF-ß receptor in single cells depending on the relative expression levels of TGFBR1 and TGFBR2. A similar control principle could govern the heterogeneity of signaling responses in other signaling pathways.

Design, synthesis and evaluation of a pyrazolo[3,4-d]pyrimidine derivative as a novel and potent TGFß1R1 inhibitor.

The transforming growth factor ß1 (TGFß1)/SMAD signaling pathway regulates many vital physiological processes. The development of potent inhibitors targeting activin receptor-like kinase 5 (ALK5) would provide potential treatment reagents for various diseases. A significant number of ALK5 inhibitors have been discovered, and they are currently undergoing clinical evaluation at various stages. However, the clinical demands were far from being met. In this study, we utilized an alternative conformation-similarity-based virtual screening (CSVS) combined with a fragment-based drug designing (FBDD) strategy to efficiently discover a potent and active hit with a novel chemical scaffold. After structural optimization in the principle of group replacement, compound 57 was identified as the most promising ALK5 inhibitor. Compound 57 demonstrated significant inhibitory effects against the TGF-ß1/SMAD signaling pathway. It could markedly attenuate the production of extracellular matrix (ECM) and deposition of collagen. Also, the lead compound showed adequate pharmacokinetic (PK) properties and good in vivo tolerance. Moreover, treatment with compound 57 in two different xerograph models showed significant inhibitory effects on the growth of pancreatic cancer cells. These results suggested that lead compound 57 refers as a promising ALK5 inhibitor both in vitro and in vivo, which merits further validation.

Targeting the TGF-ß signaling pathway: an updated patent review (2021-present).

INTRODUCTION: The TGF-ß signaling pathway is a complex network that plays a crucial role in regulating essential biological functions and is implicated in the onset and progression of multiple diseases. This review highlights the recent advancements in developing inhibitors targeting the TGF-ß signaling pathway and their potential therapeutic applications in various diseases. AREA COVERED: The review discusses patents on active molecules related to the TGF-ß signaling pathway, focusing on three strategies: TGF-ß activity inhibition, blocking TGF-ß receptor binding, and disruption of the signaling pathway using small molecule inhibitors. Combination therapies and the development of fusion proteins targeting multiple pathways are also explored. The literature search was conducted using the Cortellis Drug Discovery Intelligence database, covering patents from 2021 onwards. EXPERT OPINION: The development of drugs targeting the TGF-ß signaling pathway has made significant progress in recent years. However, addressing challenges such as specificity, systemic toxicity, and patient selection is crucial for their successful clinical application. Targeting the TGF-ß signaling pathway holds promise as a promising approach for the treatment of various diseases.

Synthesis and anti-liver fibrosis activity of imidazole and thiazole compounds containing amino acids.

Four series of imidazoles (15a-g, 20c, and 20d) and thiazoles (18a-g, 22a, and 22b) possessing various amino acids were synthesized and evaluated for activin receptor-like kinase 5 (ALK5) inhibitory activities in an enzymatic assay. Among them, compounds 15g and 18c showed the highest inhibitory activity against ALK5, with IC50 values of 0.017 and 0.025 µM, respectively. Compounds 15g and 18c efficiently inhibited extracellular matrix (ECM) deposition in TGF-ß-induced hepatic stellate cells (HSCs), and eventually suppressed HSC activation. Moreover, compound 15g showed a good pharmacokinetic (PK) profile with a favorable half-life (t1/2 = 9.14 h). The results indicated that these compounds exhibited activity targeting ALK5 and may have potential in the treatment of liver fibrosis; thus they are worthy of further study.

Radiation-induced effects on TGF-ß and PDGF receptor signaling in cancer-associated fibroblasts.

BACKGROUND: Cancer-associated fibroblasts (CAFs) consist of heterogeneous connective tissue cells and are often constituting the most abundant cell type in the tumor stroma. Radiation effects on tumor stromal components like CAFs in the context of radiation treatment is not well-described. AIM: This study explores potential changes induced by ionizing radiation (IR) on platelet-derived growth factor (PDGF)/PDGFRs and transforming growth factor-beta (TGF-ß)/TGFßRs signaling systems in CAFs. METHODS AND RESULTS: Experiments were carried out by employing primary cultures of human CAFs isolated from freshly resected non-small cell lung carcinoma tumor tissues. CAF cultures from nine donors were treated with one high (1 × 18 Gy) or three fractionated (3 × 6 Gy) radiation doses. Alterations in expression levels of TGFßRII and PDGFRα/ß induced by IR were analyzed by western blots and flow cytometry. In the presence or absence of cognate ligands, receptor activation was studied in nonirradiated and irradiated CAFs. Radiation exposure did not exert changes in expression of PDGF or TGF-ß receptors in CAFs. Additionally, IR alone was unable to trigger activation of either receptor. The radiation regimens tested did not affect PDGFRß signaling in the presence of PDGF-BB. In contrast, signaling via pSmad2/3 and pSmad1/5/8 appeared to be down-regulated in irradiated CAFs after stimulation with TGF-ß, as compared with controls. CONCLUSION: Our data demonstrate that IR by itself is insufficient to induce measurable changes in PDGF or TGF-ß receptor expression levels or to induce receptor activation in CAFs. However, in the presence of their respective ligands, exposure to radiation at certain doses appear to interfere with TGF-ß receptor signaling.

Utilizing multiscale engineered biomaterials to examine TGF-ß-mediated myofibroblastic differentiation.

Cells integrate many mechanical and chemical cues to drive cell signalling responses. Because of the complex nature and interdependency of alterations in extracellular matrix (ECM) composition, ligand density, mechanics, and cellular responses it is difficult to tease out individual and combinatorial contributions of these various factors in driving cell behavior in homeostasis and disease. Tuning of material viscous and elastic properties, and ligand densities, in combinatorial fashions would enhance our understanding of how cells process complex signals. For example, it is known that increased ECM mechanics and transforming growth factor beta (TGF-ß) receptor (TGF-ß-R) spacing/clustering independently drive TGF-ß signalling and associated myofibroblastic differentiation. However, it remains unknown how these inputs orthogonally contribute to cellular outcomes. Here, we describe the development of a novel material platform that combines microgel thin films with controllable viscoelastic properties and DNA origami to probe how viscoelastic properties and nanoscale spacing of TGF-ß-Rs contribute to TGF-ß signalling and myofibroblastic differentiation. We found that highly viscous materials with non-fixed TGF-ß-R spacing promoted increased TGF-ß signalling and myofibroblastic differentiation. This is likely due to the ability of cells to better cluster receptors on these surfaces. These results provide insight into the contribution of substrate properties and receptor localisation on downstream signalling. Future studies allow for exploration into other receptor-mediated processes.

Glycosaminoglycan modifications of betaglycan regulate ectodomain shedding to fine-tune TGF-ß signaling responses in ovarian cancer.

In pathologies including cancer, aberrant Transforming Growth Factor-ß (TGF-ß) signaling exerts profound tumor intrinsic and extrinsic consequences. Intense clinical endeavors are underway to target this pathway. Central to the success of these interventions is pinpointing factors that decisively modulate the TGF-ß responses. Betaglycan/type III TGF-ß receptor (TßRIII), is an established co-receptor for the TGF-ß superfamily known to bind directly to TGF-ßs 1-3 and inhibin A/B. Betaglycan can be membrane-bound and also undergo ectodomain cleavage to produce soluble-betaglycan that can sequester its ligands. Its extracellular domain undergoes heparan sulfate and chondroitin sulfate glycosaminoglycan modifications, transforming betaglycan into a proteoglycan. We report the unexpected discovery that the heparan sulfate glycosaminoglycan chains on betaglycan are critical for the ectodomain shedding. In the absence of such glycosaminoglycan chains betaglycan is not shed, a feature indispensable for the ability of betaglycan to suppress TGF-ß signaling and the cells' responses to exogenous TGF-ß ligands. Using unbiased transcriptomics, we identified TIMP3 as a key inhibitor of betaglycan shedding thereby influencing TGF-ß signaling. Our results bear significant clinical relevance as modified betaglycan is present in the ascites of patients with ovarian cancer and can serve as a marker for predicting patient outcomes and TGF-ß signaling responses. These studies are the first to demonstrate a unique reliance on the glycosaminoglycan chains of betaglycan for shedding and influence on TGF-ß signaling responses. Dysregulated shedding of TGF-ß receptors plays a vital role in determining the response and availability of TGF-ßs', which is crucial for prognostic predictions and understanding of TGF-ß signaling dynamics.

A New Cell Model Overexpressing sTGFBR3 for Studying Alzheimer's Disease In vitro.

BACKGROUND: Recent studies have suggested that abnormal microglial hyperactivation has an important role in the progression of Alzheimer's disease (AD). sTGFBR3 (a shed extracellular domain of the transforming growth factor type III receptor) is a newly identified target of microglia polarization dysregulation, whose overexpression can cause abnormal accumulation of transforming growth factor ß1 (TGF-ß1), promoting Aß, tau, and neuroinflammatory pathology. OBJECTIVE: The objective of this study is to develop and validate a new cell model overexpressing sTGFBR3 for studying AD in vitro. METHODS: BV2 cells (a microglial cell derived from C57/BL6 murine) were used as a cell model. Cells were then treated with different concentrations of lipopolysaccharide (LPS) (0, 1, or 0.3 µg/mL) for 12, 24, or 48h and then with or without sodium pervanadate (100 µM) for 30 min. Next, the effect surface optimization method was used to determine optimal experimental conditions. Finally, the optimized model was used to assess the effect of ZQX series compounds and vasicine on cell viability and protein expression. Expression of TGFBR3 and TNF-α was assessed using Western blot. MTT assay was used to assess cell viability, and enzyme- linked immunosorbent assay (ELISA) was employed to evaluate extracellular TGF-ß1 and sTGFBR3. RESULTS: LPS (0.3 µg/mL) treatment for 11 h at a cell density of 60% and pervanadate concentration (100 µM) incubation for 30 min were the optimal experimental conditions for increasing membrane protein TGFBR3 overexpression, as well as extracellular sTGFBR3 and TGF-ß1. Applying ZQX-5 and vasicine reversed this process by reducing extracellular TGF-ß1, promoting the phosphorylation of Smad2/3, a protein downstream of TGF-ß1, and inhibiting the release of the inflammatory factor TNF-α. CONCLUSION: This new in vitro model may be a useful cell model for studying Alzheimer's disease in vitro.

Cholesterol modulates type I/II TGF-ß receptor complexes and alters the balance between Smad and Akt signaling in hepatocytes.

Cholesterol mediates membrane compartmentalization, affecting signaling via differential distribution of receptors and signaling mediators. While excessive cholesterol and aberrant transforming growth factor-ß (TGF-ß) signaling characterize multiple liver diseases, their linkage to canonical vs. non-canonical TGF-ß signaling remained unclear. Here, we subjected murine hepatocytes to cholesterol depletion (CD) or enrichment (CE), followed by biophysical studies on TGF-ß receptor heterocomplex formation, and output to Smad2/3 vs. Akt pathways. Prior to ligand addition, raft-dependent preformed heteromeric receptor complexes were observed. Smad2/3 phosphorylation persisted following CD or CE. CD enhanced phospho-Akt (pAkt) formation by TGF-ß or epidermal growth factor (EGF) at 5 min, while reducing it at later time points. Conversely, pAkt formation by TGF-ß or EGF was inhibited by CE, suggesting a direct effect on the Akt pathway. The modulation of the balance between TGF-ß signaling to Smad2/3 vs. pAkt (by TGF-ß or EGF) has potential implications for hepatic diseases and malignancies.

Prodigiosin Inhibits Transforming Growth Factor ß Signaling by Interfering Receptor Recycling and Subcellular Translocation in Epithelial Cells.

Prodigiosin (PG) is a naturally occurring polypyrrole red pigment produced by numerous microorganisms including some Serratia and Streptomyces strains. PG has exhibited promising anticancer activity; however, the molecular mechanisms of action of PG on malignant cells remain ambiguous. Transforming growth factor-ß (TGF-ß) is a multifunctional cytokine that governs a wide array of cellular processes in development and tissue homeostasis. Malfunctions of TGF-ß signaling are associated with numerous human cancers. Emerging evidence underscores the significance of internalized TGF-ß receptors and their intracellular trafficking in initiating signaling cascades. In this study, we identified PG as a potent inhibitor of the TGF-ß pathway. PG blocked TGF-ß signaling by targeting multiple sites of this pathway, including facilitating the sequestering of TGF-ß receptors in the cytoplasm by impeding the recycling of type II TGF-ß receptors to the cell surface. Additionally, PG prompts a reduction in the abundance of receptors on the cell surface through the disruption of the receptor glycosylation. In human Caucasian lung carcinoma cells and human hepatocellular cancer cell line cells, nanomolar concentrations of PG substantially diminish TGF-ß-triggered phosphorylation of Smad2 protein. This attenuation is further reflected in the suppression of downstream target gene expression, including those encoding fibronectin, plasminogen activator inhibitor-1, and N-cadherin. SIGNIFICANCE STATEMENT: Prodigiosin (PG) emerges from this study as a potent TGF-ß pathway inhibitor, disrupting receptor trafficking and glycosylation and reducing TGF-ß signaling and downstream gene expression. These findings not only shed light on PG's potential therapeutic role but also present a captivating avenue towards future anti-TGF-ß strategies.

Inhibition of transforming growth factor-ß signals suppresses tumor formation by regulation of tumor microenvironment networks.

The tumor microenvironment (TME) consists of cancer cells surrounded by stromal components including tumor vessels. Transforming growth factor-ß (TGF-ß) promotes tumor progression by inducing epithelial-mesenchymal transition (EMT) in cancer cells and stimulating tumor angiogenesis in the tumor stroma. We previously developed an Fc chimeric TGF-ß receptor containing both TGF-ß type I (TßRI) and type II (TßRII) receptors (TßRI-TßRII-Fc), which trapped all TGF-ß isoforms and suppressed tumor growth. However, the precise mechanisms underlying this action have not yet been elucidated. In the present study, we showed that the recombinant TßRI-TßRII-Fc protein effectively suppressed in vitro EMT of oral cancer cells and in vivo tumor growth in a human oral cancer cell xenograft mouse model. Tumor cell proliferation and angiogenesis were suppressed in tumors treated with TßRI-TßRII-Fc. Molecular profiling of human cancer cells and mouse stroma revealed that K-Ras signaling and angiogenesis were suppressed. Administration of TßRI-TßRII-Fc protein decreased the expression of heparin-binding epidermal growth factor-like growth factor (HB-EGF), interleukin-1ß (IL-1ß) and epiregulin (EREG) in the TME of oral cancer tumor xenografts. HB-EGF increased proliferation of human oral cancer cells and mouse endothelial cells by activating ERK1/2 phosphorylation. HB-EGF also promoted oral cancer cell-derived tumor formation by enhancing cancer cell proliferation and tumor angiogenesis. In addition, increased expressions of IL-1ß and EREG in oral cancer cells significantly enhanced tumor formation. These results suggest that TGF-ß signaling in the TME controls cancer cell proliferation and angiogenesis by activating HB-EGF/IL-1ß/EREG pathways and that TßRI-TßRII-Fc protein is a promising tool for targeting the TME networks.

Chimeric TßRII-SE/Fc overexpression by a lentiviral vector exerts strong antitumoral activity on colorectal cancer-derived cell lines in vitro and on xenografts.

The TGF signaling pathway is a key regulator of cancer progression. In this work, we report for the first time the antitumor activity of TßRII-SE/Fc, a novel peptibody whose targeting domain is comprised of the soluble endogenous isoform of the human TGF-ß type II receptor (TßRII-SE). Overexpression of TßRIISE/Fc reduces in vitro cell proliferation and migration while inducing cell cycle arrest and apoptosis in human colorectal cancer-derived cell lines. Moreover, TßRII-SE/Fc overexpression reduces tumorigenicity in BALB/c nude athymic mice. Our results revealed that TRII-SE/Fc-expressing tumors were significantly reduced in size or were even incapable of developing. We also demonstrated that the novel peptibody has the ability to inhibit the canonical TGF-ß and BMP signaling pathways while identifying SMAD-dependent and independent proteins involved in tumor progression that are modulated by TßRII-SE/Fc. These findings provide insights into the underlying mechanism responsible for the antitumor activity of TßRII-SE/Fc. Although more studies are required to demonstrate the effectiveness and safety of the novel peptibody as a new therapeutic for the treatment of cancer, our initial in vitro and in vivo results in human colorectal tumor-derived cell lines are highly encouraging. Our results may serve as the foundation for further research and development of a novel biopharmaceutical for oncology.

Advanced biosensors for nanomaterial-based detection of transforming growth factor alpha and beta, a class of major polypeptide regulators.

Transforming growth factors (TGFs) regulate several cellular processes including, differentiation, growth, migration, extracellular matrix production, and apoptosis. TGF alpha (TGF-α) is a heterogeneous molecule containing 160 amino acid residues. It is a potent angiogenesis promoter that is activated by JAK-STAT signaling. Whereas TGF beta (TGF-ß) consists of 390-412 amino acids. Smad and non-Smad signaling both occur in TGF beta. It is linked to immune cell activation, differentiation, and proliferation. It also triggers pre-apoptotic responses and inhibits cell proliferation. Both growth factors have a promising role in the development and homeostasis of tissues. Defects such as autoimmune diseases and cancer develop mechanisms to modulate checkpoints of the immune system resulting in altered growth factors profile. An accurate amount of these growth factors is essential for normal functioning, but an exceed or fall behind the normal level is alarming as it is linked to several disorders. This demands techniques for TGF-α and TGF-ß profiling to effectively diagnose diseases, monitor their progression, and assess the efficacy of immunotherapeutic drugs. Quantitative detection techniques including the emergence of biosensing technology seem to accomplish the purpose. Until the present time, few biosensors have been designed in the context of TGF-α and TGF-ß for disease detection, analyzing receptor binding, and interaction with carriers. In this paper, we have reviewed the physiology of transforming growth factor alpha and beta, including the types, structure, function, latent/active forms, signaling, and defects caused. It involves the description of biosensors on TGF-α and TGF-ß, advances in technology, and future perspectives.

Sertoli Cell-Specific Activation of Transforming Growth Factor Beta Receptor 1 Leads to Testicular Granulosa Cell Tumor Formation.

The transforming growth factor ß (TGFß) superfamily, consisting of protein ligands, receptors, and intracellular SMAD transducers, regulates fundamental biological processes and cancer development. Our previous study has shown that sustained activation of TGFß receptor 1 (TGFBR1) driven by anti-Mullerian hormone receptor type 2 (Amhr2)-Cre in the mouse testis induces the formation of testicular granulosa cell tumors (TGCTs). As Amhr2-Cre is expressed in both Sertoli cells and Leydig cells, it remains unclear whether the activation of TGFBR1 in Sertoli cells alone is sufficient to induce TGCT formation. Therefore, the objective of this study was to determine whether Sertoli cell-activation of TGFBR1 drives oncogenesis in the testis. Our hypothesis was that overactivation of TGFBR1 in Sertoli cells would promote their transdifferentiation into granulosa-like cells and the formation of TGCTs. To test this hypothesis, we generated mice harboring constitutive activation of TGFBR1 in Sertoli cells using anti-Mullerian hormone (Amh)-Cre. Disorganized seminiferous tubules and tumor nodules were found in TGFBR1CA; Amh-Cre mice. A histological analysis showed that Sertoli cell-specific activation of TGFBR1 led to the development of neoplasms resembling granulosa cell tumors, which derailed spermatogenesis. Moreover, TGCTs expressed granulosa cell markers including FOXL2, FOXO1, and INHA. Using a dual fluorescence reporter line, the membrane-targeted tdTomato (mT)/membrane-targeted EGFP (mG) mouse, we provided evidence that Sertoli cells transdifferentiated toward a granulosa cell fate during tumorigenesis. Thus, our findings indicate that Sertoli cell-specific activation of TGFBR1 leads to the formation of TGCTs, supporting a key contribution of Sertoli cell reprogramming to the development of this testicular malignancy in our model.

Simultaneously targeting extracellular vesicle trafficking and TGF-ß receptor kinase activity blocks signaling hyperactivation and metastasis.

Metastasis is the leading cause of cancer-related deaths. Transforming growth factor beta (TGF-ß) signaling drives metastasis and is strongly enhanced during cancer progression. Yet, the use of on-target TGF-ß signaling inhibitors in the treatment of cancer patients remains unsuccessful, highlighting a gap in the understanding of TGF-ß biology that limits the establishment of efficient anti-metastatic therapies. Here, we show that TGF-ß signaling hyperactivation in breast cancer cells is required for metastasis and relies on increased small extracellular vesicle (sEV) secretion. Demonstrating sEV's unique role, TGF-ß signaling levels induced by sEVs exceed the activity of matching concentrations of soluble ligand TGF-ß. Further, genetic disruption of sEV secretion in highly-metastatic breast cancer cells impairs cancer cell aggressiveness by reducing TGF-ß signaling to nearly-normal levels. Otherwise, TGF-ß signaling activity in non-invasive breast cancer cells is inherently low, but can be amplified by sEVs, enabling invasion and metastasis of poorly-metastatic breast cancer cells. Underscoring the translational potential of inhibiting sEV trafficking in advanced breast cancers, treatment with dimethyl amiloride (DMA) decreases sEV secretion, TGF-ß signaling activity, and breast cancer progression in vivo. Targeting both the sEV trafficking and TGF-ß signaling by combining DMA and SB431542 at suboptimal doses potentiated this effect, normalizing the TGF-ß signaling in primary tumors to potently reduce circulating tumor cells, metastasis, and tumor self-seeding. Collectively, this study establishes sEVs as critical elements in TGF-ß biology, demonstrating the feasibility of inhibiting sEV trafficking as a new therapeutic approach to impair metastasis by normalizing TGF-ß signaling levels in breast cancer cells.

MicroRNA-499-5p inhibits transforming growth factor-ß1-induced Smad2 signaling pathway and suppresses fibroblast proliferation and collagen synthesis in rat by targeting TGFß-R1.

BACKGROUND: Artial fibrosis has been recognized as a typical pathological change in atrial fibrillation. Although present evidence suggests that microRNA-499-5p (miR-499-5p) plays an important role in the development of atrial fibrosis, the specific mechanism is not fully understood. Therefore, this study attempted to assess the influence of miR-499-5p on atrial fibroblasts and explore the potential molecular mechanism. METHODS: Atrial fibroblasts from sprague dawley rat were respectively transfected with miR-499-5p mimic, miR-499-5p negative control and miR-499-5p inhibitor, atrial fibroblasts without any treatment were also established. Cell counting kit-8 assay and transwell assay were used to detect the proliferation and migration of atrial fibroblasts in each group. Expressions of miR-499-5p, TGF-ß1, smad2, α-SMA, collagen-I and TGFß-R1 in mRNA and protein level were subsequently detected via quantitative real-time polymerase chain reaction and western blot. Furthermore, the prediction of the binding sites of miR-499-5p and TGFß-R1 was performed via the bioinformatics online software TargetScan and verified by dual luciferase reporter. RESULTS: By utilizing miR-499-5p-transfected atrial fibroblasts model, expression of miR-499-5p in the miR-499-5p mimic group was upregulated, while it was downregulated in the miR-499-5p inhibitors group. Upregulated miR-499-5p expression led to to a significant decrease in the proliferative and migratory ability of cultured atrial fibroblasts, while downregulated miR-499-5p expression led to a significant increase in the proliferative and migratory ability of cultured atrial fibroblasts. Additionally, upregulated miR-499-5p expression made a significant rise in TGF-ß1-induced mRNA and protein expression of TGF-ß1, TGFß-R1, smad2, α-SMA and collagen-I in atrial fibroblasts. Furthermore, results from the dual luciferase reporter conformed that miR-499-5p may repress TGFß-R1 by binding the 3'UTR of TGFß-R1 directly. CONCLUSIONS: miR-499-5p is able to inhibit the activation of transforming growth factor ß-induced Smad2 signaling and eventually suppressed the proliferation, migration and invasion of atrial fibroblasts and collagen synthesis by targeting TGFß-R1.

Design and characterization of a novel tumor-homing cell-penetrating peptide for drug delivery in TGFBR3 high-expressing tumors.

Targeted therapy has attracted more and more attention in cancer treatment in recent years. However, due to the diversity of tumor types and the mutation of target sites on the tumor surface, some existing targets are no longer suitable for tumor therapy. In addition, the long-term administration of a single targeted drug can also lead to drug resistance and attenuate drug potency, so it is important to develop new targets for tumor therapy. The expression of Type III transforming growth factor ß receptor (TGFBR3) is upregulated in colon, breast, and prostate cancer cells, and plays an important role in the occurrence and development of these cancers, so TGFBR3 may be developed as a novel target for tumor therapy, but so far there is no report on this research. In this study, the structure of bone morphogenetic protein 4 (BMP4), one of the ligands of TGFBR3 was analyzed through the docking analysis with TGFBR3 and sequence charge characteristic analysis, and a functional tumor-targeting penetrating peptide T3BP was identified. The results of fluorescent labeling experiments showed that T3BP could target and efficiently enter tumor cells with high expression of TGFBR3, especially A549 cells. When the expression of TGFBR3 on the surface of tumor cells (HeLa) was knocked down by RNA interference, the high delivery efficiency of T3BP was correspondingly reduced by 40%, indicating that the delivery was TGFBR3-dependent. Trichosanthin (TCS, a plant-derived ribosome inactivating protein) fused with T3BP can enhance the inhibitory activity of the fusion protein on A549 cells by more than 200 times that of TCS alone. These results indicated that T3BP, as a novel targeting peptide that can efficiently bind TGFBR3 and be used for targeted therapy of tumors with high expression of TGFBR3. This study enriches the supply of tumor-targeting peptides and provides a new potential application option for the treatment of tumors with high expression of TGFBR3.

Transforming Growth Factor ß1 Ameliorates Microglial Activation in Perioperative Neurocognitive Disorders.

Perioperative neurocognitive disorder (PND) is a common complication of surgery and anesthesia, especially among older patients. Microglial activation plays a crucial role in the occurrence and development of PND and transforming growth factor beta 1 (TGF-ß1) can regulate microglial homeostasis. In the present study, abdominal surgery was performed on 12-14 months-old C57BL/6 mice to establish a PND model. The expression of TGF-ß1, TGF-ß receptor 1, TGF-ß receptor 2, and phosphor-smad2/smad3 (psmad2/smad3) was assessed after anesthesia and surgery. Additionally, we examined changes in microglial activation, morphology, and polarization, as well as neuroinflammation and dendritic spine density in the hippocampus. Behavioral tests, including the Morris water maze and open field tests, were used to examine cognitive function, exploratory locomotion, and emotions. We observed decreased TGF-ß1 expression after surgery and anesthesia. Intranasally administered exogenous TGF-ß1 increased psmad2/smad3 colocalization with microglia positive for ionized calcium-binding adaptor molecule 1. TGF-ß1 treatment attenuated microglial activation, reduced microglial phagocytosis, and reduced surgery- and anesthesia-induced changes in microglial morphology. Compared with the surgery group, TGF-ß1 treatment decreased M1 microglial polarization and increased M2 microglial polarization. Additionally, surgery- and anesthesia-induced increase in interleukin 1 beta and tumor necrosis factor-alpha levels was ameliorated by TGF-ß1 treatment at postoperative day 3. TGF-ß1 also ameliorated cognitive function after surgery and anesthesia as well as rescue dendritic spine loss. In conclusion, surgery and anesthesia induced decrease in TGF-ß1 levels in older mice, which may contribute to PND development; however, TGF-ß1 ameliorated microglial activation and cognitive dysfunction in PND mice.

CUL4B enhances the malignant phenotype of esophageal squamous cell carcinoma by suppressing TGFBR3 expression.

Cullin 4B (CUL4B), which acts as a scaffold protein in CUL4B-RING ubiquitin ligase complexes (CRL4B), is frequently overexpressed in cancer and represses tumor suppressors through epigenetic mechanisms. However, the expression and function of CUL4B in esophageal squamous cell carcinoma (ESCC) have not been well illustrated. In this study, we show that upregulation of CUL4B in ESCC cells enhances proliferation, invasion and cisplatin (CDDP)-resistance, while knockdown of CUL4B significantly represses the malignant activities. Mechanistically, we demonstrate that CUL4B promotes proliferation and migration of ESCC cells through inhibiting expression of transforming growth factor beta receptor III (TGFBR3). CRL4B complex binds to the promoter of TGFBR3, and represses its transcription by catalyzing monoubiquitination at H2AK119 and coordinating with PRC2 and HDAC complexes. Taken together, our findings establish a critical role for the CUL4B/TGFBR3 axis in the regulation of ESCC malignancy.