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Triple-negative breast cancer (TNBC), is diagnosed as the most lethal molecular subtype of breast cancer (BC) preceded by an extremely poor prognosis. For enabling effective TNBC therapy, the identification of novel druggable biomarkers is an earnest need. Multigene paneling and genomewide association studies identify multiple genes with high-to-moderate penetrance in TNBC. Modern computer-aided drug designing techniques, thus aim to design more cost-effective natural small molecule inhibitors for TNBC prevention and diagnosis. Here Amygdalin, a natural glycosidic inhibitor is docked and simulated against three such high-to-moderate penetrance genes identified in TNBC, BARD1, RAD51, and PALB2. The preliminary result of the analysis, reports a highest, intermediate, and least binding energy score of - 6.69 kcal/mol, - 5.09 kcal/mol, and - 4.89 kcal/mol in BARD1, RAD51, and PALB2, respectively. The best-docked protein-ligand complex (BARD1-Amygdalin) was then simulated and compared with an approved drug for TNBC treatment, Olaparib. A comparable binding energy score of - 8.53 kcal/mol was obtained by docking olaparib with BARD1. A 100 ns MD simulation revealed, Amygdalin forms more H-bonds, providing more stable and compact protein-ligand complex with BARD1 than compared to Olaparib. The result was also supported by calculation of solvent accessible surface area and analysis of radius of gyration. Thus, our findings suggest that role of Amygdalin can further be studied in details for TNBC therapeutics, which was found to target the BRCT domain of the BARD1 receptor in stable manner. Please check and confirm that the authors and their respective affiliations have been correctly identified and amend if necessary. Name and affiliations are correctly identified., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

We recently presented Stafia-1 as the first chemical entity that inhibits the transcription factor STAT5a with selectivity over the highly homologous STAT5b. Stafia-1, which was identified from a series of symmetrically substituted m-terphenyl phosphates, binds to the interface between the SH2 domain and the linker domain of STAT5a. Here, we outline a synthetic strategy for the synthesis of asymmetrically substituted m-terphenyl phosphates, which can be tailored to address their asymmetric STAT5a binding site in a more specific manner. The asymmetrically substituted m-terphenyl phosphate with the highest activity against STAT5a was converted to a phosphatase-stable monofluoromethylene phosphonate. The synthetic methodology and activity analysis described here provide first insights into the structure-activity relationships of m-terphenyl phosphates for use as selective STAT5a inhibitors., (© 2021 The Authors. ChemBioChem published by Wiley-VCH GmbH.)

We successfully repurpose the DNA repair protein methylguanine methyltransferase (MGMT) as an inducible degron for protein fusions. MGMT is a suicide protein that removes alkyl groups from the O 6 position of guanine (O 6 G) and is thereafter quickly degraded by the ubiquitin proteasome pathway (UPP). Starting with MGMT pseudosubstrates (benzylguanine and lomeguatrib), we first demonstrate that these lead to potent MGMT depletion while affecting little else in the proteome. We then show that fusion proteins of MGMT undergo rapid UPP-dependent degradation in response to pseudosubstrates. Mechanistic studies confirm the involvement of the UPP, while revealing that at least two E3 ligase classes can degrade MGMT depending on cell-line and expression type (native or ectopic). We also demonstrate the technique's versatility with two clinically relevant examples: degradation of KRAS G12C and a chimeric antigen receptor.

O 6 -Methylguanine-DNA-methyltransferase (MGMT) is a key DNA repair enzyme involved in chemoresistance to DNA-alkylating anti-cancer drugs such as Temozolomide (TMZ) through direct repair of drug-induced O 6 -methylguanine residues in DNA. MGMT substrate analogues, such as O 6 -benzylguanine (BG), efficiently inactivate MGMT in vitro and in cells; however, these drugs failed to reach the clinic due to adverse side effects. Here, we designed hybrid drugs combining a BG residue covalently linked to a DNA-interacting moiety (6-chloro-2-methoxy-9-aminoacridine). Specifically, two series of hybrids, encompassing three compounds each, were obtained by varying the position of the attachment point of BG (N 9 of guanine vs. the benzyl group) and the length and nature of the linker. UV/vis absorption and fluorescence data indicate that all six hybrids adopt an intramolecularly stacked conformation in aqueous solutions in a wide range of temperatures. All hybrids interact with double-stranded DNA, as clearly evidenced by spectrophotometric titrations, without intercalation of the acridine ring and do not induce thermal stabilization of the duplex. All hybrids, as well as the reference DNA intercalator (6-chloro-2-methoxy-9-aminoacridine 8), irreversibly inhibit MGMT in vitro with variable efficiency, comparable to that of BG. In a multidrug-resistant glioblastoma cell line T98G, benzyl-linked hybrids 7a-c and the N 9 -linked hybrid 19b are moderately cytotoxic (GI 50  ≥ 15 μM after 96 h), while N 9 -linked hybrids 19a and 19c are strongly cytotoxic (GI 50  = 1-2 μM), similarly to acridine 8 (GI 50  = 0.6 μM). Among all compounds, hybrids 19a and 19c, similarly to BG, display synergic cytotoxic effect upon co-treatment with subtoxic doses of TMZ, with combination index (CI) values as low as 0.2-0.3. In agreement with in vitro results, compound 19a inactivates cellular MGMT but, unlike BG, does not induce significant levels of DNA damage, either alone or in combination with TMZ, as indicated by the results of γH2AX immunostaining experiments. Instead, and unlike BG, compound 19a alone induces significant apoptosis of T98G cells, which is not further increased in a combination with TMZ. These results indicate that molecular mechanisms underlying the cytotoxicity of 19a and its combination with TMZ are distinct from that of BG. The strongly synergic properties of this combination represent an interesting therapeutic opportunity in treating TMZ-resistant cancers., 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 © 2021 Elsevier Masson SAS. All rights reserved.)

Taxol is commonly used chemotherapy regimen for esophageal squamous cell carcinoma (ESCC). Study of the underlying mechanisms of Taxol chemoresistance provides better understanding of esophageal cancer treatment and may provide a rational molecular target for diagnosis and intervention. Here we showed FBXO31, which was reported to be highly expressed in ESCC and significantly associated with poor prognosis, could regulate ESCC chemosensitivity to Taxol. Silencing of FBXO31 in ESCC cells sensitized cells to Taxol treatment, evidenced by FACS analysis and TUNEL assay, showing as an increased apoptotic population in FBXO31-knockdown cells compared to the control cells. The mass spectrometry data and coimmunoprecipitation results showed FBXO31 could bind with cofilin-1. Cofilin-1 knockdown in FBXO31-overexpression cells reversed FBXO31-induced suppression of cell apoptosis, suggesting FBXO31-mediated Taxol chemoresistance is associated with cofilin-1. Furthermore, in vivo experiments confirmed that knockdown of FBXO31 sensitized ESCC to Taxol treatment. This finding substantiated a pivotal role of FBOX31 in ESCC chemoresistance, indicating that FBXO31 may be a potential indicator or target for drug resistance in ESCC., 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 © 2021 Elsevier Inc. All rights reserved.)

Chloroethylnitrosoureas (CENUs) are an important family of chemotherapies in clinical treatment of cancers, which exert antitumor activity by inducing the formation of DNA interstrand crosslinks (dG-dC ICLs). However, the drug resistance mediated by O 6 -alkylguanine-DNA alkyltransferase (AGT) and absence of tumor-targeting ability largely decrease the antitumor efficacy of CENUs. In this study, we synthesized an azobenzene-based hypoxia-activated combi-nitrosourea prodrug, AzoBGNU, and evaluated its hypoxic selectivity and antitumor activity. The prodrug was composed of a CENU pharmacophore and an O 6 -benzylguanine (O 6 -BG) analog moiety masked by a N,N-dimethyl-4-(phenyldiazenyl)aniline segment as a hypoxia-activated trigger, which was designed to be selectively reduced via azo bond break in hypoxic tumor microenvironment, accompanied with releasing of an O 6 -BG analog to inhibit AGT and a chloroethylating agent to induce dG-dC ICLs. AzoBGNU exhibited significantly increased cytotoxicity and apoptosis-inducing ability toward DU145 cells under hypoxia compared with normoxia, indicating the hypoxia-responsiveness as expected. Predominant higher cytotoxicity was observed in the cells treated by AzoBGNU than those by traditional CENU chemotherapy ACNU and its combination with O 6 -BG. The levels of dG-dC ICLs in DU145 cells induced by AzoBGNU was remarkably enhanced under hypoxia, which was approximately 6-fold higher than those in the AzoBGNU-treated groups under normoxia and those in the ACNU-treated groups. The results demonstrated that azobenzene-based combi-nitrosourea prodrug possessed desirable tumor-hypoxia targeting ability and eliminated chemoresistance compared with the conventional CENUs., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Multiple noncoding natural antisense transcripts (ncNAT) are known to modulate key biological events such as cell growth or differentiation. However, the actual impact of ncNATs on cancer progression remains largely unknown. In this study, we identified a complete list of differentially expressed ncNATs in hepatocellular carcinoma. Among them, a previously undescribed ncNAT HNF4A-AS1L suppressed cancer cell growth by regulating its sense gene HNF4A , a well-known cancer driver, through a promoter-specific mechanism. HNF4A-AS1L selectively activated the HNF4A P1 promoter via HNF1A, which upregulated expression of tumor suppressor P1-driven isoforms, while having no effect on the oncogenic P2 promoter. RNA-seq data from 23 tissue and cancer types identified approximately 100 ncNATs whose expression correlated specifically with the activity of one promoter of their associated sense gene. Silencing of two of these ncNATs ENSG00000259357 and ENSG00000255031 (antisense to CERS2 and CHKA , respectively) altered the promoter usage of CERS2 and CHKA . Altogether, these results demonstrate that promoter-specific regulation is a mechanism used by ncNATs for context-specific control of alternative isoform expression of their counterpart sense genes. SIGNIFICANCE: This study characterizes a previously unexplored role of ncNATs in regulation of isoform expression of associated sense genes, highlighting a mechanism of alternative promoter usage in cancer., (©2021 The Authors; Published by the American Association for Cancer Research.)

Herein we describe a native mass spectromery protein-peptide model as a competent surrogate for the HOP-HSP90 protein-protein interaction (PPI), application of which led to the qualititive identification of two new peptides capable of in vitro PPI disruption. This proof of concept study offers a viable alternative for PPI inhibitor screening.

Particulate matters (PMs) increase oxidative stress and inflammatory response in different tissues. PMs disrupt the formation of primary cilia in various skin cells, including keratinocytes and melanocytes. In this study, we found that 2-isopropylmalic acid (2-IPMA) promoted primary ciliogenesis and restored the PM2.5-induced dysgenesis of primary cilia in dermal fibroblasts. Moreover, 2-IPMA inhibited the generation of excessive reactive oxygen species and the activation of stress kinase in PM2.5-treated dermal fibroblasts. Further, 2-IPMA inhibited the production of pro-inflammatory cytokines, including IL-6 and TNF-α, which were upregulated by PM2.5. However, the inhibition of primary ciliogenesis by IFT88 depletion reversed the downregulated cytokines by 2-IPMA. Moreover, we found that PM2.5 treatment increased the MMP-1 expression in dermal fibroblasts and a human 3-D-skin model. The reduced MMP-1 expression by 2-IPMA was further reversed by IFT88 depletion in PM2.5-treated dermal fibroblasts. These findings suggest that 2-IPMA ameliorates PM2.5-induced inflammation by promoting primary ciliogenesis in dermal fibroblasts.

Mammalian cell cytoskeletal reorganization for efficient directional movement requires tight coordination of actomyosin and microtubule networks. In this study, we show that LRAP35a potentiates microtubule stabilization by promoting CLASP2/EB1 interaction besides its complex formation with MRCK/MYO18A for retrograde actin flow. The alternate regulation of these two networks by LRAP35a is tightly regulated by a series of phosphorylation events that dictated its specificity. Sequential phosphorylation of LRAP35a by Protein Kinase A (PKA) and Glycogen Synthase Kinase-3β (GSK3β) initiates the association of LRAP35a with CLASP2, while subsequent binding and further phosphorylation by Casein Kinase 1δ (CK1δ) induce their dissociation, which facilitates LRAP35a/MRCK association in driving lamellar actomyosin flow. Importantly, microtubule dynamics is directly moderated by CK1δ activity on CLASP2 to regulate GSK3β phosphorylation of the SxIP motifs that blocks EB1 binding, an event countered by LRAP35a interaction and its competition for CK1δ activity. Overall this study reveals an essential role for LRAP35a in coordinating lamellar contractility and microtubule polarization in cell migration., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objective: Prostate cancer (PCa) is considered the most serious cancer in the world. Nevertheless, the accuracy of current biomarkers, such as pathological staging, Gleason's score, and serum prostate-specific antigen (PSA) levels, is limited. FOXO1 is a key downstream effector of PTEN and a tumor suppressor in PCA, which has been reported extensively. However, the clinical relevance of FOXO1 in PCa remains unclear., Methods: In this study, we first detected its expression in four public databases to explore the clinical role of FOXO1. Verification of the knockdown effect of FOXO1 siRNA was performed by real-time PCR analysis. Changes in cell viability were assessed using cell counting kit-8 (CCK-8) assays. In addition, we verified the effect of FOXO1 on the PCa cell cycle using a cell cycle assay., Results: Herein, we found that FOXO1 was significantly downregulated in PCa tissues and was significantly associated with Gleason's score, age, biochemical recurrence (BCR), and lymph node (LN) status, while FOXO1 expression was independent of pathological staging and preoperative PSA levels. The Kaplan-Meier survival analysis showed that PCA patients with high FOXO1 expression were less likely to develop BCR compared with patients with low FOXO1 expression. In terms of function, FOXO1 inhibition significantly promoted the proliferation and cell cycle progression of PCa cells., Conclusions: In summary, our study suggests that FOXO1 may be one of the prognostic factors that describe the risk of PCa for BCR. These results suggest that FOXO1 may be a therapeutic target for PCa., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2021 Ning Yang et al.)

Inhibitor of growth 3 (ING3) is one of five members of the ING tumour suppressor family, characterized by a highly conserved plant homeodomain (PHD) as a reader of the histone mark H3K4me3. ING3 was reported to act as a tumour suppressor in many different cancer types to regulate apoptosis. On the other hand, ING3 levels positively correlate with poor survival prognosis of prostate cancer (PCa) patients. In PCa cells, ING3 acts rather as an androgen receptor (AR) co-activator and harbours oncogenic properties in PCa. Here, we show the identification of a novel ING3 splice variant in both the human PCa cell line LNCaP and in human PCa patient specimen. The novel ING3 splice variant lacks exon 11, ING3∆ex11, which results in deletion of the PHD, providing a unique opportunity to analyse functionally the PHD of ING3 by a natural splice variant. Functionally, overexpression of ING3Δex11 induced morphological changes of LNCaP-derived 3D spheroids with generation of lumen and pore-like structures within spheroids. Since these structures are an indicator of epithelial-mesenchymal transition (EMT), key regulatory factors and markers for EMT were analysed. The data suggest that in contrast to ING3, ING3Δex11 specifically modulates the expression of key EMT-regulating upstream transcription factors and induces the expression of EMT markers, indicating that the PHD of ING3 inhibits EMT. In line with this, ING3 knockdown also induced the expression of EMT markers, confirming the impact of ING3 on EMT regulation. Further, ING3 knockdown induced cellular senescence via a pathway leading to cell cycle arrest, indicating an oncogenic role for ING3 in PCa. Thus, the data suggest that the ING3Δex11 splice variant lacking functional PHD exhibits oncogenic characteristics through triggering EMT in PCa cells.

Omics analyses are powerful methods to obtain an integrated view of complex biological processes, disease progression, or therapy efficiency. However, few studies have compared different disease forms and different therapy strategies to define the common molecular signatures representing the most significant implicated pathways. In this study, we used RNA sequencing and mass spectrometry to profile the transcriptomes and proteomes of mouse models for three forms of centronuclear myopathies (CNMs), untreated or treated with either a drug (tamoxifen), antisense oligonucleotides reducing the level of dynamin 2 (DNM2), or following modulation of DNM2 or amphiphysin 2 (BIN1) through genetic crosses. Unsupervised analysis and differential gene and protein expression were performed to retrieve CNM molecular signatures. Longitudinal studies before, at, and after disease onset highlighted potential disease causes and consequences. Main pathways in the common CNM disease signature include muscle contraction, regeneration and inflammation. The common therapy signature revealed novel potential therapeutic targets, including the calcium regulator sarcolipin. We identified several novel biomarkers validated in muscle and/or plasma through RNA quantification, western blotting, and enzyme-linked immunosorbent assay (ELISA) assays, including ANXA2 and IGFBP2. This study validates the concept of using multi-omics approaches to identify molecular signatures common to different disease forms and therapeutic strategies., Competing Interests: Declaration of interests J.L. is co-founder of Dynacure (Illkirch, France). The remaining authors declare no competing interests., (Copyright © 2021 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.)

The FMS-like tyrosine kinase 3 ( FLT3 ) gene is mutated in one-third of patients with de novo acute myeloid leukemia (AML). Mutated FLT3 variants are constitutively active kinases signaling via AKT kinase, MAP kinases, and STAT5. FLT3 inhibitors have been approved for the treatment of FLT3 -mutated AML. However, treatment response to FLT3 inhibitors may be short-lived, and resistance may emerge. Compounds targeting STAT5 may enhance and prolong effects of FLT3 inhibitors in this subset of patients with FLT3 -mutated AML. Here STAT5-inhibitor AC-4-130, FLT3 inhibitor midostaurin (PKC412), BMI-1 inhibitor PTC596, MEK-inhibitor trametinib, MCL1-inhibitor S63845, and BCL-2 inhibitor venetoclax were assessed as single agents and in combination for their ability to induce apoptosis and cell death in leukemic cells grown in the absence or presence of bone marrow stroma. Synergistic effects on cell viability were detected in both FLT3 -mutated and FLT3 -wild-type AML cells treated with AC-4-130 in combination with the MCL1 inhibitor S63845. AML patient samples with a strong response to AC-4-130 and S63845 combination treatment were characterized by mutated FLT3 or mutated TET2 genes. Susceptibility of AML cells to AC-4-130, PTC596, trametinib, PKC412, and venetoclax was altered in the presence of HS-5 stroma. Only the MCL1 inhibitor S63845 induced cell death with equal efficacy in the absence or presence of bone marrow stroma. The combination of the STAT5-inhibitor AC-4-130 and the MCL1 inhibitor S63845 may be an effective treatment targeting FLT3 -mutated or TET2 -mutated AML.

Deleted in liver cancer 1 (DLC1) is a recognized tumor suppressor gene that negatively regulates Rho family proteins by hydrolyzing the active GTP-bound state to its inactive GDP-bound state. Active Rho proteins play a positive role in tumorigenesis. Numerous in vitro and in vivo experiments have shown that DLC1 is downregulated or inactivated in various solid tumors, which may be due to the following five reasons: genomic deletion, epigenetic modification and ubiquitin-dependent proteasomal degradation may cause DLC1 underexpression; phosphorylation at the post-translation level may cause DLC1 inactivation; and failure to localize at focal adhesions (FAs) may prevent DLC1 from exerting full activity. All of the causes could be attributed to molecular binding. Experimental evidence suggests that direct or indirect targeting of DLC1 is feasible for cancer treatment. Therefore, elucidating the interaction of DLC1 with its binding partners might provide novel targeted therapies for cancer. In this review, we summarized the binding partners of DLC1 at both the gene and protein levels and expounded a variety of anticancer drugs targeting DLC1 to provide information about DLC1 as a cancer diagnostic indicator or therapeutic target., (Copyright © 2021 Elsevier B.V. All rights reserved.)

It has been reported that the gut microbiome modulates postoperative cognitive dysfunction (POCD), and that administration of probiotics (VSL#3) may effectively relieve POCD. In this study, we aimed to identify the underlying mechanism of VSL#3 in POCD. A mouse model of POCD was constructed in adult male C57BL/6 mice, which were then treated with VSL#3. VSL#3 exerted a protective role against POCD and resultant neuronal apoptosis. The expression of miR-146a was found to be downregulated in hippocampal tissues of POCD mice, while VSL#3 could restore its expression. Loss- and gain-function approaches were conducted to determine the roles of microRNA (miR)-146a, B-cell translocation gene 2 (BTG2), and Bcl-2-associated X protein (Bax) in post-operative effects on cognitive function and neuronal apoptosis. The levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD) were measured to determine oxidative stress in brain tissue. The dual-luciferase reporter gene assay identified that miR-146a could target BTG2 and negatively regulate its expression. BTG2 knockdown suppressed neuronal apoptosis and contributed to shortened time of latency, prolonged time of mice spent in the target quadrant, and reduced oxidative stress through downregulating Bax expression. Finally, VSL#3 treatment upregulated the expression of miR-146a to block BTG2/Bax axis and consequently inhibited neuronal apoptosis and reduced oxidative stress in POCD mice. Taken together, the study suggested that miR-146a-mediated suppression of BTG2/Bax contributed to the protective role of probiotics treatment against POCD.

Aspirin can efficiently inhibit the glycolysis and proliferation of cancer cells, however, the underlying mechanism is poorly understood. Here, we report that aspirin attenuates the glycolysis and proliferation of hepatoma cells through modulating the levels of lysine 2-hydroxyisobutyrylation (Khib) of enolase 1 (ENO1). We found that aspirin decreased the levels of glucose consumption and lactate production in hepatoma cells. Moreover, 4 mM aspirin reduced the activities of ENO1, a key enzyme of glycolysis, and decreased the levels of ENO1 Khib in the cells. Interestingly, we identified that 4 mM aspirin could decrease the levels of Khib on many proteins by using pan Khib antibody in the cells. Interestingly, the activities of ENO1 could be rescued by the transient overexpression of ENO1, but not by ENO1 mutant (K281R). Moreover, we identified that the C646, an inhibitor of p300 which is a writer of Khib, could reduce the levels of ENO1 Khib, resulting in the decrease of ENO1 activities. The treatment with PDTC, an inhibitor of NF-κB which is a target of aspirin, could work well as C646 in the cells. Both of aspirin and C646 (or PDTC) displayed a stronger effect than the single treatment in the system. Functionally, ENO1, but not ENO1 mutant (K281R), could rescue the aspirin-induced inhibition of proliferation of liver cancer cells in vitro, suggesting that ENO1K281 is involved in the aspirin-mediated inhibition of liver cancer. Our finding provides new insights into the mechanism by which aspirin attenuates the glycolysis and proliferation of hepatoma cells., Competing Interests: Declaration of competing interest Ying Yuan, Hong-feng Yuan and Yu Geng performed experiments, and analyzed the data. Li-na Zhao, Hao-lin Yun, Yu-fei Wang contributed to the interpretation of the results. Xiao-dong Zhang designed, conceived, wrote the manuscript, and supervised the study. All authors read and approved the manuscript., (Copyright © 2021. Published by Elsevier Inc.)

Down-regulated in renal cell carcinoma 1 (DRR1), a unique stress-induced protein, is highly expressed in the nervous system. This study investigated the roles of DRR1 in the brain by examining its expression pattern at different developmental stages of a rat brain and in cultured primary hippocampal neurons. High expression of DRR1 was observed in all developmental stages of a rat brain and cultured primary hippocampal neurons. We then focused on the role of DRR1 in promoting neurite outgrowth during the early stage of hippocampal neuron development. Results showed that down-regulation of DRR1 suppressed axon outgrowth. Mass spectrometry analysis revealed that tropomodulin-2 (Tmod2) is a novel binding partner of DRR1. Our results showed that both DRR1 and Tmod2 mediate axon formation during the early stage of hippocampal neuron development. Suppression of TMOD2 expression rescued the abnormal axon outgrowth induced by DRR1 knockdown during the early stage of hippocampal neuron development., 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 © 2021 Elsevier Inc. All rights reserved.)

Hippo signaling is an evolutionarily conserved pathway that restricts growth and regeneration predominantly by suppressing the activity of the transcriptional coactivator Yap. Using a high-throughput phenotypic screen, we identified a potent and non-toxic activator of Yap. In vitro kinase assays show that the compound acts as an ATP-competitive inhibitor of Lats kinases-the core enzymes in Hippo signaling. The substance prevents Yap phosphorylation and induces proliferation of supporting cells in the murine inner ear, murine cardiomyocytes, and human Müller glia in retinal organoids. RNA sequencing indicates that the inhibitor reversibly activates the expression of transcriptional Yap targets: upon withdrawal, a subset of supporting-cell progeny exits the cell cycle and upregulates genes characteristic of sensory hair cells. Our results suggest that the pharmacological inhibition of Lats kinases may promote initial stages of the proliferative regeneration of hair cells, a process thought to be permanently suppressed in the adult mammalian inner ear.

Polo-like kinase 1 (Plk1) is an important regulator of the cell cycle and it is frequently overexpressed in cancer cells. Several small molecule inhibitors have been developed to target Plk1 and some of them have reached clinical trials in adults with acute myeloid leukemia (AML). Pediatric AML patients have a poor prognosis and survivors suffer from long-term side effects. As adult AML cells have an elevated expression of Plk1, AML is a disease candidate for Plk1 inhibition. However, the relative success of clinical trials have been hampered by adverse reactions. Herein, PLK1-targeting RNA interference (RNAi) prodrugs that enter cells without a transfection reagent are used to target PLK1 selectively in primary cells from pediatric AML patients. We show that PLK1 and PLK4 mRNA expression are significantly higher in pediatric AML patients when compared to healthy donors and that PLK1 is downregulated by on average 50% using RNAi prodrugs without a significant effect on other PLK family members. In addition, the RNAi prodrug-induced decrease in PLK1 can be used to potentiate the effect of cytarabine. In summary, PLK1-targeting RNAi prodrugs can decrease the elevated levels of PLK1 in primary cells from pediatric AML patients and sensitize pediatric AML cells to chemotherapeutics., (© 2021 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

An elastic network model (ENM) represents a molecule as a matrix of pairwise atomic interactions. Rich in coded information, ENMs are hereby proposed as a novel tool for the prediction of the activity of series of molecules, with widely different chemical structures, but a common biological activity. The new approach is developed and tested using a set of 183 inhibitors of serine/threonine-protein kinase enzyme (Plk3) which is an enzyme implicated in the regulation of cell cycle and tumorigenesis. The elastic network (EN) predictive model is found to exhibit high accuracy and speed compared to descriptor-based machine-trained modeling. EN modeling appears to be a highly promising new tool for the high demands of industrial applications such as drug and material design.

The development of resistance poses a serious problem in the therapy of cancer due to the necessity of a multiple-drug and unlimited treatment of affected patients. In chronic myeloid leukemia (CML), the introduction of imatinib has revolutionized the therapy. The persistence of an untreatable cancer stem cell pool and other resistance-causing factors, however, also impede the cure of this malignancy. New therapeutic approaches are therefore essential to overcome current treatment drawbacks. In this regard, an intervention in the STAT5 signaling pathway can significantly improve drug response, as this central signaling node induces the formation of highly resistant CML cells. In the present study, we continued the design of efficient chemosensitizers derived from the partial PPARγ agonist telmisartan. The developed 2-carbonitriles or 2-carboxymethyl esters showed improved potency in sensitizing K562-resistant cells to imatinib treatment, even at concentrations, which are considered patient-relevant. At 5 μM, for instance, 2d sensitized the cells in such a manner that the resistance was fully overcome and the recovered efficacy of imatinib resulted in >76% cell death. Importantly, all compounds were non-cytotoxic per se. A transactivation experiment showed that only the carbonitriles are partial agonists of PPARγ, which does not seem to be involved in the mode of action. Yet, immunoassays revealed a suppression of the STAT5 phosphorylation status by co-application of the most active derivatives with imatinib. This mechanism consequently resulted in reduced cell proliferation and induction of cell death in resistant CML cells., 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 © 2021 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)

Epstein-Barr virus (EBV) is the first identified human tumor-related DNA virus, and has a high infection among people worldwide. Recent studies have showed that nearly 10% of gastric cancers have shown EBV infection and this kind of gastric cancer has been identified as a new subtype: EBV associated Gastric cancer (EBVaGC). Furthermore, it has been reported that tumor related genes in the EBVaGC showed frequent methylation modifications compared to those in the EBV negative gastric cancer (EBVnGC). To fully understand the role of EBV in EBVaGC, we analyzed and found that 16.67% of gastric carcinoma samples showed positive EBER1 signals. Mechanically, EBV-encoded Latent membrane protein 1 (LMP1) inhibited the expression of RASSF10, and promoted tumorigenesis by recruiting DNMT1 and inducing the DNA methylation of RASSF10. Altogether, it allows us a better understanding of the possible mechanism of EBV-induced gene hypermethylation in gastric cancer genome. Targeting EBV-induced DNA methylation is a potential therapeutic modality of EBVaGC., Competing Interests: Declaration of competing interest The authors declare no potential conflicts of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Atopic Dermatitis is an inflammatory skin disease associated with broad defects in skin barrier function caused by increased levels of type-2 cytokines (IL-4 and IL-13) that repress keratinocyte (KC) differentiation. Although crucial in mediating allergic disease, the mechanisms for gene repression induced by type-2 cytokines remain unclear. In this study, we determined that gene repression requires the master regulator of the epidermal differentiation program, p63. We found that type-2 cytokine-mediated inhibition of the expression of genes involved in early KC differentiation, including keratin 1, keratin 10, and DSC-1, is reversed by p63 blockade. Type-2 cytokines, through p63, also regulate additional genes involved in KC differentiation, including CHAC-1, STC2, and CALML5. The regulation of the expression of these genes is ablated by p63 small interfering RNA as well. In addition, we found that IL-4 and IL-13 and Staphylococcus aureus lipoteichoic acid work in combination through p63 to further suppress the early KC differentiation program. Finally, we found that IL-4 and IL-13 also inhibit the activity of Notch, a transcription factor required to induce early KC differentiation. In conclusion, type-2 cytokine-mediated gene repression and blockade of KC differentiation are multifactorial, involving pathways that converge on transcription factors critical for epidermal development, p63 and Notch., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Gallbladder carcinoma (GBC) is a considerable challenge because of its high metastatic potential. The tumor microenvironment is characterized by nutrient starvation, which promotes tumor metastasis. Stathmin1, an important microtubuleregulating protein, is overexpressed and promotes metastasis in GBC. It remains unclear how the harsh tumor microenvironment regulates stathmin1 expression to affect GBC metastasis. We employed glucose deficiency to mimic nutrient starvation and found that glucose deficiency upregulated stathmin1 transcription. However, glucose deficiency also promoted p27 degradation. There was a significant negative correlation between stathmin1 and p27 protein levels under glucose deficiency. Further study revealed that, under glucose deficiency, human kinase interacting with stathmin (hKIS) induced phosphorylation at Ser10 of p27 and its translocation to the cytoplasm for degradation, which upregulated the transcription factor E2F1 to promote stathmin1 transcription. hKIS knockdown significantly inhibited p27 cytoplasmic translocation and its consequent degradation. Stathmin1 knockdown significantly inhibited GBC cell migration and invasion in vitro. Our study revealed the role of the hKIS/p27/E2F1 axis in upregulating stathmin1 transcription to promote GBC cell migration and invasion under glucose deficiency conditions., (Copyright © 2021. Published by Elsevier Inc.)

Signal transducers and activators of transcription 5A and 5B (STAT5A and STAT5B) are two closely related STAT family members that are crucial downstream effectors of tyrosine kinase oncoproteins such as FLT3-ITD in acute myeloid leukemia (AML) and BCR-ABL in chronic myeloid leukemia (CML). We recently developed and reported the synthesis of a first molecule called 17 f that selectively inhibits STAT5 signaling in myeloid leukemia cells and overcomes their resistance to chemotherapeutic agents. To improve the antileukemic effect of 17 f, we synthesized ten analogs of this molecule and analyzed their impact on cell growth, survival, chemoresistance and STAT5 signaling. Two compounds, 7 a and 7 a', were identified as having similar or higher antileukemic effects in various AML and CML cell lines. Both molecules were found to be more effective than 17 f at inhibiting STAT5 activity/expression and suppressing the chemoresistance of CML., (© 2020 Wiley-VCH GmbH.)

Background: Glioblastoma (GB) remains an incurable and deadly brain malignancy that often proves resistant to upfront treatment with temozolomide. Nevertheless, temozolomide remains the most commonly prescribed FDA-approved chemotherapy for GB. The DNA repair protein methylguanine-DNA methyl transferase (MGMT) confers resistance to temozolomide. Unsurprisingly temozolomide-resistant tumors tend to possess elevated MGMT protein levels or lack inhibitory MGMT promotor methylation. In this study, cultured human temozolomide resistance GB (43RG) cells were introduced to the MGMT inhibitor O 6 -benzylguanine combined with temozolomide and either LY2835219 (CDK 4/6 inhibitor) or LY2157299 (TGF-βRI inhibitor) seeking to overcome GB treatment resistance., Methods: Treatment effects were assessed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, western blot, cell viability, and cell cycle progression., Results: Our in vitro study demonstrated that sequential treatment of O 6 -Benzylguanine with either LY2385219 or LY2157299-enhanced temozolomide enhanced sensitivity in MGMT+ 43RG cells. Importantly, normal human neurons and astrocytes remained impervious to the drug therapies under these conditions. Furthermore, LY2835219 has additional anti-proliferative effects on cell cycling, including induction of an RB-associated G (1) arrest via suppression of cyclin D-CDK4/6-Rb pathway. LY2157299 enhances anti-tumor effect by disrupting TGF-β-dependent HIF-1α signaling and by activating both Smad and PI3K-AKT pathways towards transcription of S/G2 checkpoints., Conclusion: This study establishes the groundwork for the development of a combinatorial pharmacologic approach by using either LY2385219 or LY2157299 inhibitor plus O 6 -Benzylguanine to augment temozolomide response in temozolomide-resistant GB cells.

Regulation of sphingolipid metabolism plays a role in cellular homeostasis, and dysregulation of these pathways is involved in cancer progression. Previously, our reports identified ceramide as an anti-metastatic lipid. In the present study, we investigated the biochemical alterations in ceramide-centered metabolism of sphingolipids that were associated with metastatic potential. We established metastasis-prone sublines of SKOV3 ovarian cancer cells using an in vivo selection method. These cells showed decreases in ceramide levels and ceramide synthase (CerS) 2 expression. Moreover, CerS2 downregulation in ovarian cancer cells promoted metastasis in vivo and potentiated cell motility and invasiveness. Moreover, CerS2 knock-in suppressed the formation of lamellipodia required for cell motility in this cell line. In order to define specific roles of ceramide species in cell motility controlled by CerS2, the effect of exogenous long- and very long-chain ceramide species on the formation of lamellipodia was evaluated. Treatment with distinct ceramides increased cellular ceramides and had inhibitory effects on the formation of lamellipodia. Interestingly, blocking the recycling pathway of ceramides by a CerS inhibitor was ineffective in the suppression of exogenous C 24:1 -ceramide for the formation of lamellipodia. These results suggested that C 24:1 -ceramide, a CerS2 metabolite, predominantly suppresses the formation of lamellipodia without the requirement for deacylation/reacylation. Moreover, knockdown of neutral ceramidase suppressed the formation of lamellipodia concomitant with upregulation of C 24:1 -ceramide. Collectively, the CerS2-C 24:1 -ceramide axis, which may be countered by neutral ceramidase, is suggested to limit cell motility and metastatic potential. These findings may provide insights that lead to further development of ceramide-based therapy and biomarkers for metastatic ovarian cancer., (© 2021 Federation of American Societies for Experimental Biology.)

Purpose: Glioma, especially glioblastoma (GBM), is the most aggressive malignant brain tumor and its standard therapy is often ineffective because of temozolomide (TMZ) resistance. Reversal of the TMZ resistance might improve the prognosis of glioma patients. We previously found that interferon-α (IFN-α) and anti-epileptic drug levetiracetam (LEV) could sensitize glioma to TMZ, respectively. In this study, we further investigated the efficiency of combining of LEV and IFN-α for improving the efficacy of TMZ., Methods: We evaluated whether LEV and IFN-α could increase TMZ efficacy using colony formation assay and cell viability assay with MGMT-positive and MGMT-negative glioma cell lines in vitro. Subcutaneous xenografts and orthotopic xenografts mice models were used in vivo to observe the tumor growth and mice survival upon treatments with TMZ, TMZ + IFN-α, TMZ + LEV, or TMZ + LEV + IFN-α. The expression levels of MGMT, markers of pro-apoptotic and anti-apoptotic in tumor samples were analyzed by Western blotting., Results: The combinational use of IFN-α, LEV, and TMZ showed the best anti-tumor activity in MGMT-positive cell lines (U138, GSC-1, U118, and T98 G). TMZ + LEV + IFN-α further obviously increased TMZ + LEV or TMZ + IFN-α efficiency in MGMT-positive cell lines, while not in negative cell lines (SKMG-4, U87, U373, and U251) in vitro, which were also observed in subcutaneous mice models (U138, GSC-1 compared to SKMG-4, U87) and orthotopic models (GSC-1) in vivo. Strikingly, the combination of LEV and IFN-α together with TMZ significantly prolonged the survival of mice with orthotopic GSC-1 glioma. Furthermore, we confirmed that the combination of LEV and IFN-α enhanced the inhibition of MGMT and the activation of apoptosis in U138 tumor on the basis of TMZ treatment., Conclusions: The combination use of LEV and IFN-α could be an optimal method to overcome TMZ resistance through obvious MGMT inhibition in MGMT-positive glioma.

A novel 2-thiopyrimidine/chalcone hybrid was designed, synthesised, and evaluated for their cytotoxic activities against three different cell lines, K-562, MCF-7, and HT-29. The most active cytotoxic derivatives were 9d , 9f , 9n , and 9p (IC 50 =0.77-1.74 µM, against K-562 cell line), 9a and 9r (IC 50 =1.37-3.56 µM against MCF-7 cell line), and 9a , 9l , and 9n (IC 50 =2.10 and 2.37 µM against HT-29 cell line). Compounds 9a , 9d , 9f , 9n , and 9r were further evaluated for their cytotoxicity against normal fibroblast cell line WI38. Moreover, STAT3 and STAT5a inhibitory activities were determined for the most active derivatives 9a , 9d , 9f , 9n , and 9r . Dual inhibitory activity was observed in compound 9n (IC 50 =113.31 and 50.75 µM, against STAT3 and STAT5a, respectively). Prediction of physicochemical properties, drug likeness score, pharmacokinetic and toxic properties was detected.

In pancreatic cancer, the activities of inhibitory agents were investigated using docking, since the inhibition of TP63, which plays an important role in the spread of cancer with metastasis, in preventing the proliferation and proliferation of this type of cancer. It has been shown that the active ingredients in some plants used as traditional medicines have an inhibitory effect on this cancer type in preventing growth, reproduction and spread. These computational results guide experimental studies, preventing time and item loss; It is an important study in terms of choosing and using the right active substances.

Spleen tyrosine kinase (SYK) is an important oncogene and signaling mediator activated by cell surface receptors crucial for acute myeloid leukemia (AML) maintenance and progression. Genetic or pharmacologic inhibition of SYK in AML cells leads to increased differentiation, reduced proliferation, and cellular apoptosis. Herein, we addressed the consequences of SYK inhibition to leukemia stem-cell (LSC) function and assessed SYK-associated pathways in AML cell biology. Using gain-of-function MEK kinase mutant and constitutively active STAT5A, we demonstrate that R406, the active metabolite of a small-molecule SYK inhibitor fostamatinib, induces differentiation and blocks clonogenic potential of AML cells through the MEK/ERK1/2 pathway and STAT5A transcription factor, respectively. Pharmacological inhibition of SYK with R406 reduced LSC compartment defined as CD34 + CD38 - CD123 + and CD34 + CD38 - CD25 + in vitro, and decreased viability of LSCs identified by a low abundance of reactive oxygen species. Primary leukemic blasts treated ex vivo with R406 exhibited lower engraftment potential when xenotransplanted to immunodeficient NSG/J mice. Mechanistically, these effects are mediated by disturbed mitochondrial biogenesis and suppression of oxidative metabolism (OXPHOS) in LSCs. These mechanisms appear to be partially dependent on inhibition of STAT5 and its target gene MYC, a well-defined inducer of mitochondrial biogenesis. In addition, inhibition of SYK increases the sensitivity of LSCs to cytarabine (AraC), a standard of AML induction therapy. Taken together, our findings indicate that SYK fosters OXPHOS and participates in metabolic reprogramming of AML LSCs in a mechanism that at least partially involves STAT5, and that SYK inhibition targets LSCs in AML. Since active SYK is expressed in a majority of AML patients and confers inferior prognosis, the combination of SYK inhibitors with standard chemotherapeutics such as AraC constitutes a new therapeutic modality that should be evaluated in future clinical trials.

Pancreatic cancer is a malignant cancer with poor prognosis. This study aimed to explore how O6-methylguanine-DNA methyltransferase (MGMT) affects the gemcitabine resistance of pancreatic cancer cells by the regulatory role of SHH/GLI signaling pathway. MGMT inhibition induced by lomeguatrib (LM) suppressed the proliferation, invasion, migration and autophagy, promoted the apoptosis of PanC-1/GEM cells and up-regulated the GEM inhibition rates for PanC-1/GEM cells. Moreover, MGMT inhibition increased the expression of Caspase-3 and Bax and decreased the expression of Bcl-2, Beclin1 and Atg5 in PanC-1/GEM cells. PVT1 silencing could also produce the similar effects of MGMT inhibition induced by LM on PanC-1/GEM cells. And, PVT1 silencing could inhibit the SHH/GLI signaling pathway in PanC-1/GEM cells by regulating the MGMT expression. miR-409 was demonstrated to be a potential target of PVT1 and SHH was demonstrated to be a potential target of miR-409. Furthermore, GLI overexpression could reverse the effects of PVT1 silencing. In the xenograft model of pancreatic cancer, nude mice were treated with GEM. MGMT inhibition suppressed the tumor growth and autophagy and promoted the apoptosis in tumor tissues. And, PVT1 silencing could inhibit the SHH/GLI signaling pathway in tumor tissues. In conclusion, MGMT inhibition could suppress the proliferation, invasion, migration and autophagy and promote the apoptosis of PanC-1/GEM cells in vitro and in vivo. PVT1 silencing may affect the PanC-1/GEM cells through changing the MGMT expression by inhibiting the SHH/GLI signaling pathway., Competing Interests: Declaration of competing interest The authors declare they have no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

p62/SQSTM1, encoded by gene SQSTM1 , is widely known as an adaptor protein of selective autophagy to promote aggregate-prone proteins for degradation. It is also a stress-induced scaffold protein involved in Nrf2 activation to resist oxidative stress. Multiple domains of p62 interact with several essential pathways implicated in cell differentiation and proliferation, placing p62 at a significant position to mediate cell survival and apoptosis. The p62 protein has been suggested as a potential target in recent years, since its abnormal expression or SQSTM1 gene mutation is tightly associated with various diseases including cancer such as hepatocellular carcinoma and prostate cancer, neurodegenerative disorders such as Alzheimer's disease and amyotrophic lateral sclerosis, atherosclerosis, and Paget's disease of bone. In this review, we will discuss the relationship between p62 and these diseases, and we attempt to put forward novel methods for current diagnosis or therapy by regulating the p62 expression level.

The inhibition of the DNA damage response (DDR) pathway in the treatment of cancer has recently gained interest, and different DDR inhibitors have been developed. Among them, the most promising ones target the WEE1 kinase family, which has a crucial role in cell cycle regulation and DNA damage identification and repair in both nonmalignant and cancer cells. This review recapitulates and discusses the most recent findings on the biological function of WEE1/PKMYT1 during the cell cycle and in the DNA damage repair, with a focus on their dual role as tumor suppressors in nonmalignant cells and pseudo-oncogenes in cancer cells. We here report the available data on the molecular and functional alterations of WEE1/PKMYT1 kinases in both hematological and solid tumors. Moreover, we summarize the preclinical information on 36 chemo/radiotherapy agents, and in particular their effect on cell cycle checkpoints and on the cellular WEE1/PKMYT1-dependent response. Finally, this review outlines the most important pre-clinical and clinical data available on the efficacy of WEE1/PKMYT1 inhibitors in monotherapy and in combination with chemo/radiotherapy agents or with other selective inhibitors currently used or under evaluation for the treatment of cancer patients.

The Hippo pathway regulates myriad biological processes in diverse species and is a key cancer signaling network in humans. Although Hippo has been linked to multiple aspects of cancer, its role in this disease is incompletely understood. Large-scale pan-cancer analyses of core Hippo pathway genes reveal that the pathway is mutated at a high frequency only in select human cancers, including malignant mesothelioma and meningioma. Hippo pathway deregulation is also enriched in squamous epithelial cancers. We discuss cancer-related functions of the Hippo pathway and potential explanations for the cancer-restricted mutation profile of core Hippo pathway genes. Greater understanding of Hippo pathway deregulation in cancers will be essential to guide the imminent use of Hippo-targeted therapies., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Glioblastoma is the most common primary tumor of the central nervous system that develops chemotherapy resistance. Previous studies showed that Allicin could inhibit multiple cancer cells including glioblastoma, but the function of Allicin in glioblastoma is still unclear. Our work aimed to investigate the underlying molecular mechanism. The results showed that miR-486-3p levels were greatly increased in glioblastoma during Allicin treatment. Overexpression of miR-486-3p increased chemosensitivity to temozolomide (TMZ) in vitro and in vivo. O6-methylguanine-DNA methyltransferase (MGMT) was identified as a direct target of miR-486-3p, and miR-486-3p overexpression prevented the protein translation of MGMT. Moreover, overexpression of MGMT restored miR-486-3p-induced chemosensitivity to TMZ. Taken together, our studies revealed that Allicin could upregulate miR-486-3p and enhance TMZ sensitivity in glioblastoma. The results suggested that in the future, Allicin can be used as an adjuvant therapy with TMZ to improve the prognosis of patients, and miR-486-3p may be a potential target for glioblastoma treatment to improve the curative effects.

We recently reported, for the first time, the expression and regulation of the PDZ polarity proteins Scrib and Dlg1 in human APCs, and also described the viral targeting of these proteins by NS1 of influenza A virus in human dendritic cells (DCs). Scrib plays an important role in reactive oxygen species (ROS) production in Mϕs and uropod formation and migration in T cells, while Dlg1 is important for T cell downstream activation after Ag recognition. Nevertheless, the functions of these proteins in human DCs remain unknown. Here, we knocked-down the expression of both Scrib and Dlg1 in human DCs and then evaluated the expression of co-stimulatory molecules and cytokine production during maturation. We demonstrated that Scrib is necessary for adequate CD86 expression, while Dlg1 is important for CD83 up-regulation and IL-6 production upon maturation, suggesting that Scrib and Dlg1 participate in separate pathways in DCs. Additionally, both proteins are required for adequate IL-12 production after maturation. Furthermore, we showed that the inefficient maturation of DCs induced by Scrib or Dlg1 depletion leads to impaired T cell activation. Our results revealed the previously unknown contribution of Scrib and Dlg1 in human DCs pivotal functions, which may be able to impact innate and adaptive immune response., (©2020 Society for Leukocyte Biology.)

HUWE1 is an E3 ubiquitin ligase that is involved in cancer cell proliferation by regulating MCL-1 stability. The HECT domain has been shown to be required for the ubiquitin ligase activity of HUWE1. To identify efficient drugs that impair the activity of HUWE1, and thus decrease MCL-1 accumulation, we screened 2000 candidate compounds that might suppress HUWE1 activity. To evaluate these 2000 candidates, the HECT domain of HUWE1, which is the catalytic domain responsible for MCL1 ubiquitination, was selected as a conjugation site, and putative binding candidates were filtrated. Tobramycin emerged as one of the compounds that show efficient binding ability with the HECT domain of HUWE1. The surface plasmon resonance (SPR) results validated the specific binding of Tobramycin with the HECT domain. Subsequent analyses demonstrated its potential to inhibit cancer cell proliferation by binding to the HECT domain of HUWE1 and impeding the HUWE1-mediated ubiquitination of MCL-1. Consequently, the accumulation of MCL-1 inhibited the proliferation of tumour cells, while the apoptosis rates were not significantly altered after Tobramycin treatment. In vitro experiments showed that Tobramycin could inhibit cell proliferation by regulating the G2/M transition in cancer cell models, including A549 and HeLaCaco2 cell lines. Our results indicated that Tobramycin could be a potential new probe to develop targeted therapies for the prevention or treatment of HUWE1-overexpressing cancers., (© 2020 John Wiley & Sons Australia, Ltd.)

N 6 -methyladenosine (m 6 A) is the most abundant mRNA modification and is catalyzed by the methyltransferase complex, in which methyltransferase-like 3 (METTL3) is the sole catalytic subunit. Accumulating evidence in recent years reveals that METTL3 plays key roles in a variety of cancer types, either dependent or independent on its m 6 A RNA methyltransferase activity. While the roles of m 6 A modifications in cancer have been extensively reviewed elsewhere, the critical functions of METTL3 in various types of cancer, as well as the potential targeting of METTL3 as cancer treatment, have not yet been highlighted. Here we summarize our current understanding both on the oncogenic and tumor-suppressive functions of METTL3, as well as the underlying molecular mechanisms. The well-documented protein structure of the METTL3/METTL14 heterodimer provides the basis for potential therapeutic targeting, which is also discussed in this review.

Renal cell carcinoma (RCC) is one of the most common malignancies worldwide, and metabolic reprogramming has a profound effect on RCC tumorigenesis. mTORC1 inhibitors are widely used in RCC treatment, yet some types of RCC cells are resistant to these compounds. Thus, clarification of the metabolic mechanism of mTORC1 inhibitors and exploration of new therapeutic approaches are urgently needed. In this study, we found that the mTORC1 pathway was hyperactive in RCC. Immunohistochemistry and western blot analysis showed that phosphorylation of the mTORC1 substrate 4EBP1 at threonine 37/46 increased in RCC tissues compared with that in normal renal tissues. It was also found that mTORC1 inhibitor everolimus suppressed glucose consumption, lactate production, and multiple catalytic enzymes involved in glycolysis in 786-O and ACHN cells, but the accumulation of HIF1α induced by CoCl 2 blocked the inhibitory effect of everolimus on aerobic glycolysis. Interestingly, western blot and metabolite analysis showed that the tumor suppressor NDRG2 (N-Myc downstream regulated gene 2) was able to inhibit mTORC1 activity and cooperate with an mTOR inhibitor to decrease aerobic glycolysis in 786-O and ACHN cells. These results demonstrate that NDRG2 may potentially synergize with mTORC1 inhibitors to suppress malignant phenotype of RCC. Taken together, these data provided preclinical evidence that the combination of NDRG2 and mTORC1 inhibitors might be a promising strategy for RCC therapy.

Proteasome inhibitors have provided a significant advance in the treatment of multiple myeloma (MM). Consequently, there is increasing interest in developing strategies to target E3 ligases, de-ubiquitinases, and/or ubiquitin receptors within the ubiquitin proteasome pathway, with an aim to achieve more specificity and reduced side-effects. Previous studies have shown a role for the E3 ligase HUWE1 in modulating c-MYC, an oncogene frequently dysregulated in MM. Here we investigated HUWE1 in MM. We identified elevated expression of HUWE1 in MM compared with normal cells. Small molecule-mediated inhibition of HUWE1 resulted in growth arrest of MM cell lines without significantly effecting the growth of normal bone marrow cells, suggesting a favorable therapeutic index. Studies using a HUWE1 knockdown model showed similar growth inhibition. HUWE1 expression positively correlated with MYC expression in MM bone marrow cells and correspondingly, genetic knockdown and biochemical inhibition of HUWE1 reduced MYC expression in MM cell lines. Proteomic identification of HUWE1 substrates revealed a strong association of HUWE1 with metabolic processes in MM cells. Intracellular glutamine levels are decreased in the absence of HUWE1 and may contribute to MYC degradation. Finally, HUWE1 depletion in combination with lenalidomide resulted in synergistic anti-MM activity in both in vitro and in vivo models. Taken together, our data demonstrate an important role of HUWE1 in MM cell growth and provides preclinical rationale for therapeutic strategies targeting HUWE1 in MM.

Background: The resistance of lung cancer to epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) is one of the unconquered frontiers in chemotherapy. Mitogen-inducible gene 6 (Mig-6) is known to inhibit the kinase activity of epidermal growth factor receptor (EGFR). Similarly, numerous studies of mouse models suggested tumor suppressive function of Mig-6 in lung cancer. On the contrary, the results of clinical investigations revealed that lung cancer patients with elevated expression of Mig-6 are associated with a poor prognosis. More recent work showed that unlike wild type (WT) EGFR, mutant EGFR phosphorylates Mig-6 and phosphorylated Mig-6 negatively regulates the degradation of EGFR mutants in lung adenocarcinoma. Here, we tried to untangle the controversies surrounding Mig-6 function as a protagonist or an antagonist of EGFR-TKI resistant lung cancer., Methods: We compared the expression and phosphorylation status of Mig-6 in the EGFR-TKI resistant lung adenocarcinoma (PC9/GR cells) to EGFR-TKI sensitive lung adenocarcinoma (PC9 cells). We investigated the function of Mig-6 by either depletion or overexpression of Mig-6 in those cells and evaluated the efficacy of combining of Mig-6 knock-down and EGFR-TKI treatment in PC9/GR. The correlation between Mig-6 expressions and the prognoses of lung adenocarcinoma was examined by The Cancer Genome Atlas (TCGA) data and clinical samples., Results: Our results indicated that the expression of Mig-6 was significantly increased in PC9/GR cells compared to that of PC9 cells. The significant portion of Mig-6 existed as a phosphorylated form in PC9 and PC9/GR cells. Moreover, overexpression of Mig-6 significantly increased the cell proliferation, invasion and epithelial mesenchymal transition (EMT) in PC9 cells. Combination of Mig-6 knock-down and EGFR-TKI treatment significantly overcame the EGFR-TKI resistance of PC9/GR cells. In addition, our analyses of clinical samples confirmed that high Mig-6 expressions positively correlate with a poor prognosis and EGFR-TKI resistance in lung adenocarcinoma., Conclusion: Our findings reinforce scientific notion of Mig-6 as an oncoprotein in the context of EGFR-TKI resistant lung adenocarcinoma. We propose that targeting Mig-6 may be a promising strategy to overcome the EGFR-TKI resistance in lung cancer.

Targeting the MAPK pathway by combined inhibition of BRAF and MEK has increased overall survival in advanced BRAF-mutant melanoma in both therapeutic and adjuvant clinical settings. However, a significant proportion of tumors develop acquired resistance, leading to treatment failure. We have previously shown p63 to be an important inhibitor of p53-induced apoptosis in melanoma following genotoxic drug exposure. Here, we investigated the role of p63 in acquired resistance to MAPK inhibition and show that p63 isoforms are upregulated in melanoma cell lines chronically exposed to BRAF and MEK inhibition, with consequent increased resistance to apoptosis. This p63 upregulation was the result of its reduced degradation by the E3 ubiquitin ligase FBXW7. FBXW7 was itself regulated by MDM2, and in therapy-resistant melanoma cell lines, nuclear accumulation of MDM2 caused downregulation of FBXW7 and consequent upregulation of p63. Consistent with this, both FBXW7-inactivating mutations and MDM2 upregulation were found in melanoma clinical samples. Treatment of MAPK inhibitor-resistant melanoma cells with MDM2 inhibitor Nutlin-3A restored FBXW7 expression and p63 degradation in a dose-dependent manner and sensitized these cells to apoptosis. Collectively, these data provide a compelling rationale for future investigation of Nutlin-3A as an approach to abrogate acquired resistance of melanoma to MAPK inhibitor targeted therapy. SIGNIFICANCE: Upregulation of p63, an unreported mechanism of MAPK inhibitor resistance in melanoma, can be abrogated by treatment with the MDM2 inhibitor Nutlin-3A, which may serve as a strategy to overcome resistance., (©2020 American Association for Cancer Research.)

Docetaxel-based chemotherapy is recommended for metastatic castration-resistant prostate cancer (mCRPC). However, chemoresistance is inevitable and eventually progresses after several rounds of chemotherapy. Therefore, exploration of new therapeutic targets and molecular mechanisms that contribute to chemoresistance remains necessary. Our previous study accidentally demonstrated that expression of nitrogen permease regulator-like 2 (NPRL2), which is defined as a tumor suppressor, is upregulated in prostate cancer (PCa) and linked to poor prognosis, particularly in CRPC. The aim of this study was to investigate the role of NPRL2 in the chemoresistant CRPC cells. We found that NPRL2 was significantly overexpressed in docetaxel-resistant CRPC cells, while autophagy was enhanced and mTOR signaling was inhibited. Inhibiting NPRL2 increased the sensitivity to docetaxel in docetaxel-resistant CRPC cells, enhanced apoptosis and inhibited autophagy, and the opposite trends were observed when the mTOR inhibitor torin 1 was added to NPRL2-silenced cells. We further found that NPRL2 silenced docetaxel-resistant CRPC cells were sensitive to docetaxel in vivo. Briefly, our research reveals that overexpression of NPRL2 promotes chemoresistance by regulating autophagy via mTOR signaling and inhibits apoptosis in CRPC cells., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: The adhesion molecule, FAT4, has a tumor suppressor function with a critical role in the epithelial-to-mesenchymal-transition (EMT) and anti-malignant growth in several cancers. No study has investigated yet its role in epithelial ovarian cancer (EOC) progression. In the present study, we examined the role of FAT4 in proliferation and metastasis, and its mechanisms of interaction in these processes., Methods: We have performed cell viability, colony formation, and invasion assays in ovarian cancer cells treated with siRNA to knockdown FAT4 gene expression. The regulatory effects of FAT4 on proteins involved in apoptotic, Wnt, Hippo, and retinoblastoma signaling pathways were evaluated by Western blotting following FAT4 repression. Also, 426 ovarian tumor samples and 88 non-tumor samples from the Gene Expression Profiling Interactive Analysis (GEPIA) database were analyzed for the expression of FAT4. Pearson's correlation was performed to determine the correlation between FAT4 and the E2F5, cyclin D1, cdk4, and caspase 9 expressions., Results: Lower expression of FAT4 was observed in ovarian cancer cell lines and human samples as compared to non-malignant tissues. This down-regulation seems to enhance cell viability, invasion, and colony formation. Silencing FAT4 resulted in the upregulation of E2F5, vimentin, YAP, β-catenin, cyclin D1, cdk4, and Bcl2, and in the downregulation of GSK-3-β, and caspase 9 when compared to control. Furthermore, regulatory effects of FAT4 on the EMT and aggressive phenotype seem to occur through Hippo, Wnt, and cell cycle pathways., Conclusion: FAT4 downregulation promotes increased growth and invasion through the activation of Hippo and Wnt-β-catenin pathways.

Sophoridine is a quinolizidine natural product and the exploration of its derivatives has been carried out, and the potent anticancer compound IMB-HDC was acquired. Although previous studies have revealed that some sophoridine derivatives could induce DNA breakage, the underlying mechanisms of inhibition of DNA damage repair (ATR inactivation) and the apoptosis independent of p53, have not been elucidated. Our research reveals a novel DNA response mechanism different from general DNA-damaging agents, and that sophoridine derivate inhibits the phosphorylation of Tyr694 and Ser780 of STAT5a to induce the lessened shuttle from the cytoplasm to the nucleus, and leads to the decreased nuclear STAT5a and subsequently inhibits the expression of STAT5a target gene RAD51 that contributes to the checkpoint activation, thus inhibiting ATR activation. Meanwhile, IMB-HDC that induced the diminished expression of STAT5a target gene contributes to proliferation and leads to apoptosis. More importantly, we give the first evidence that promoting the effect of Tyr694 phosphorylation on nuclear location and subsequent STAT5a target gene transcription depends on Ser780 increased or unchanged phosphorylation and was not correlated with Ser726 phosphorylation.

Cisplatin (DDP) is the first-line drug for the treatment of gastric cancer (GC). However, DDP resistance is common. Autophagy, which is closely related to chemoresistance, is a process of resolving and recycling proteins and damaged cellular organs. Additionally, O-6-methylguanine-DNA methyltransferase (MGMT) is responsible for alkylating drug resistance. However, the relationship between autophagy and MGMT in response to DDP in GC is still unknown. In the present study, we determined that autophagy induced by DDP decreases chemosensitivity in GC cell lines. DDP may have induced autophagy in GC by inhibiting MGMT to increase autophagy-related gene (ATG) 4B. Inhibition of MGMT-mediated ATG4B suppression resulted in autophagy induction and DDP resistance. In vivo, combined DDP and autophagy inhibitor chloroquine (CQ) enhanced the anti-tumor effect of DDP; additionally, the negative correlation of MGMT and ATG4B was confirmed. High expression of MGMT and low expression of ATG4B were significantly correlated with favorable five-year survival rate (P <  0.05) in 66 clinicopathologically characterized GC cases. Our study demonstrate that DDP inhibits MGMT-mediated autophagy suppression to decrease chemosensitivity in GC, which provides a novel therapeutic strategy to promote DDP chemosensitivity in GC., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2020 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)