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Background: Ulcerative colitis (UC) is a chronic inflammatory condition affecting the colon, with a global incidence that is rising. Despite the increasing prevalence, effective treatment options for UC remain limited., Methods: We utilized an NF-κB promoter dual fluorescence reporter system to screen for compounds that could inhibit p65 and IκBα phosphorylation. The anti-hypertension drug lacidipine was identified as a candidate. Its efficacy was further evaluated in a murine model of dextran sulfate sodium (DSS)-induced colitis. The analysis included the assessment of colon lesions, inflammation markers, and signal pathway activation, with a focus on NF-κB and Notch signaling., Results: Lacidipine effectively inhibited p65 and IκBα phosphorylation in the reporter system. In the DSS-induced colitis murine model, lacidipine treatment led to a reduction in colon lesions and inflammatory markers. Target analysis showed significant enrichment of the Notch signaling pathway. Additionally, lacidipine inhibited both NF-κB and Notch activation in DSS-stimulated colons., Conclusion: Lacidipine demonstrated a protective effect in UC, reducing inflammation and modulating key signaling pathways. These findings suggest that lacidipine could be a promising candidate for the treatment of UC., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Abstract: We report on the antileukemic activity of homoharringtonine (HHT) in T-cell acute lymphoblastic leukemia (T-ALL). We showed that HHT inhibited the NOTCH/MYC pathway and induced significantly longer survival in mouse and patient-derived T-ALL xenograft models, supporting HHT as a promising agent for T-ALL., (© 2024 American Society of Hematology. Published by Elsevier Inc. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Aberration of Notch signaling is one of the key events involved in the development and progression of head and neck squamous cell carcinoma (HNSCC). The Notch pathway controls the tissue-specific differentiation of normal squamous epithelial cells and is frequently altered in squamous carcinomas, thus affecting their proliferation, growth, survival, and chemosensitivity or resistance against anti-cancer agents. In this study, we show that the use of novel, small-molecule inhibitors of Notch signaling, such as FLI-06, can have a beneficial effect on increasing the chemosensitivity of HNSCC to taxane-based chemotherapy. Inhibition of Notch signaling by FLI-06 alone virtually blocks the proliferation and growth of HNSCC cells in both 2D and 3D cultures and the zebrafish model, which is accompanied by down-regulation of key Notch target genes and proteins. Mechanistically, FLI-06 treatment causes cell cycle arrest in the G 1 -phase and induction of apoptosis in HNSCC, which is accompanied by increased c-Jun S63 phosphorylation. Combining FLI-06 with Docetaxel shows a synergistic effect and partially blocks the cell growth of aggressive HNSCC cells via enhanced apoptosis and modification of c-Jun S243 phosphorylation via GSK-3β inhibition. In conclusion, inhibition of Notch signaling in HNSCC cells that retain active Notch signaling significantly supports taxane-based anticancer activities via modulation of both the GSK-3β and the c-Jun., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

With the increased prevalence of nonalcoholic steatohepatitis (NASH) in the world, effective pharmacotherapy in clinical practice is still lacking. Previous studies have shown that dibenzazepine (DBZ), a Notch inhibitor, could alleviate NASH development in a mouse model. However, low bioavailability, poor water solubility, and extrahepatic side effects restrict its clinical application. To overcome these barriers, we developed a reactive oxygen species (ROS)-sensitive nanoparticle based on the conjugation of bilirubin to poly(ethylene glycol) (PEG) chains, taking into account the overaccumulation of hepatic ROS in the pathologic state of nonalcoholic steatohepatitis (NASH). The PEGylated bilirubin can self-assemble into nanoparticles in an aqueous solution and encapsulate insoluble DBZ into its hydrophobic cavity. DBZ nanoparticles (DBZ Nps) had good stability, rapidly released DBZ in response to H 2 O 2 , and effectively scavenged intracellular ROS of hepatocytes. After systemic administration, DBZ Nps could accumulate in the liver of the NASH mice, extend persistence in circulation, and improve the bioavailability of DBZ. Furthermore, DBZ Nps significantly improved glucose intolerance, relieved hepatic lipid accumulation and inflammation, and ameliorated NASH-induced liver fibrosis. Additionally, DBZ Nps had no significant extrahepatic side effects. Taken together, our results highlight the potential of the ROS-sensitive DBZ nanoparticle as a promising therapeutic strategy for NASH.

Objective: Lifelong insulin replacement remains the mainstay of type 1 diabetes treatment. Genetic FoxO1 ablation promotes enteroendocrine cell (EECs) conversion into glucose-responsive β-like cells. Here, we tested whether chemical FoxO1 inhibitors can generate β-like gut cells., Methods: We used Ngn3-or Villin-driven FoxO1 ablation to capture the distinctive developmental effects of FoxO1 on EEC pool. We combined FoxO1 ablation with Notch inhibition to enhance the expansion of EEC pool. We tested the ability of an orally available small molecule of FoxO1 inhibitor, Cpd10, to phenocopy genetic ablation of FoxO1. We evaluated the therapeutic impact of genetic ablation or chemical inhibition of FoxO1 on insulin-deficient diabetes in Ins2 Akita/+ mice., Results: Pan-intestinal epithelial FoxO1 ablation expanded the EEC pool, induced β-like cells, and improved glucose tolerance in Ins2 Akita/+ mice. This genetic effect was phenocopied by Cpd10. Cpd10 induced β-like cells that released insulin in response to glucose in gut organoids, and this effect was enhanced by the Notch inhibitor, DBZ. In Ins2 Akita/+ mice, a five-day course of either Cpd10 or DBZ induced intestinal insulin-immunoreactive β-like cells, lowered glycemia, and increased plasma insulin levels without apparent adverse effects., Conclusion: These results provide proof of principle of gut cell conversion into β-like cells by a small molecule FoxO1 inhibitor, paving the way for clinical applications., (Copyright © 2022 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Background and Aims: The mechanisms involved in liver regeneration after partial hepatectomy (pHx) are complicated. Cellular senescence, once linked to aging, plays a pivotal role in wound repair. However, the regulatory effects of cellular senescence on liver regeneration have not been fully elucidated., Approach and Results: Mice subjected to pHx were analyzed 14 days after surgery. The incomplete remodeling of liver sinusoids affected shear stress-induced endothelial nitric oxide synthase (eNOS) signaling on day 14, resulting in the accumulation of senescent LSECs. Removing macrophages to augment LSEC senescence led to a malfunction of the regenerating liver. A dynamic fluctuation in Notch activity accompanied senescent LSEC accumulation during liver regeneration. Endothelial Notch activation by using Cdh5-CreERT NIC eCA mice triggered LSEC senescence and senescence-associated secretory phenotype, which disrupted liver regeneration. Blocking the Notch by γ-secretase inhibitor (GSI) diminished senescence and promoted LSEC expansion. Mechanically, Notch-hairy and enhancer of split 1 signaling inhibited sirtuin 1 (Sirt1) transcription by binding to its promoter region. Activation of Sirt1 by SRT1720 neutralized the up-regulation of P53, P21, and P16 caused by Notch activation and eliminated Notch-driven LSEC senescence. Finally, Sirt1 activator promoted liver regeneration by abrogating LSEC senescence and improving sinusoid remodeling., Conclusions: Shear stress-induced LSEC senescence driven by Notch interferes with liver regeneration after pHx. Sirt1 inhibition accelerates liver regeneration by abrogating Notch-driven senescence, providing a potential opportunity to target senescent cells and facilitate liver repair after injury., (© 2021 American Association for the Study of Liver Diseases.)

Although it is thought that there is a close relationship between Notch signal and preterm birth, the functioning of this mechanism in the cervix is unknown. The efficacy of surfactants and prostaglandin inhibitors in preterm labor is also still unclear. In this study, 48 female CD-1 mice were distributed to pregnant control (PC), Sham, PBS, indomethacin (2 mg/kg; intraperitoneally), lipopolysaccharides (LPS) (25 μg/100 μl; intrauterine), LPS + IND, and Surfactant Protein A Block (SP-A Block: SP-A B; the anti-SP-A antibody was applied 20 µg/100μl; intrauterine) groups. Tissues were examined by immunohistochemistry, immunofluorescence, and Western blot analysis. LPS administration increased the expression of N1 Dll-1 and Jagged-2 (Jag-2). Although Toll-like receptor (Tlr)-2 significantly increased in the LPS-treated and SP-A-blocked groups, Tlr-4 significantly increased only in the LPS-exposed groups. It was observed that Jag-2 is specifically expressed by mast cells. Overall, this experimental model shows that some protein responses increase throughout the uterus, starting at a specific point on the cervix epithelium. Surfactant Protein A, which we observed to be significantly reduced by LPS, may be associated with the regulation of the epithelial response, especially during preterm delivery due to infection. On the contrary, prostaglandin inhibitors can be considered an option to delay infection-related preterm labor with their dose-dependent effects. Finally, the link between mast cells and Jag-2 could potentially be a control switch for preterm birth.

Chronic lymphocytic leukemia (CLL) is characterized by disordered DNA methylation, suggesting these epigenetic changes might play a critical role in disease onset and progression. The methyltransferase DNMT3A is a key regulator of DNA methylation. Although DNMT3A somatic mutations in CLL are rare, we found that low DNMT3A expression is associated with more aggressive disease. A conditional knockout mouse model showed that homozygous depletion of Dnmt3a from B cells results in the development of CLL with 100% penetrance at a median age of onset of 5.3 months, and heterozygous Dnmt3a depletion yields a disease penetrance of 89% with a median onset at 18.5 months, confirming its role as a haploinsufficient tumor suppressor. B1a cells were confirmed as the cell of origin of disease in this model, and Dnmt3a depletion resulted in focal hypomethylation and activation of Notch and Myc signaling. Amplification of chromosome 15 containing the Myc gene was detected in all CLL mice tested, and infiltration of high- Myc -expressing CLL cells in the spleen was observed. Notably, hyperactivation of Notch and Myc signaling was exclusively observed in the Dnmt3a CLL mice, but not in three other CLL mouse models tested ( Sf3b1-Atm , Ikzf3 , and MDR ), and Dnmt3a -depleted CLL were sensitive to pharmacologic inhibition of Notch signaling in vitro and in vivo . Consistent with these findings, human CLL samples with lower DNMT3A expression were more sensitive to Notch inhibition than those with higher DNMT3A expression. Altogether, these results suggest that Dnmt3a depletion induces CLL that is highly dependent on activation of Notch and Myc signaling. SIGNIFICANCE: Loss of DNMT3A expression is a driving event in CLL and is associated with aggressive disease, activation of Notch and Myc signaling, and enhanced sensitivity to Notch inhibition., (©2021 American Association for Cancer Research.)

Necrotizing enterocolitis (NEC) is an often lethal, inflammatory disease of the preterm intestine. The underdeveloped immune system plays an important role; however, the initial trigger for NEC development is likely a damaged intestinal epithelial layer. We hypothesize that due to incomplete maturation of different epithelial cell lineages, nutrients and bacteria are able to damage the epithelial cells and cause the (immature) inflammatory response, food intolerance and malabsorption seen in NEC. Intestinal organoid research has shown that maturation of intestinal epithelial cell lineages is orchestrated by two key signaling pathways: Wnt and Notch. In NEC, these pathways are dysregulated by hyperactivation of Toll-like-receptor-4. Breastfeeding decreases the risk of developing NEC compared to formula milk. Here, we review the intricate link between breast milk components, Wnt and Notch signaling and intestinal epithelial maturation. We argue that (nutritional) interventions regulating these pathways may decrease the risk of NEC development in preterm infants., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Notch signaling involves cell to cell contact. It is an ancient signaling mechanism that is conserved throughout the animal kingdom. The basic function of Notch signaling is to decide cell fate and execute asymmetrical division. Notch signaling is indispensable for embryo growth. Aberrant Notch signaling involves in cancer progression by altering cell proliferation rate, tumor micro-environment, stem cell activities. The role of Notch signaling in cancer progression is context-dependent. In breast cancer and T cell lymphoma Notch signaling is highly active, whereas in squamous cell carcinoma (SCC) as oral and skin cancer, the signaling is suppressed. It is believed that in SCC, Notch-mediated tumor growth is due to the cell non-autonomous function. Oral cancer is the 6th most risky cancer worldwide. In many patients, oral cancer is preceded by pre-cancer conditions. In this review, we have summarized the research knowledge related to the role of Notch signaling in oral cancer and pre-cancer conditions and the therapeutic options available targeting different components of Notch pathways., (© 2021. Springer Science+Business Media, LLC, part of Springer Nature.)

Air-liquid interface (ALI) cultures are frequently used in lung research but require substantial cell numbers that cannot readily be obtained from patients. We explored whether organoid expansion [three-dimensional (3D)] can be used to establish ALI cultures from clinical samples with low epithelial cell numbers. Airway epithelial cells were obtained from tracheal aspirates (TA) from preterm newborns and from bronchoalveolar lavage (BAL) or bronchial tissue (BT) from adults. TA and BAL cells were 3D-expanded, whereas cells from BT were expanded in 3D and 2D. Following expansion, cells were cultured at ALI to induce differentiation. The impact of cell origin and 2D or 3D expansion was assessed with respect to 1 ) cellular composition, 2 ) response to cigarette smoke exposure, and 3 ) effect of Notch inhibition or IL-13 stimulation on cellular differentiation. We established well-differentiated ALI cultures from all samples. Cellular compositions (basal, ciliated, and goblet cells) were comparable. All 3D-expanded cultures showed a similar stress response following cigarette smoke exposure but differed from the 2D-expanded cultures. Higher peak levels of antioxidant genes HMOX1 and NQO1 and a more rapid return to baseline, and a lower unfolded protein response was observed after cigarette smoke exposure in 3D-derived cultures compared to 2D-derived cultures. In addition, TA- and BAL-derived cultures were less sensitive to modulation by DAPT or IL-13 than BT-derived cultures. Organoid-based expansion of clinical samples with low cell numbers, such as TA from preterm newborns is a valid method and tool to establish ALI cultures.

Systemic inhibition of Notch with γ-secretase inhibitors (GSI) decreases multiple myeloma tumor growth, but the clinical use of GSI is limited due to its severe gastrointestinal toxicity. In this study, we generated a GSI Notch inhibitor specifically directed to the bone (BT-GSI). BT-GSI administration decreased Notch target gene expression in the bone marrow, but it did not alter Notch signaling in intestinal tissue or induce gut toxicity. In mice with established human or murine multiple myeloma, treatment with BT-GSI decreased tumor burden and prevented the progression of multiple myeloma-induced osteolytic disease by inhibiting bone resorption more effectively than unconjugated GSI at equimolar doses. These findings show that BT-GSI has dual anti-myeloma and anti-resorptive properties, supporting the therapeutic approach of bone-targeted Notch inhibition for the treatment of multiple myeloma and associated bone disease. SIGNIFICANCE: Development of a bone-targeted Notch inhibitor reduces multiple myeloma growth and mitigates cancer-induced bone destruction without inducing the gastrointestinal toxicity typically associated with inhibition of Notch., (©2021 American Association for Cancer Research.)

Some individuals develop prediabetes and/or diabetes following acute pancreatitis (AP). AP-induced beta-cell injury and the limited regenerative capacity of beta cells might account for pancreatic endocrine insufficiency. Previously, we found that only a few pancreatic cytokeratin 5 positive (Krt5 + ) cells differentiated into beta cells in the murine AP model, which was insufficient to maintain glucose homeostasis. Notch signaling determines pancreatic progenitor differentiation in pancreas development. This study aimed to examine whether Notch signaling inhibition could promote pancreatic Krt5 + cell differentiation into beta cells and improve glucose homeostasis following AP. Pancreatic tissues from patients with acute necrotizing pancreatitis (ANP) were used to evaluate beta-cell injury, Krt5 + cell activation and differentiation, and Notch activity. The murine AP model was induced by cerulein, and the effect of Notch inhibition on Krt5 + cell differentiation was evaluated both in vivo and in vitro. The results demonstrated beta-cell loss in ANP patients and AP mice. Krt5 + cells were activated in ANP pancreases along with persistently elevated Notch activity, which resulted in the formation of massive duct-like structures. AP mice that received Notch inhibitor showed that impaired glucose tolerance was reversed 7 and 15 days following AP, and increased numbers of newborn small islets due to increased differentiation of Krt5 + cells to beta cells to some extent. In addition, Krt5 + cells isolated from AP mice showed increased differentiation to beta cells by Notch inhibition. Collectively, these findings suggest that beta-cell loss contributes to pancreatic endocrine insufficiency following AP, and inhibition of Notch activity promotes pancreatic Krt5 + cell differentiation to beta cells and improves glucose homeostasis. The findings from this study may shed light on the potential treatment of prediabetes/diabetes following AP., (© 2021. The Author(s).)

Functional loss of myosin Vb (MYO5B) induces a variety of deficits in intestinal epithelial cell function and causes a congenital diarrheal disorder, microvillus inclusion disease (MVID). The impact of MYO5B loss on differentiated cell lineage choice has not been investigated. We quantified the populations of differentiated epithelial cells in tamoxifen-induced, epithelial cell-specific MYO5B-knockout (VilCreERT2 Myo5bfl/fl) mice utilizing digital image analysis. Consistent with our RNA-sequencing data, MYO5B loss induced a reduction in tuft cells in vivo and in organoid cultures. Paneth cells were significantly increased by MYO5B deficiency along with expansion of the progenitor cell zone. We further investigated the effect of lysophosphatidic acid (LPA) signaling on epithelial cell differentiation. Intraperitoneal LPA significantly increased tuft cell populations in both control and MYO5B-knockout mice. Transcripts for Wnt ligands were significantly downregulated by MYO5B loss in intestinal epithelial cells, whereas Notch signaling molecules were unchanged. Additionally, treatment with the Notch inhibitor dibenzazepine (DBZ) restored the populations of secretory cells, suggesting that the Notch pathway is maintained in MYO5B-deficient intestine. MYO5B loss likely impairs progenitor cell differentiation in the small intestine in vivo and in vitro, partially mediated by Wnt/Notch imbalance. Notch inhibition and/or LPA treatment may represent an effective therapeutic approach for treatment of MVID.

Cell signaling is considered a part of a network for communication that regulates basic cellular activities. The ability of cells to communicate correctly to the surrounding environment has an important role in development, tissue repair, and immunity as well as normal tissue homeostasis. Dysregulated activation and crosstalk between many intracellular signaling pathways are implicated in the pathogenesis of rheumatoid arthritis (RA), such as the Janus Kinase/signal transducers and activators of transcription (JAK/STAT), Toll-like receptor/nuclear factor kappa B (TLR/NF-κB), phosphatidylinositide-3Kinase/protein kinase B/mammalian target of rapamycin (PI-3K/AKT/mTOR), the stress activated protein kinase/mitogen-activated protein kinase (SAPK/MAPK), and spleen tyrosine kinase (SYK) pathways. Other interrelated pathways that can be targeted to halt the inflammatory status in the disease are purinergic 2X7 receptor (P2X7R)/nucleotide binding oligomerization domain-like receptor family pyrin domain containing 3 or inflammasome (NLRP-3)/NF-κB and Notch pathways. In this review, we will show the orchestrated modulation in the pathogenesis of RA via the crossregulation between dysregulated signaling pathways which can mediate a sustained loop of activation for these signaling pathways as well as aggrevate the inflammatory condition. Also, this review will highlight many targets that can be useful in the development of more effective therapeutic options., (© 2021. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Interleukin (IL)-17 is a prominent cytokine that promotes pancreatic intraepithelial neoplasia (PanIN) and pancreatic ductal adenocarcinoma (PDAC) and is associated with the oncogenic pathways in tumor progression. However, the mechanism and therapeutic value of the IL-17 axis remain unclear. In this study, we verified the activation of the IL-17 and Notch pathways in PanIN/PDAC via complementary approaches and validated their pro-tumor effects on tumor progression. Additionally, we found a positive correlation between IL-17 and Notch; the IL-17 axis can upregulate Notch activity via the canonical NF-κB pathway in vitro, thus synergistically promoting PanIN/PDAC. Furthermore, we observed that the co-inhibition of IL-17 and the Notch pathway can enhance the therapeutic effect by restricting tumor growth in vivo. Our study highlights the synergistic effect of the IL-17 axis and Notch pathway in promoting PanIN/PDAC and further suggests that IL-17-Notch co-inhibition is a novel therapeutic strategy with superior potential in treating PDAC., (Copyright © 2021. Published by Elsevier B.V.)

The etiology for liver cancer has been clearly defined. Unfortunately, therapeutic approaches for liver cancer are rather limited, and liver cancer is insensitive to chemotherapy and radiotherapy. Traditional Chinese medicine (TCM) has become a promising strategy for cancer treatment as TCM elicits broad spectrum anticancer activity. In the present study, we evaluated the anticancer efficacy of AB4, an extract from the medical herb Pulsatilla chinensis (Bunge) Regel, in liver cancer in vitro and in vivo . We found that AB4 readily dose‑ and time‑dependently inhibited liver cancer HepG2 and Huh‑7 cell proliferation and colony formation. Western blot and flow cytometry analyses suggested that AB4 treatment induced liver cancer cell apoptosis. Moreover, these findings could be readily recaptured in vivo , in which the AB4 regimen resulted in tumor suppression and cancer cell apoptosis in xenograft tumor‑bearing nude mice. Importantly, we noted that treatment with a Notch signaling inhibitor DAPT produced very similar anticancer efficacy in both HepG2 and Huh‑7 cell lines, and administration of DAPT also efficiently suppressed HepG2 xenograft outgrowth. To this end, we anticipated that AB4 and DAPT may act on the same signaling pathway, probably through inhibition of the Notch pathway. Indeed, we found decreased expression of Notch1 protein, as well as downstream targets Hes1 and Hey1, after AB4 treatment. Immunohistochemistry analysis further confirmed the suppression of Notch signaling in HepG2 xenograft‑bearing mice. Taken together, our study highlighted the anticancer efficacy of AB4 in liver cancer. We also provided preliminary data showing Notch as a therapeutic target of AB4. It would be interesting to investigate the anticancer efficacy of AB4 in other types of cancer with elevated Notch activity.

Aberrant NOTCH3 signaling and overexpression is oncogenic, associated with cancer stem cells and drug resistance, yet therapeutic targeting remains elusive. Here, we develop NOTCH3-targeted antibody drug conjugates (NOTCH3-ADCs) by bioconjugation of an auristatin microtubule inhibitor through a protease cleavable linker to two antibodies with differential abilities to inhibit signaling. The signaling inhibitory antibody rapidly induces ligand-independent receptor clustering and internalization through both caveolin and clathrin-mediated pathways. The non-inhibitory antibody also efficiently endocytoses via clathrin without inducing receptor clustering but with slower lysosomal co-localization kinetics. In addition, DLL4 ligand binding to the NOTCH3 receptor mediates transendocytosis of NOTCH3-ADCs into ligand-expressing cells. NOTCH3-ADCs internalize into receptor and ligand cells independent of signaling and induce cell death in both cell types representing an atypical mechanism of ADC cytotoxicity. Treatment of xenografts with NOTCH3-ADCs leads to sustained tumor regressions, outperforms standard-of-care chemotherapy, and allows targeting of tumors that overexpress NOTCH3 independent of signaling inhibition., Competing Interests: All authors are/were employees and shareholders of Pfizer. US patents associated with the development of NOTCH3-ADCs include 8,828,401 and 9,433,687., (© 2021 The Authors.)

Although advances in osteosarcoma treatment have been made in recent decades, the survival rate for patients suffering from metastatic disease, especially lung metastasis, remains disappointing. Previous studies have confirmed that epithelial-to-mesenchymal transition (EMT) is associated with tumor metastasis, and several studies have suggested that osteosarcoma cells also exhibit EMT-like characteristics. In addition, Notch signaling is known to be related to the development and progression of human malignancies, including osteosarcoma. However, whether chemotherapy affects the EMT-like events and whether these events are medicated by Notch signaling remain to be elucidated. To address these issues, in the current work, osteosarcoma 143B cells were exposed to sublethal concentrations of the first-line chemotherapeutic agent cisplatin (DDP), which promoted cell migration, in vitro invasion, and in vivo lung metastasis. Furthermore, low concentrations of DDP upregulated mesenchymal phenotype-related genes and proteins and promoted EMT-like properties in osteosarcoma cells. In addition, low concentrations of DDP could activate the Notch receptor and its target genes. Finally, combined treatment of DDP with the Notch signaling pathway inhibitor DAPT, which can effectively downregulate mesenchymal phenotype-related genes and proteins, inhibited cell migration and invasion in vitro, and it decreased pulmonary metastatic nodules in vivo. The results of the current study supported the idea that low concentrations of DDP could induce EMT-like characteristics in osteosarcoma cells and could promote cell mobility in vitro, as well as pulmonary metastasis in vivo. Importantly, however, these biological processes are mediated by the Notch signaling pathway. Blocking the Notch signaling pathway can effectively attenuate the osteosarcoma EMT-like phenotype and its associated migration, invasion, and metastasis., (Copyright © 2021. Published by Elsevier B.V.)

The Notch signaling pathway regulates developmental cell-fate decisions and has recently also been linked to inflammatory diseases. Although therapies targeting Notch signaling in inflammation in theory are attractive, their design and implementation have proven difficult, at least partly due to the broad involvement of Notch signaling in regenerative and homeostatic processes. In this review, we summarize the supporting role of Notch signaling in various inflammation-driven diseases, and highlight efforts to intervene with this pathway by targeting Notch ligands and/or receptors with distinct therapeutic strategies, including antibody designs. We discuss this in light of lessons learned from Notch targeting in cancer treatment. Finally, we elaborate on the impact of individual Notch members in inflammation, which may lay the foundation for development of therapeutic strategies in chronic inflammatory diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Christopoulos, Gjølberg, Krüger, Haraldsen, Andersen and Sundlisæter.)

Background This phase 1, single-center, nonrandomized, single-arm, open-label, dose-escalation study, evaluated the tolerability of crenigacestat, a γ-secretase inhibitor as an oral Notch inhibitor in Japanese patients with advanced solid tumors. Methods The study consisted of 2 dose levels of crenigacestat (25 mg and 50 mg), administered orally 3 times per week (TIW) over a 28-day cycle until disease progression, development of unacceptable toxicity, or any other discontinuation criteria were met. The primary objective was to evaluate the tolerability and determine the recommended dose of crenigacestat for Japanese patients. Secondary objectives were to characterize the safety and toxicity, the pharmacokinetic parameters, and to document any antitumor activity of crenigacestat. Results Eleven Japanese patients with advanced solid tumors were enrolled; 4 patients (median age of 64 years) received 25 mg of crenigacestat, and 7 patients (median age of 72 years) received 50 mg of crenigacestat. Median treatment duration was 8 weeks in the 25-mg treatment arm and 4 weeks in the 50-mg treatment arm. There were no dose-limiting toxicities or dose-limiting equivalent toxicities observed. None of the patients had a complete or partial response to the treatment. One patient (14.3%) with a desmoid tumor in the 50-mg treatment arm showed tumor size shrinkage of 22.4% and had stable disease for 22.5 months. Frequent (>14%) treatment-related-adverse events in both treatment arms included diarrhea, malaise, and vomiting. Conclusions Crenigacestat was tolerated in Japanese patients but with limited clinical activity. The recommended crenigacestat dose in Japanese patients is 50 mg TIW.Trial registration: NCT02836600 ( ClinicalTrials.gov ) registered on July 19, 2016.

Notch signaling inhibitors with the potential of immune suppressor production by pathogenic bacteria for easy host infection were searched from extracts of Nocardia sp. Nocobactin NA-a (compound 1) and nocobactin NA-b (compound 2), which have been suggested as pathogenesis factors, were isolated from N. farcinica IFM 11523 isolated from the sputum of a Japanese patient with chronic bronchitis. Compounds 1 and 2 showed Notch inhibitory activities with IC 50 values of 12.4 and 17.6 μM, respectively. Compound 1 and 2 decreased of Notch1 protein, Notch intracellular domain, and hairy and enhancer of split 1, which is a Notch signaling target protein. In addition, compounds 1 and 2 showed cytotoxicity against mouse macrophage-like cell line RAW264.7 with IC 50 values of 18.9 and 21.1 μM, respectively. These results suggested that the Notch inhibitors production by pathogenic bacteria may increase pathogen infectivity.

The dysregulation of Notch signaling is associated with a wide variety of different human cancers. Notch signaling activation mostly relies on the activity of the γ-secretase enzyme that cleaves the Notch receptors and releases the active intracellular domain. It is well-documented that γ-secretase inhibitors (GSIs) block the Notch activity, mainly by inhibiting the oncogenic activity of this pathway. To date, several GSIs have been introduced clinically for the treatment of various diseases, such as Alzheimer's disease and various cancers, and their impacts on Notch inhibition have been found to be promising. Therefore, GSIs are of great interest for cancer therapy. The objective of this review is to provide a systematic review of in vitro and in vivo studies for investigating the effect of GSIs on various cancer stem cells (CSCs), mainly by modulation of the Notch signaling pathway. Various scholarly electronic databases were searched and relevant studies published in the English language were collected up to February 2020. Herein, we conclude that GSIs can be potential candidates for CSC-targeting therapy. The outcome of our study also indicates that GSIs in combination with anticancer drugs have a greater inhibitory effect on CSCs.

Mucoepidermoid carcinoma (MEC) is the most common type of salivary gland cancers and patients with advanced, metastatic, and recurrent MECs have limited therapeutic options and poor treatment outcomes. MEC is commonly associated with a chromosomal translocation t(11;19) (q14-21;p12-13) that encodes the CRTC1-MAML2 oncogenic fusion. The CRTC1-MAML2 fusion is required for MEC growth in part through inducing autocrine AREG-EGFR signaling. Growing evidence suggests that MEC malignancy is maintained by cancer stem-like cells. In this study, we aimed to determine critical signaling for maintaining MEC stem-like cells and the effect of combined targeting of stem cell signaling and CRTC1-MAML2-induced EGFR signaling on blocking MEC growth. First, we evaluated the significance of Notch signaling in regulating MEC stem-like cells. Aberrantly activated Notch signaling was detected in human fusion-positive MEC cells. The inhibition of Notch signaling with genetic or pharmacological inhibitors reduced oncosphere formation and ALDH-bright population in vitro and blocked the growth of MEC xenografts in vivo. Next, we investigated the effect of co-targeting Notch signaling and EGFR signaling, and observed enhanced inhibition on MEC growth in vivo. Collectively, this study identified a critical role of Notch signaling in maintaining MEC stem-like cells and tumor growth, and revealed a novel approach of co-targeting Notch and EGFR signaling as a potential effective anti-MEC treatment.

P-type ATPase inhibitors are among the most successful and widely prescribed therapeutics in modern pharmacology. Clinical transition has been safely achieved for H + /K + ATPase inhibitors such as omeprazole and Na + /K + -ATPase inhibitors like digoxin. However, this is more challenging for Ca 2+ -ATPase modulators due to the physiological role of Ca 2+ in cardiac dynamics. Over the past two decades, sarco-endoplasmic reticulum Ca 2+ -ATPase (SERCA) modulators have been studied as potential chemotherapy agents because of their Ca 2+ -mediated pan-cancer lethal effects. Instead, recent evidence suggests that SERCA inhibition suppresses oncogenic Notch1 signaling emerging as an alternative to γ-secretase modulators that showed limited clinical activity due to severe side effects. In this review, we focus on how SERCA inhibitors alter Notch1 signaling and show that Notch on-target-mediated antileukemia properties of these molecules can be achieved without causing overt Ca 2+ cellular overload.

We here investigated molecular basis of notch receptor GLP-1 in controlling simulated microgravity stress in Caenorhabditis elegans. glp-1 expression was decreased by simulated microgravity. Meanwhile, glp-1 mutation caused resistance to toxicity of simulated microgravity. GLP-1 acted in germline cells to control toxicity of simulated microgravity. In germline cells, RNAi knockdown of glp-1 increased daf-16 expression. RNAi knockdown of daf-16 suppressed resistance to toxicity of simulated microgravity in glp-1 mutant. In simulated microgravity treated worms, germline RNAi knockdown of glp-1 decreased expressions of daf-28, ins-39, and ins-8 encoding insulin peptides, and resistance to simulated microgravity toxicity could be detected in daf-28(RNAi), ins-39(RNAi), and ins-8(RNAi) worms. In simulated microgravity treated worms, RNAi knockdown of daf-28, ins-39, or ins-8 in germline cells further increased expression and nucleus localization of transcriptional factor DAF-16 in intestinal cells. Therefore, the GLP-1-activated germline-intestine communication of insulin signaling is required for control of simulated microgravity toxicity in C. elegans., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The crosstalk between Notch and MAPK pathway plays a role in MEK inhibitor resistance in BRAF V600E metastatic melanoma (MM) and promotes migration in GNAQ Q209L uveal melanoma (UM) cells. We determined the cytotoxicity of combinatorial inhibition of MEK and Notch by cobimetinib and γ-secretase inhibitor (GSI) nirogacestat, in BRAF V600E and BRAF wt MM and GNAQ Q209L UM cells displaying different Erk1/2 and Notch activation status, with the aim to elucidate the impact of Notch signaling in the response to MEK inhibitor. Overall the combination was synergic in BRAF V600E MM and GNAQ Q209L UM cells and antagonistic in BRAF wt one. Focusing on UM cells, we found that cobimetinib resulted in G0/G1 phase arrest and apoptosis induction, whereas the combination with GSI increased treatment efficacy by inducing a senescent-like state of cells and by blocking migration towards liver cancer cells. Mechanistically, this was reflected in a strong reduction of cyclin D1, in the inactivation of retinoblastoma protein and in the increase of p27 KIP1 expression levels. Of note, each drug alone prevented Notch signaling activation resulting in inhibition of c-jun(Ser63) and Hes-1 expression. The combination achieved the strongest inhibition on Notch signaling and on both c-jun(Ser63) and Erk1/2 activation level. In conclusion we unveiled a coordinate action of MAPK and Notch signaling in promoting proliferation of BRAF V600E MM and GNAQ Q209L UM cells. Remarkably, the simultaneous inhibition of MEK and Notch signaling highlighted a role for the second pathway in protecting cells against senescence in GNAQ Q209L UM cells treated with the MEK inhibitor., (Copyright © 2020 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Notch is a key evolutionary conserved pathway, which has fascinated and engaged the work of investigators in an uncountable number of biological fields, from development of metazoans to immunotherapy for cancer. The study of Notch has greatly contributed to the understanding of cancer biology and a substantial effort has been spent in designing Notch-targeting therapies. Due to its broad involvement in cancer, targeting Notch would allow to virtually modulate any aspect of the disease. However, this means that Notch-based therapies must be highly specific to avoid off-target effects. This review will present the newest mechanistic and therapeutic advances in the Notch field and discuss the promises and challenges of this constantly evolving field.

Melanoma is one of the most serious forms of skin cancer, and its increasing incidence coupled with nonlasting therapeutic options for metastatic disease highlights the need for additional novel approaches for its management. In this study, we determined the potential interactions between polo-like kinase 1 (PLK1, a serine/threonine kinase involved in mitotic regulation) and NOTCH1 (a type I transmembrane protein deciding cell fate during development) in melanoma. Employing an in-house human melanoma tissue microarray (TMA) containing multiple cases of melanomas and benign nevi, coupled with high-throughput, multispectral quantitative fluorescence imaging analysis, we found a positive correlation between PLK1 and NOTCH1 in melanoma. Furthermore, The Cancer Genome Atlas database analysis of patients with melanoma showed an association of higher mRNA levels of PLK1 and NOTCH1 with poor overall, as well as disease-free, survival. Next, utilizing small-molecule inhibitors of PLK1 and NOTCH (BI 6727 and MK-0752, respectively), we found a synergistic antiproliferative response of combined treatment in multiple human melanoma cells. To determine the molecular targets of the overall and synergistic responses of combined PLK1 and NOTCH inhibition, we conducted RNA-sequencing analysis employing a unique regression model with interaction terms. We identified the modulations of several key genes relevant to melanoma progression/metastasis, including MAPK, PI3K , and RAS , as well as some new genes such as Apobec3G, BTK , and FCER1G , which have not been well studied in melanoma. In conclusion, our study demonstrated a synergistic antiproliferative response of concomitant targeting of PLK1 and NOTCH in melanoma, unraveling a potential novel therapeutic approach for detailed preclinical/clinical evaluation., (©2020 American Association for Cancer Research.)

During brain development, progenitor cells need to balanceproliferation and differentiation in order to generate different neurons in the correct numbers and proportions. Currently, the patterns of multipotent progenitor divisions that lead to neurogenic entry and the factors that regulate them are not fully understood. We here use the zebrafish retina to address this gap, exploiting its suitability for quantitative live-imaging. We show that early neurogenic progenitors arise from asymmetric divisions. Notch regulates this asymmetry, as when inhibited, symmetric divisions producing two neurogenic progenitors occur. Surprisingly however, Notch does not act through an apicobasal activity gradient as previously suggested, but through asymmetric inheritance of Sara-positive endosomes. Further, the resulting neurogenic progenitors show cell biological features different from multipotent progenitors, raising the possibility that an intermediate progenitor state exists in the retina. Our study thus reveals new insights into the regulation of proliferative and differentiative events during central nervous system development., Competing Interests: EN, MR, CN No competing interests declared, (© 2020, Nerli et al.)

Liver X receptor (LXR) activation can achieve satisfactory anti-atherosclerotic activity, but can also lead to the development of fatty liver and hypertriglyceridemia. In contrast, Notch inhibition can suppress both atherosclerosis and the hepatic accumulation of lipids. In the present study, we sought to assess whether combining LXR ligand agonists (T317) with Notch receptor inhibitors (DAPT) would lead to enhanced anti-atherosclerotic activity while overcoming the adverse events associated with LXR ligand agonist therapy. The impact of the combined T317 + DAPT therapeutic regimen on atherosclerosis, fatty liver development, and hypertriglyceridemia was assessed using ApoE deficient (ApoE -/- ) mice. The results of this analysis suggested that DAPT was able to improve the anti-atherosclerotic activity of T317 without reducing the stability of lesion plaques while simultaneously reducing blood lipids in treated ApoE -/- mice. This combination T317 + DAPT treatment was also linked with a significant upregulation of ABCA1 and the stimulation of reverse cholesterol transport (RCT), as well as with decreases in the levels of intercellular cell adhesion molecule-1 (ICAM-1) and p-p65, and with altered M1/M2 macrophage proportions within atherosclerotic plaques. Importantly, DAPT was also able to reduce T317-mediated lipid accumulation within the liver owing to its ability to reduce SREBP-1 expression while simultaneously increasing that of Pi-AMPKα and PPARα. Together, our results suggest that administering Notch receptor inhibitors to ApoE -/- mice may be an effective means of enhancing the anti-atherosclerotic activity of LXR ligand agonists while simultaneously limiting associated fatty liver and hypertriglyceridemia development in these animals., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Breast cancer is a heterogeneous disease that can be subdivided into unique molecular subtypes based on protein expression of the Estrogen Receptor, Progesterone Receptor, and/or the Human Epidermal Growth Factor Receptor 2. Therapeutic approaches are designed to inhibit these overexpressed receptors either by endocrine therapy, targeted therapies, or combinations with cytotoxic chemotherapy. However, a significant percentage of breast cancers are inherently resistant or acquire resistance to therapies, and mechanisms that promote resistance remain poorly understood. Notch signaling is an evolutionarily conserved signaling pathway that regulates cell fate, including survival and self-renewal of stem cells, proliferation, or differentiation. Deregulation of Notch signaling promotes resistance to targeted or cytotoxic therapies by enriching of a small population of resistant cells, referred to as breast cancer stem cells, within the bulk tumor; enhancing stem-like features during the process of de-differentiation of tumor cells; or promoting epithelial to mesenchymal transition. Preclinical studies have shown that targeting the Notch pathway can prevent or reverse resistance through reduction or elimination of breast cancer stem cells. However, Notch inhibitors have yet to be clinically approved for the treatment of breast cancer, mainly due to dose-limiting gastrointestinal toxicity. In this review, we discuss potential mechanisms of Notch-mediated resistance in breast cancer cells and breast cancer stem cells, and various methods of targeting Notch through γ-secretase inhibitors, Notch signaling biologics, or transcriptional inhibitors. We also discuss future plans for identification of novel Notch-targeted therapies, in order to reduce toxicity and improve outcomes for women with resistant breast cancer.

Introduction: The aim of this study was to determine the role of Notch in indoxyl sulfate (IS)-induced vascular calcification (VC). Materials and methods: VC and expression of Notch-related and osteogenic molecules were examined in Dahl salt-sensitive (DS), DS hypertensive (DH), and DH IS-treated rats (DH+IS). The effects of IS on expression of Notch receptors, apoptotic activity, and calcification were examined in cultured aortic smooth muscle cells (SMCs). Results: Medial calcification was noted only in aortas and coronary arteries of DH+IS rats. Notch1, Notch3, and Hes-1 were expressed in aortic SMCs of all rats, but only weakly in the central areas of the media and around the calcified lesions in DH+IS rats. RT-PCR and western blotting of DH+IS rat aortas showed downregulation of Notch ligands, Notch1 and Notch3, downstream transcriptional factors, and SM22, and conversely, overexpression of osteogenic markers. Expression of Notch1 and Notch3 in aortic SMCs was highest in incubation under 500 μM IS for 24hrs, and then decreased time- and dose-dependently. Coupled with this decrease, IS increased caspase 3/7 activity and TUNEL-positive aortic SMCs. In addition, pharmacological Notch signal inhibition with DAPT induced apoptosis in aortic SMCs. ZVAD, a caspase inhibitor abrogated IS-induced and DAPT-induced in vitro vascular calcification. Knockdown of Notch1 and Notch3 cooperatively increased expression of osteogenic transcriptional factors and decreased expression of SM22. Conclusion: Our results suggested that IS-induced VC is mediated through suppression of Notch activity in aortic SMCs, induction of osteogenic differentiation and apoptosis., Competing Interests: Competing Interests: S.G. is employed by Kureha Co. All other authors have no competing interests to declare. Kureha Co had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results., (© The author(s).)

Purpose: Inhibition of Notch signaling has been recently demonstrated to promote beige adipocyte biogenesis. However, most γ-secretase inhibitors (GSIs) used to achieve pharmacological inhibition of Notch signaling are at the basic research or preclinical stage, limiting the translation of fundamental findings into clinical practice. This present study aimed to evaluate the potential of several clinical candidates of GSIs as browning agents for the treatment of obesity., Methods: Seven GSIs that are clinical candidates for the treatment of Alzheimer's disease or cancer were selected and their impacts on Notch inhibition as well as promoting beige biogenesis were compared using in vitro culture of 3T3-L1 preadipocytes., Results: Four compounds (i.e.RO4929097, PF-03084014, LY3039478, and BMS-906024) that efficiently inhibited the expression of Notch target genes in 3T3-L1 preadipocytes were identified. Moreover, these compounds were optimized for dose-dependent effects at three gradient concentrations (0.5, 1, and 10 μM) to promote beige adipogenesis and mitochondrial biogenesis in 3T3-L1 preadipocytes without causing severe cytotoxicity., Conclusions: Our findings not only highlight the potential of cross-therapeutic application of these GSIs for obesity treatment via inhibition of γ-secretase-mediated processing of Notch signaling, but also provide important experimental evidence to support further design and development of clinically translatable Notch-inhibiting drug delivery systems.

Activation of Notch signaling is associated with tumor aggressiveness, poor clinical outcome and drug resistance in breast cancer patients. Targeting Notch signaling with small molecule inhibitors may be a better strategy for anticancer drug development. We identified 3-O-(E)-p-Coumaroylbetulinic acid (CB) as a lead compound and potent inhibitor of Notch signaling pathway. Treatment of human breast cancer MBA-MD-231 and T47D cells with CB resulted in a dose- and time-dependent inhibition of cell viability and G0/G1-phase cell cycle arrest. This effect was associated with a marked decrease in the expression of cyclin D1 and its activating partner, cyclin-dependent kinase 2 with concomitant increase in cyclin kinase inhibitor p21, operative in G1-phase of the cell cycle. CB treatment induced early apoptosis in breast cancer cells as evident by increase in cleaved caspase-3, decrease in Bcl2 and survivin, surge in reactive oxygen species and disruption of mitochondrial membrane potential. CB treatment altered Notch target genes viz. Hes1, Hey1 and E-cadherin at mRNA and protein level in time-dependent manner along with decrease in Notch promoter activity at IC 50 concentration. Furthermore, CB treatment decreased mammosphere formation in MCF-7 cells through down-modulation of the Notch signaling pathway and suppression of self-renewal markers such as c-Myc, SOX-2 and CD44. Our findings demonstrate that CB possess anticancer activity in breast cancer cells and suppresses self-renewal ability in the mammosphere as a result of modulation in cell-cycle machinery, disruption of mitochondrial function, induction of apoptosis, and Notch inhibition., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Notch signaling is often involved in cancer cell initiation and proliferation. Aberrant Notch activation underlies more than 50% of T-cell acute lymphoblastic leukemia (T-ALL); accordingly, chemicals disrupting Notch signaling are of potential to treat Notch-dependent cancer. Here, we developed a flow cytometry-based high-throughput assay to identify compounds that disrupt the interactions of DNA and RBPJ, the major downstream effector of Notch signaling. From 1492 compounds, we identified 18 compounds that disrupt RBPJ-DNA interactions in a dose-dependent manner. Cell-based assays further revealed that auranofin downregulates Notch-dependent transcription and decreases RBPJ-chromatin interactions in cells. Most strikingly, T-ALL cells that depend on Notch signaling for proliferation are more sensitive to auranofin treatment, supporting the notion that auranofin downregulates Notch signaling by disrupting RBPJ-DNA interaction. These results validate the feasibility of our assay scheme to screen for additional Notch inhibitors and provide a rationale to further test the use of auranofin in treating Notch-dependent cancer.

Since Notch signaling plays important roles in cell proliferation and differentiation, aberrant activation of this signaling contributes to cancer progression. In neural stem cells, Notch signaling inhibits differentiation by activating HES1 expression. Therefore, Notch signaling inhibitors may be candidates for new anticancer drugs or have applications in neural regenerative medicine. In this study, six naturally occurring Notch inhibitors were isolated from the methanol (MeOH) extract of Lansium domesticum using our novel cell-based assay. Hongherin (2), a cardiac glycoside, demonstrated potent Notch inhibitory activity with an IC 50 of 0.62 μM and was found to be cytotoxic in HPB-ALL human T cell acute lymphoblastic leukemia cells. Hongherin (2) also induced the differentiation of C17.2 neural stem cells to neurons, causing a 65% increase in differentiation compared to the control. Mechanistically, hongherin (2) reduced the amount of Notch1 (full length) and mastermind-like protein (MAML). This indicates that hongherin (2) inhibits Notch signaling through a dual mechanism involving the reduction of both Notch1 and MAML protein levels.

Understanding and targeting Notch signaling effectively has long been valued in the field of cancer and other immune disorders. Here, we discuss key discoveries at the intersection of Notch signaling, cancer and immunology. While there is a plethora of Notch targeting agents tested in vitro , in vivo and in clinic, undesirable off-target effects and therapy-related toxicities have been significant obstacles. We make a case for the clinical application of ligand-derived and affinity modifying compounds as novel therapeutic agents and discuss major research findings with an emphasis on Notch ligand-specific modulation of immune responses., (Copyright © 2020 Goruganthu, Shanker, Dikov and Carbone.)

As an evolutionarily conserved pathway, Hippo signaling pathway impacts different pathology and physiology processes such as wound healing, tissue repair/size and regeneration. When some components of Hippo signaling dysregulated, it affects cancer cells proliferation. Moreover, the relation Hippo pathway with other signaling including Wnt, TGFβ, Notch, and EGFR signaling leaves effect on the proliferation of cancer cells. Utilizing a number of therapeutic approaches, such as siRNAs and long noncoding RNA (lncRNA) to prevent cancer cells through the targeting of Hippo pathways, can provide new insights into cancer target therapy. The purpose of present review, first of all, is to demonstrate the importance of Hippo signaling and its relation with other signaling pathways in cancer. It also tries to demonstrate targeting Hippo signaling progress in cancer therapy., (© 2020 International Union of Biochemistry and Molecular Biology.)

Abdominal aortic aneurysm (AAA) is a localized pathological dilation of the aorta exceeding the normal diameter (∼20 mm) by more than 50% of its original size (≥30 mm), accounting for approximately 150000-200000 deaths worldwide per year. We previously reported that Notch inhibition does not decrease the size of pre-established AAA at late stage of the disease. Here, we examined whether a potent pharmacologic inhibitor of Notch signaling (DAPT (N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester)), regresses an actively growing AAA. In a mouse model of an aneurysm (Apoe-/- mice; n=44); DAPT (n=17) or vehicle (n=17) was randomly administered at day 14 of angiotensin II (AngII; 1 µg/min/kg), three times a week and mice were killed on day 42. Progressive increase in aortic stiffness and maximal intraluminal diameter (MILD) was observed in the AngII + vehicle group, which was significantly prevented by DAPT (P<0.01). The regression of aneurysm with DAPT was associated with reduced F4/80+Cd68+ (cluster of differentiation 68) inflammatory macrophages. DAPT improved structural integrity of aorta by reducing collagen fibrils abnormality and restoring their diameter. Mechanistically, C-C chemokine receptor type 7 (Ccr7)+F4/80- dendritic cells (DCs), implicated in the regression of aneurysm, were increased in the aorta of DAPT-treated mice. In the macrophages stimulated with AngII or lipopolysaccharide (LPS), DAPT reverted the expression of pro-inflammatory genes Il6 and Il12 back to baseline within 6 h compared with vehicle (P<0.05). DAPT also significantly increased the expression of anti-inflammatory genes, including c-Myc, Egr2, and Arg1 at 12-24 h in the LPS-stimulated macrophages (P<0.05). Overall, these regressive effects of Notch signaling inhibitor emphasize its therapeutic implications to prevent the progression of active AAAs., (© 2020 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Platinum-based chemoradiotherapy is a mainstay of organ-preserving therapy for patients with head and neck squamous cell carcinoma cancer (HNSCC). However, the disease eventually becomes resistant to treatment necessitating new therapies. Checkpoint kinase 1 and 2 (CHK1/2) are serine/threonine kinases that activate cell-cycle checkpoints and serve a critical role in the DNA-damage response (DDR). As resistance to cisplatin and radiation may involve a heightened DDR, we hypothesized that prexasertib, an inhibitor of CHK1/2, may enhance the cytotoxicity induced by cisplatin and irradiation in HNSCC. In this study, we found that combining prexasertib with cisplatin and radiation significantly decreased the in vitro survival fraction in HNSCC cell lines both with and without radiotherapy. Reduced survival was accompanied by inhibition of DNA repair checkpoint activation, which resulted in persistent DNA damage and increased apoptosis. In addition, NanoString analysis with the PanCancer Pathways Panel revealed that prexasertib downregulated NOTCH signaling target genes ( NOTCH1, NOTCH2 , and NOTCH3 ) and their associated ligands (JAG1, JAG2, SKP2, MAML2, and DLL1). Prexasertib also reduced NOTCH1, NOTCH3 and HES1 protein expression. Importantly, a significant tumor growth delay was observed in vivo in both human papillomavirus (HPV)-positive UM-SCC47 and HPV-negative UM-SCC1 cell line xenografts treated with prexasertib, cisplatin, and radiotherapy without increased toxicity as measured by mouse body weight. Taken together, prexasertib reduced NOTCH signaling and enhanced the in vitro and in vivo response of HNSCCs to cisplatin and radiation, suggesting combination therapy may increase clinical benefit. A clinical trial has recently completed accrual (NCT02555644)., (©2020 American Association for Cancer Research.)

In hepatocellular carcinoma (HCC), the poor response to the chemotherapeutic agents is partially attributed to the chemoresistance property of cancer stem cells (CSCs). NOTCH signaling pathway plays a crucial role in the chemoresistance through the maintenance of the CSCs. We observed that the NOTCH pathway was activated in HCC CD133 + cells treated with vincristine (VIN) 1 and 5-fluorouracil (5-FU) 2 . Therefore, we examined whether inhibition of the NOTCH can improve sensitization of HCC CD133 + cells to VIN and 5-FU. The Huh7 cell line was pre-incubated γ-secretase DAPT, as a NOTCH inhibitor, and then treated with IC 50 dose of VIN or 5-FU. The CD133 + cells were then isolated and analyzed for the cell viability, apoptosis, migration and spheroid formation capacities, and gene and protein expression. It was observed that pre-incubation with DAPT significantly downregulated the expression of NOTCH-related genes and led to a significant reduction in VIN- and 5-FU-CD133 + population. In addition, DAPT pre-incubated VIN- and 5-FU-treated-CD133 + cells formed fewer spheroids in 3D culture and had a lesser migration capacity in 2D culture. Importantly, DAPT enhanced the apoptosis rate of VIN- and 5-FU-treated CD133 + cells for 3- and 2-fold, which was correlated with the enhanced expression of pro-apoptotic BBC3 (BCL-2-binding component 3) and decreased expression of HES1 that was reported to regulate BBC3 negatively. Collectively, it was observed that NOTCH inhibition sensitized the HCC CD133 + cells to VIN and 5-FU through enhancing BBC3-mediated apoptosis. The results highlighted the role of NOTCH/HES1/BBC3 axis in resistance of CD133 + cells to VIN and 5-FU. Understanding the molecular mechanisms underlying chemoresistance in HCC CD133 + cells may help in designing the novel targeted therapies to chemosensitize them., Competing Interests: Declaration of competing interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Cell signaling pathways play key roles in coordinating cellular events in development. The Notch signaling pathway is highly conserved across all multicellular animals and is known to coordinate a multitude of diverse cellular events, including proliferation, differentiation, fate specification, and cell death. Specific functions of the pathway are, however, highly context-dependent and are not well characterized in post-traumatic regeneration. Here, we use a small-molecule inhibitor of the pathway (DAPT) to demonstrate that Notch signaling is required for proper arm regeneration in the brittle star Ophioderma brevispina, a highly regenerative member of the phylum Echinodermata. We also employ a transcriptome-wide gene expression analysis (RNA-seq) to characterize the downstream genes controlled by the Notch pathway in the brittle star regeneration. We demonstrate that arm regeneration involves an extensive cross-talk between the Notch pathway and other cell signaling pathways. In the regrowing arm, Notch regulates the composition of the extracellular matrix, cell migration, proliferation, and apoptosis, as well as components of the innate immune response. We also show for the first time that Notch signaling regulates the activity of several transposable elements. Our data also suggests that one of the possible mechanisms through which Notch sustains its activity in the regenerating tissues is via suppression of Neuralized1., Competing Interests: The authors have declared that no competing interests exist.

Endometrial cancer (EC) is the most common gynecologic malignancy in world. It has been reported that the mutation rate of FBXW7 is frequent in EC, but the specific functions of FBXW7 remain unknown in EC. In the present study, we revealed the role and mechanism of FBXW7 in EC cells. Compared with adjacent nontumor tissues, the FBXW7 expression level was lower in EC tissues. However, the level of STYX was in contrast with the expression of FBXW7 in EC tissues. And STYX interacted with FBXW7 and then down-regulated its expression level in EC. Over-expression of FBXW7 inhibited cell proliferation and facilitated apoptosis in EC cells, whereas silencing FBXW7 acted an opposite effect on EC cells. And the process of FBXW7 participated the proliferation and apoptosis in EC was regulated by STYX. FBXW7 suppressed the expression of Notch pathway related protein, and further inhibited the phosphorylation of mTOR. In addition, we also found that mTOR activitor (MHY1485) and Notch activator (Jagged-1) reversed the effect of over-expressing FBXW7 on cell proliferation and cell apoptosis. And Notch inhibitor (DAPT) counteracted the impact of over-expressing STYX on cell proliferation and cell apoptosis. Collectively, the present study verified that STYX inhibited the expression level of FBXW7 in EC, and then promoted cell proliferation but suppressed apoptosis through Notch-mTOR signaling pathway, which promoted carcinogenesis and progression of EC., (© 2020 The Author(s).)

Background: Systemic sclerosis (SSc) is characterised by autoimmune activation, tissue and vascular fibrosis in the skin and internal organs. Tissue fibrosis is driven by myofibroblasts, that are known to maintain their phenotype in vitro, which is associated with epigenetically driven trimethylation of lysine 27 of histone 3 (H3K27me3)., Methods: Full-thickness skin biopsies were surgically obtained from the forearms of 12 adult patients with SSc of recent onset. Fibroblasts were isolated and cultured in monolayers and protein and RNA extracted. HOX transcript antisense RNA (HOTAIR) was expressed in healthy dermal fibroblasts by lentiviral induction employing a vector containing the specific sequence. Gamma secretase inhibitors were employed to block Notch signalling. Enhancer of zeste 2 (EZH2) was blocked with GSK126 inhibitor., Results: SSc myofibroblasts in vitro and SSc skin biopsies in vivo display high levels of HOTAIR, a scaffold long non-coding RNA known to direct the histone methyltransferase EZH2 to induce H3K27me3 in specific target genes. Overexpression of HOTAIR in dermal fibroblasts induced EZH2-dependent increase in collagen and α-SMA expression in vitro, as well as repression of miRNA-34A expression and consequent NOTCH pathway activation. Consistent with these findings, we show that SSc dermal fibroblast display decreased levels of miRNA-34a in vitro. Further, EZH2 inhibition rescued miRNA-34a levels and mitigated the profibrotic phenotype of both SSc and HOTAIR overexpressing fibroblasts in vitro., Conclusions: Our data indicate that the EZH2-dependent epigenetic phenotype of myofibroblasts is driven by HOTAIR and is linked to miRNA-34a repression-dependent activation of NOTCH signalling., Competing Interests: Competing interests: H.Y.C. is a co-founder of Accent Therapeutics, Boundless Bio, and advisor to 10x Genomics, Arsenal Biosciences, and Spring Discovery., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Current treatments for clear cell renal cell cancer (ccRCC) are insufficient because two-thirds of patients with metastases progress within two years. Here we report the identification and characterization of a cancer stem cell (CSC) population in ccRCC. CSCs are quantitatively correlated with tumor aggressiveness and metastasis. Transcriptional profiling and single cell sequencing reveal that these CSCs exhibit an activation of WNT and NOTCH signaling. A significant obstacle to the development of rational treatments has been the discrepancy between model systems and the in vivo situation of patients. To address this, we use CSCs to establish non-adherent sphere cultures, 3D tumor organoids, and xenografts. Treatment with WNT and NOTCH inhibitors blocks the proliferation and self-renewal of CSCs in sphere cultures and organoids, and impairs tumor growth in patient-derived xenografts in mice. These findings suggest that our approach is a promising route towards the development of personalized treatments for individual patients.

Rheumatoid arthritis affects individuals commonly during the most productive years of adulthood. Poor response rates and high costs associated with treatment mandate the search for new therapies. Here we show that targeting a specific G-protein coupled receptor promotes senescence in synovial fibroblasts, enabling amelioration of joint inflammation. Following activation of the melanocortin type 1 receptor (MC 1 ), synovial fibroblasts acquire a senescence phenotype characterized by arrested proliferation, metabolic re-programming and marked gene alteration resembling the remodeling phase of wound healing, with increased matrix metalloproteinase expression and reduced collagen production. This biological response is attained by selective agonism of MC 1 , not shared by non-selective ligands, and dependent on downstream ERK1/2 phosphorylation. In vivo, activation of MC 1 leads to anti-arthritic effects associated with induction of senescence in the synovial tissue and cartilage protection. Altogether, selective activation of MC 1 is a viable strategy to induce cellular senescence, affording a distinct way to control joint inflammation and arthritis.

Myocardial infarction is a frequent cardiovascular event and a major cause for cardiomyocyte loss. In adult mammals, cardiomyocytes are traditionally considered to be terminally differentiated cells, unable to proliferate. Therefore, the wound-healing response in the infarct area typically yields scar tissue rather than newly formed cardiomyocytes. In the last decade, several lines of evidence have challenged the lack of proliferative capacity of the differentiated cardiomyocyte: studies in zebrafish and neonatal mammals have convincingly demonstrated the regenerative capacity of cardiomyocytes. Moreover, multiple signaling pathways have been identified in these models that-when activated in adult mammalian cardiomyocytes-can reactivate the cell cycle in these cells. However, cardiomyocytes frequently exit the cell cycle before symmetric division into daughter cells, leading to polyploidy and multinucleation. Now that there is more insight into the reactivation of the cell cycle machinery, other prerequisites for successful symmetric division of cardiomyocytes, such as the control of sarcomere disassembly to allow cytokinesis, require more investigation. This review aims to discuss the signaling pathways involved in cardiomyocyte proliferation, with a specific focus on wingless/int-1 protein signaling. Comparing the conflicting results from in vitro and in vivo studies on this pathway illustrates that the interaction with other cells and structures around the infarct is likely to be essential to determine the outcome of these interventions. The extensive crosstalk with other pathways implicated in cardiomyocyte proliferation calls for the identification of nodal points in the cell signaling before cardiomyocyte proliferation can be moved forward toward clinical application as a cure of cardiac disease. SIGNIFICANCE STATEMENT: Evidence is mounting that proliferation of pre-existing cardiomyocytes can be stimulated to repair injury of the heart. In this review article, an overview is provided of the different signaling pathways implicated in cardiomyocyte proliferation with emphasis on wingless/int-1 protein signaling, crosstalk between the pathways, and controversial results obtained in vitro and in vivo., (Copyright © 2020 by The Author(s).)