Tafenoquine and its derivatives as inhibitors for the severe acute respiratory syndrome coronavirus 2.

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has severely affected human lives around the world as well as the global economy. Therefore, effective treatments against COVID-19 are urgently needed. Here, we screened a library containing Food and Drug Administration (FDA)-approved compounds to identify drugs that could target the SARS-CoV-2 main protease (Mpro), which is indispensable for viral protein maturation and regard as an important therapeutic target. We identified antimalarial drug tafenoquine (TFQ), which is approved for radical cure of Plasmodium vivax and malaria prophylaxis, as a top candidate to inhibit Mpro protease activity. The crystal structure of SARS-CoV-2 Mpro in complex with TFQ revealed that TFQ noncovalently bound to and reshaped the substrate-binding pocket of Mpro by altering the loop region (residues 139-144) near the catalytic Cys145, which could block the catalysis of its peptide substrates. We also found that TFQ inhibited human transmembrane protease serine 2 (TMPRSS2). Furthermore, one TFQ derivative, compound 7, showed a better therapeutic index than TFQ on TMPRSS2 and may therefore inhibit the infectibility of SARS-CoV-2, including that of several mutant variants. These results suggest new potential strategies to block infection of SARS-CoV-2 and rising variants.

A novel quinoline derivative, DFIQ, sensitizes NSCLC cells to ferroptosis by promoting oxidative stress accompanied by autophagic dysfunction and mitochondrial damage.

BACKGROUND: The development of nonapoptotic programmed cell death inducers as anticancer agents has emerged as a cancer therapy field. Ferroptosis, ferrous ion-driven programmed cell death that is induced by redox imbalance and dysfunctional reactive oxygen species (ROS) clearance, is triggered during sorafenib and PD-1/PD-L1 immunotherapy. DFIQ, a quinoline derivative, promotes apoptosis by disrupting autophagic flux and promoting ROS accumulation. Our pilot experiments suggest that DFIQ participates in ferroptosis sensitization. Thus, in this study, we aimed to reveal the mechanisms of DFIQ in ferroptosis sensitization and evaluate the clinical potential of DFIQ. METHODS: We treated the non-small cell lung cancer (NSCLC) cell lines H1299, A549, and H460 with the ferroptosis inducer (FI) DFIQ and analyzed viability, protein expression, ROS generation, and fluorescence staining at different time points. Colocalization analysis was performed with ImageJ. RESULTS: DFIQ sensitized cells to FIs such as erastin and RSL3, resulting in a decrease in IC50 of at least 0.5-fold. Measurement of ROS accumulation to explore the underlying mechanism indicated that DFIQ and FIs treatment promoted ROS accumulation and SOD1/SOD2 switching. Mitochondria, known ROS sources, produced high ROS levels during DFIQ/FI treatment. RSL3 treatment promoted mitochondrial damage and mitophagy, an autophagy-associated mitochondrial recycling system, and cotreatment with DFIQ induced accumulation of mitochondrial proteins, which indicated disruption of mitophagic flux. Thus, autophagic flux was measured in cells cotreated with DFIQ. DFIQ treatment was found to disrupt autophagic flux, leading to accumulation of damaged mitochondria and eventually inducing ferroptosis. Furthermore, the influence of DFIQ on the effects of clinical FIs, such as sorafenib, was evaluated, and DFIQ was discovered to sensitize NSCLC cells to sorafenib and promote ferroptosis. CONCLUSIONS: This study indicates that DFIQ not only promotes NSCLC apoptosis but also sensitizes cells to ferroptosis by disrupting autophagic flux, leading to accumulation of dysfunctional mitochondria and thus to ferroptosis. Ferroptosis is a novel therapeutic target in cancer therapy. DFIQ shows the potential to enhance the effects of FIs in NSCLC and act as a potential therapeutic adjuvant in ferroptosis-mediated therapy.

Crystal structure and functional implication of a bacterial cyclic AMP-AMP-GMP synthetase.

Mammalian cyclic GMP-AMP synthase (cGAS) and its homologue dinucleotide cyclase in Vibrio cholerae (VcDncV) produce cyclic dinucleotides (CDNs) that participate in the defense against viral infection. Recently, scores of new cGAS/DncV-like nucleotidyltransferases (CD-NTases) were discovered, which produce various CDNs and cyclic trinucleotides (CTNs) as second messengers. Here, we present the crystal structures of EcCdnD, a CD-NTase from Enterobacter cloacae that produces cyclic AMP-AMP-GMP, in its apo-form and in complex with ATP, ADP and AMPcPP, an ATP analogue. Despite the similar overall architecture, the protein shows significant structural variations from other CD-NTases. Adjacent to the donor substrate, another nucleotide is bound to the acceptor binding site by a non-productive mode. Isothermal titration calorimetry results also suggest the presence of two ATP binding sites. GTP alone does not bind to EcCdnD, which however binds to pppApG, a possible intermediate. The enzyme is active on ATP or a mixture of ATP and GTP, and the best metal cofactor is Mg2+. The conserved residues Asp69 and Asp71 are essential for catalysis, as indicated by the loss of activity in the mutants. Based on structural analysis and comparison with VcDncV and RNA polymerase, a tentative catalytic pathway for the CTN-producing EcCdnD is proposed.

Synthesis and Anticancer Evaluation of 4-Anilinoquinolinylchalcone Derivatives.

A series of 4-anilinoquinolinylchalcone derivatives were synthesized and evaluated for antiproliferative activities against the growth of human cancer cell lines (Huh-7 and MDA-MB-231) and normal lung cells (MRC-5). The results exhibited low cytotoxicity against human lung cells (MRC-5). Among them, (E)-3-{4-{[4-(benzyloxy)phenyl]amino}quinolin-2-yl}-1-(4-methoxyphenyl) prop-2-en-1-one (4a) was found to have the highest cytotoxicity in breast cancer cells and low cytotoxicity in normal cells. Compound 4a causes ATP depletion and apoptosis of breast cancer MDA-MB-231 cells and triggers reactive oxygen species (ROS)-dependent caspase 3/7 activation. In conclusion, it is worth studying 4-anilinoquinolinylchalcone derivatives further as new potential anticancer agents for the treatment of human cancers.

Individual and Neighborhood Characteristics Associated With Confirmatory and Follow-up Blood Lead Testing Among Children With Elevated Blood Lead Levels.

OBJECTIVES: The aim of this study was to assess the proportions and likelihood of children who receive confirmatory and follow-up blood lead testing within the recommended time frames after an initial capillary elevated blood lead level (EBLL) and confirmed EBLL, respectively, by individual and neighborhood-level sociodemographic characteristics. DESIGN: We linked and used blood testing and sociodemographic characteristics data from a Pennsylvania birth cohort including children born between 2017 and 2018. Generalized linear mixed models were constructed to examine the associations between sociodemographic factors and having recommended confirmatory and follow-up testing. SETTING: A population-based, retrospective cohort study. PARTICIPANTS: In this birth cohort, children who underwent at least 1 BLL test were followed up to 24 months of age. Children with a first unconfirmed (n = 6259) and confirmed BLL (n = 4213) ≥ 5 µg/dL were included in the analysis. MAIN OUTCOME MEASURE: Children had confirmatory and follow-up testing within the recommended time frames. RESULTS: Of the children with unconfirmed and confirmed EBLLs, 3555 (56.8%) and 1298 (30.8%) received confirmatory and follow-up testing, respectively. The proportions of the 2 outcome measures were lower among children experiencing certain sociodemographic disadvantages. In the univariate analyses, lower initial BLLs, older age, non-Hispanic Blacks, lower maternal educational levels, maternal Medicaid, The Special Supplemental Nutrition Program for Women, Infants, and Children (WIC) enrollment, maternal smoking, and higher quartiles of neighborhood poverty and old housing were associated with lower odds of having confirmatory and follow-up testing. However, in multivariate models, children with lower initial BLLs, older age, maternal smoking, and non-Hispanic Blacks were significantly less likely to have confirmatory and follow-up testing. CONCLUSIONS: There were deficiencies in having recommended confirmatory and follow-up blood lead testing among children, especially those with sociodemographic disadvantages. Public health agencies and stakeholders should finetune policies to improve follow-up testing in conjunction with primary and secondary preventions for early detection and reduction of lead exposure among targeted children at risk of lead poisoning.

The Wnt1-Cre2 transgene is active in the male germline.

The Wnt1-Cre transgenic mouse line is widely used to express the CRE recombinase in neural crest lineages, but it overexpresses WNT1 itself, which can cause undesired phenotypes. To address this, we and others previously developed a Wnt1-Cre2 line based on the same regulatory elements as Wnt1-Cre but without ectopic Wnt1 expression. However, while Wnt1-Cre2 exhibits normal activity when transmitted from female mice, it exhibits unexpected activity in the male germline. The Wnt1-Cre2 transgene was previously mapped to the E2f1 locus. Several genes in this genomic region exhibit significant expression in spermatogonia or spermatocytes, suggesting that local regulatory elements may be driving ectopic transgene expression. The Wnt1-Cre2 line can therefore be used both as a neural crest specific and a general deleter, and care should be taken when setting up genetic crosses.

Inhibition of gut microbial ß-glucuronidase effectively prevents carcinogen-induced microbial dysbiosis and intestinal tumorigenesis.

The bidirectional interaction between carcinogens and gut microbiota that contributes to colorectal cancer is complicated. Reactivation of carcinogen metabolites by microbial ß-glucuronidase (ßG) in the gut potentially plays an important role in colorectal carcinogenesis. We assessed the chemoprotective effects and associated changes in gut microbiota induced by pre-administration of bacterial-specific ßG inhibitor TCH-3511 in carcinogen azoxymethane (AOM)-treated APCMin/+ mice. AOM induced intestinal ßG activity, which was reflected in increases in the incidence, formation, and number of tumors in the intestine. Notably, inhibition of gut microbial ßG by TCH-3511 significantly reduced AOM-induced intestinal ßG activity, decreased the number of polyps in both the small and large intestine to a frequency that was similar in mice without AOM exposure. AOM also led to lower diversity and altered composition in the gut microbiota with a significant increase in mucin-degrading Akkermansia genus. Conversely, mice treated with TCH-3511 and AOM exhibited a more similar gut microbiota structure as mice without AOM administration. Importantly, TCH-3511 treatment significant decreased Akkermansia genus and produced a concomitant increase in short-chain fatty acid butyrate-producing gut commensal microbes Lachnoospiraceae NK4A136 group genus in AOM-treated mice. Taken together, our results reveal a key role of gut microbial ßG in promoting AOM-induced gut microbial dysbiosis and intestinal tumorigenesis, indicating the chemoprotective benefit of gut microbial ßG inhibition against carcinogens via maintaining the gut microbiota balance and preventing cancer-associated gut microbial dysbiosis. Thus, the bacterial-specific ßG inhibitor TCH-3511 is a potential chemoprevention agent for colorectal cancer.

Investigation of Toxoplasma infection in zoo animals using multispecies ELISA and GRA7 nested PCR.

BACKGROUND: Toxoplasma is an obligate intracellular protozoan that causes an important zoonotic disease with a worldwide distribution. Felids are the definitive hosts of this parasite, while virtually all warm-blooded animals, including birds, serve as intermediate hosts. Four ring-tailed lemurs (Lemur catta) in the Taipei Zoo died of acute Toxoplasma infection in June 2019. Since then, Toxoplasma has occasionally been identified in this Zoo during necropsy of dead animals and PCR of animal blood samples. Therefore, a general survey of Toxoplasma infection in animals in the Zoo seems to be needed. METHODS AND RESULTS: An indirect multispecies ELISA was used for the first time to screen for Toxoplasma infection in 326 serum samples collected from 75 species of animals. The infection rate of Toxoplasma was 27% (88/326). A commercial latex agglutination (LAT) assay was used to re-examine the samples with doubtful and uncertain ELISA results (151 samples from 42 species). The infection rate increased to 36.2% (118/326), and the indirect multispecies ELISA appeared to be applicable to 31 of 75 species animals included in this study. Nested PCR assays targeting the dense granule protein 7 (GRA7) gene and B1 gene were also used to detect Toxoplasma in DNA samples extracted from 10 liver or blood specimens from 8 animals. GRA7 gene fragments were amplified from 8 samples from 7 animals, while B1 gene fragments were amplified from only 4 samples from 4 animals. From the B1 nested PCR and the sequence data of GRA7 fragments amplified from infectious specimens, the animals in the Zoo were speculated to have been infected by at least three different Toxoplasma variants. CONCLUSIONS: According to the serological investigation, we speculated that over one-third (36.2%) of animals in Taipei Zoo presented the infection of Toxoplasma, and the indirect multispecies ELISA we used can be applied to detect Toxoplasma infection in 31 animal species included in this study. Sequence analysis revealed that at least three Toxoplasma variants were infecting the animals of Taipei Zoo.

Consideration of the adaptive randomization allocation ratio in the presence of treatment group heteroscedasticity in clinical trials.

For randomized clinical trials, subjects' variance structures may vary over time among treatment groups, resulting in the heteroscedasticity of residuals in a regression analysis. Commonly used methods that assume equal variance among all treatment groups may not be able to control for a type I error. When the variances are indeed the same across treatment groups, an equal randomization allocation ratio will yield the greatest study power. However, out of ethical concern or urgent need for rare disease clinical trials, more patients may have to be allocated to the study drug arm. In these situations, an unequal randomization ratio should be considered. We propose a group variance-covariance and structures-based method to adapt the randomization ratio after interim analysis. We use simulations to compare commonly used statistical methods for continuous endpoints in assessing the impact of heteroscedasticity in equal and unequal randomization ratios and examine the extent to which the findings are affected by missing data.

Challenges and potential strategies utilizing external data for efficacy evaluation in small-sized clinical trials.

In clinical trials for diseases with very small patient populations, trial investigators may encounter recruitment difficulties. It can be challenging to conduct clinical trials with enough power to detect a treatment effect, and randomization may not be feasible due to timeline, budget, and ethical concerns. To bring breakthrough therapies to the market quickly, it is important to come up with efficient approaches to utilizing individual patient data through improved study design and sound statistical methods. Emerging topics in this area include the use of Bayesian approaches to flexibly incorporate prior information into the current clinical trials, the use of historical controls to efficiently conduct trials that will reduce the number of subjects recruited and ease ethical considerations, and the use of innovative study designs, such as a platform design, to improve the efficiency and speed of the medical therapy development progress. In this paper, we describe three scenarios which highlight some of the challenges encountered in small-sized clinical trial development and provide potential statistical approaches to overcome the aforementioned challenges.

Toward Smart Home Authentication Using PUF and Edge-Computing Paradigm.

The smart home is a crucial embodiment of the internet of things (IoT), which can facilitate users to access smart home services anytime and anywhere. Due to the limited resources of cloud computing, it cannot meet users' real-time needs. Therefore, edge computing emerges as the times require, providing users with better real-time access and storage. The application of edge computing in the smart home environment can enable users to enjoy smart home services. However, users and smart devices communicate through public channels, and malicious attackers may intercept information transmitted through public channels, resulting in user privacy disclosure. Therefore, it is a critical issue to protect the secure communication between users and smart devices in the smart home environment. Furthermore, authentication protocols in smart home environments also have some security challenges. In this paper, we propose an anonymous authentication protocol that applies edge computing to the smart home environment to protect communication security between entities. To protect the security of smart devices, we embed physical unclonable functions (PUF) into each smart device. Real-or-random model, informal security analysis, and ProVerif are adopted to verify the security of our protocol. Finally, we compare our protocol with existing protocols regarding security and performance. The comparison results demonstrate that our protocol has higher security and slightly better performance.

A Structural Comparison of SARS-CoV-2 Main Protease and Animal Coronaviral Main Protease Reveals Species-Specific Ligand Binding and Dimerization Mechanism.

Animal coronaviruses (CoVs) have been identified to be the origin of Severe Acute Respiratory Syndrome (SARS)-CoV, Middle East respiratory syndrome (MERS)-CoV, and probably SARS-CoV-2 that cause severe to fatal diseases in humans. Variations of zoonotic coronaviruses pose potential threats to global human beings. To overcome this problem, we focused on the main protease (Mpro), which is an evolutionary conserved viral protein among different coronaviruses. The broad-spectrum anti-coronaviral drug, GC376, was repurposed to target canine coronavirus (CCoV), which causes gastrointestinal infections in dogs. We found that GC376 can efficiently block the protease activity of CCoV Mpro and can thermodynamically stabilize its folding. The structure of CCoV Mpro in complex with GC376 was subsequently determined at 2.75 Å. GC376 reacts with the catalytic residue C144 of CCoV Mpro and forms an (R)- or (S)-configuration of hemithioacetal. A structural comparison of CCoV Mpro and other animal CoV Mpros with SARS-CoV-2 Mpro revealed three important structural determinants in a substrate-binding pocket that dictate entry and release of substrates. As compared with the conserved A141 of the S1 site and P188 of the S4 site in animal coronaviral Mpros, SARS-CoV-2 Mpro contains N142 and Q189 at equivalent positions which are considered to be more catalytically compatible. Furthermore, the conserved loop with residues 46-49 in animal coronaviral Mpros has been replaced by a stable α-helix in SARS-CoV-2 Mpro. In addition, the species-specific dimerization interface also influences the catalytic efficiency of CoV Mpros. Conclusively, the structural information of this study provides mechanistic insights into the ligand binding and dimerization of CoV Mpros among different species.

Glycyrrhizic Acid Derivatives Bearing Amino Acid Residues in the Carbohydrate Part as Dengue Virus E Protein Inhibitors: Synthesis and Antiviral Activity.

Dengue virus (DENV) is one of the most geographically distributed mosquito-borne flaviviruses, like Japanese encephalitis virus (JEV), and Zika virus (ZIKV). In this study, a library of the known and novel Glycyrrhizic acid (GL) derivatives bearing amino acid residues or their methyl/ethyl esters in the carbohydrate part were synthesized and studied as DENV inhibitors in vitro using the cytopathic effect (CPE), viral infectivity and virus yield assays with DENV1 and DENV-2 in Vero E6 and A549 cells. Among the GL conjugates tested, compound hits GL-D-ValOMe 3, GL-TyrOMe 6, GL-PheOEt 11, and GL-LysOMe 21 were discovered to have better antiviral activity than GL, with IC50 values ranging from <0.1 to 5.98 µM on the in vitro infectivity of DENV1 and DENV2 in Vero E6 and A549 cells. Compound hits 3, 6, 11, and 21 had a concentration-dependent inhibition on the virus yield in Vero E6, in which GL-D-ValOMe 3 and GL-PheOEt 11 were the most active inhibitors of DENV2 yield. Meanwhile, the time-of-addition assay indicated that conjugates GL-D-ValOMe 3 and GL-PheOEt 11 exhibited a substantial decrease in the DENV2 attachment stage. Subsequently, chimeric single-round infectious particles (SRIPs) of DENV2 C-prM-E protein/JEV replicon and DENV2 prM-E/ZIKV replicon were utilized for the DENV envelope I protein-mediated attachment assay. GL conjugates 3 and 11 significantly reduced the attachment of chimeric DENV2 C-prM-E/JEV and DENV2 prM-E/ZIKV SRIPs onto Vero E6 cells in a concentration-dependent manner but did not impede the attachment of wild-type JEV CprME/JEV and ZIKV prM-E/ZIKV SRIPs, indicating the inhibition of Compounds 3 and 11 on DENV2 E-mediated attachment. Molecular docking data revealed that Compounds 3 and 11 have hydrophobic interactions within a hydrophobic pocket among the interfaces of Domains I, II, and the stem region of the DENV2 envelope (E) protein. These results displayed that Compounds 3 and 11 were the lead compounds targeting the DENV E protein. Altogether, our findings provide new insights into the structure−activity relationship of GL derivatives conjugated with amino acid residues and can be the new fundamental basis for the search and development of novel flavivirus inhibitors based on natural compounds.

Ribonuclease 7-driven activation of ROS1 is a potential therapeutic target in hepatocellular carcinoma.

BACKGROUND & AIMS: There are currently limited therapeutic options for hepatocellular carcinoma (HCC), particularly when it is diagnosed at advanced stages. Herein, we examined the pathophysiological role of ROS1 and assessed the utility of ROS1-targeted therapy for the treatment of HCC. METHODS: Recombinant ribonucleases (RNases) were purified, and the ligand-receptor relationship between RNase7 and ROS1 was validated in HCC cell lines by Duolink, immunofluorescence, and immunoprecipitation assays. Potential interacting residues between ROS1 and RNase7 were predicted using a protein-protein docking approach. The oncogenic function of RNase7 was analyzed by cell proliferation, migration and invasion assays, and a xenograft mouse model. The efficacy of anti-ROS1 inhibitor treatment was evaluated in patient-derived xenograft (PDX) and orthotopic models. Two independent patient cohorts were analyzed to evaluate the pathological relevance of RNase7/ROS1. RESULTS: RNase7 associated with ROS1's N3-P2 domain and promoted ROS1-mediated oncogenic transformation. Patients with HCC exhibited elevated plasma RNase7 levels compared with healthy individuals. High ROS1 and RNase7 expression were strongly associated with poor prognosis in patients with HCC. In both HCC PDX and orthotopic mouse models, ROS1 inhibitor treatment markedly suppressed RNase7-induced tumorigenesis, leading to decreased plasma RNase7 levels and tumor shrinkage in mice. CONCLUSIONS: RNase7 serves as a high-affinity ligand for ROS1. Plasma RNase7 could be used as a biomarker to identify patients with HCC who may benefit from anti-ROS1 treatment. LAY SUMMARY: Receptor tyrosine kinases are known to be involved in tumorigenesis and have been targeted therapeutically for a number of cancers, including hepatocellular carcinoma. ROS1 is the only such receptor with kinase activity whose ligand has not been identified. Herein, we show that RNase7 acts as a ligand to activate ROS1 signaling. This has important pathophysiological and therapeutic implications. Anti-ROS1 inhibitors could be used to treatment patients with hepatocellular carcinoma and high RNase7 levels.

Crystal structure of the N-terminal domain of TagH reveals a potential drug targeting site.

Bacterial wall teichoic acids (WTAs) are synthesized intracellularly and exported by a two-component transporter, TagGH, comprising the transmembrane and ATPase subunits TagG and TagH. Here the dimeric structure of the N-terminal domain of TagH (TagH-N) was solved by single-wavelength anomalous diffraction using a selenomethionine-containing crystal, which shows an ATP-binding cassette (ABC) architecture with RecA-like and helical subdomains. Besides significant structural differences from other ABC transporters, a prominent patch of positively charged surface is seen in the center of the TagH-N dimer, suggesting a potential binding site for the glycerol phosphate chain of WTA. The ATPase activity of TagH-N was inhibited by clodronate, a bisphosphonate, in a non-competitive manner, consistent with the proposed WTA-binding site for drug targeting.

Proteasome inhibitors restore the STAT1 pathway and enhance the expression of MHC class I on human colon cancer cells.

BACKGROUND: A new strategy, particularly a novel combination, for immunotherapy in microsatellite stable metastatic colorectal cancer (mCRC) treatment needs to be formulated. Studies on the interferon-γ (IFN-γ)/ Janus kinase (JAK)/ signal transducer and activator of transcription (STAT)1 pathway provide new directions in this regard. METHODS: Our study applies three colon cancer cell lines, including microsatellite stable (MSS) cell lines, which are SW480 and SW620, and microsatellite instability-high (MSI-H) cell line, which is DLD-1. We compared the expressions of immune surface markers on colon cancer cells in response to IFN-γ. We elucidated these mechanisms, which involved the upregulation of immune surface markers. Furthermore, we examined real-world clinical samples using the PerkinElmer Opal multiplex system and NanoString analysis. RESULTS: We established that the baseline expression of major histocompatibility complex (MHC) class I alleles and programmed death-ligand 1 (PD-L1) were generally low in cell line models. The immune surface markers were significantly increased after IFN-γ stimulation on SW480 but were notably unresponsive on the SW620 cell line. We discovered that STAT1 and phosphorylated STAT1 (pSTAT1) were downregulated in the SW620 cell line. We verified that the STAT1/pSTAT1 could be restored through the application of proteasome inhibitors, especially bortezomib. The expression of MHC class I as downstream signals of STAT1 was also up-regulated by proteasome inhibitors. The similar results were reproduced in DLD-1 cell line, which was also initially unresponsive to IFN-γ. In real-world samples of patients with mCRC, we found that higher STAT1 expression in tumor cells was strongly indicative of a highly immunogenic microenvironment, with significantly higher expression levels of MHC class I and PD-L1, not only on tumor cells but also on non-tumor cells. Furthermore, tumor infiltrating lymphocytes (TILs) were increased in the positive-STAT1 group. Through NanoString analysis, we confirmed that the mRNA expressions of IFN-γ, human leukocyte antigen (HLA)-A, HLA-E, and HLA-G were also significantly higher in the positive-STAT1 group than those in the negative-STAT1 group. CONCLUSION: Our study provides a novel rationale for the addition of bortezomib, a proteasome inhibitor, into new immunotherapy combinations.

Enhancement of Anticancer Potential of Pterostilbene Derivative by Chalcone Hybridization.

Pterostilbene, a natural metabolite of resveratrol, has been indicated as a potent anticancer molecule. Recently, several pterostilbene derivatives have been reported to exhibit better anticancer activities than that of the parent pterostilbene molecule. In the present study, a series of pterostilbene derivatives were designed and synthesized by the hybridization of pterostilbene, chalcone, and cinnamic acid. The cytotoxic effect of these hybrid molecules was determined using two oral cancer cell lines, HSC-3 and OECM-1. (E)-3-(2-((E)-4-Hydroxystyryl)-4,6-dimethoxyphenyl)-1-(2-methoxyphenyl)prop-2-en-1-one (4d), with IC50 of 16.38 and 18.06 µM against OECM-1 and HSC-3, respectively, was selected for further anticancer mechanism studies. Results indicated that compound 4d effectively inhibited cell proliferation and induced G2/M cell cycle arrest via modulating p21, cyclin B1, and cyclin A2. Compound 4d ultimately induced cell apoptosis by reducing the expression of Bcl-2 and surviving. In addition, cleavage of PARP and caspase-3 were enhanced following the treatment of compound 4d with increased dose. To conclude, a number of pterostilbene derivatives were discovered to possess potent anticancer potentials. Among them, compound 4d was the most active, more active than the parent pterostilbene.

MT4-MMP promotes invadopodia formation and cell motility in FaDu head and neck cancer cells.

Hypoxia-inducible factor-1α (HIF-1α) induces cancer metastasis. We previously demonstrated that HIF-1α-induced membrane-type 4 matrix metalloproteinase (MT4-MMP) is involved in hypoxia-mediated metastasis in head and neck squamous cell carcinoma (HNSCC). However, the functions and detailed mechanisms of MT4-MMP in cancer metastasis are not well understood. In this study, we investigated whether MT4-MMP regulates invadopodia formation or individual cell movement-both critical to cancer migration and invasion-in three-dimensional (3D) environments. By expressing MT4-MMP in the HNSCC cell line FaDu, we demonstrated that MT4-MMP increases invadopodia formation and gelatin degradation. Furthermore, the amoeboid-like cell movement on collagen gel was increased by MT4-MMP expression in FaDu cells. Mechanistically, MT4-MMP may induce invadopodia formation by binding with Tks5 and PDGFRα to result in Src activation and promote amoeboid-like movement by stimulating the small GTPases Rho and Cdc42. Altogether, our data indicate that MT4-MMP induces two crucial mechanisms of cancer dissemination, invadopodia formation and amoeboid movement, and elucidate the prometastatic role of MT4-MMP in hypoxia-mediated cancer metastasis.

Structural evolution of in situ boron-doped SiGe ultrathin film analyzed by multi-optical methods.

In situ boron (B)-doped SiGe (BSG) layer is extensively used in the source (S)/(D) drain of metal-oxide-semiconductor field-effect transistors. An unexpected structural evolution occurs in BSG during metallization and activation annealing during actual fabrication, which involves a correlated interaction between B and SiGe. Herein, the complicated phenomena of the structural evolution of BSG were analyzed by 325 nm micro-Raman spectroscopy, x-ray photoelectron spectroscopy (XPS), reflective second harmonic generation (RSHG), and synchrotron x-ray diffraction (XRD). Optical inspection was integrated into these processes to establish a multi-optical method. 325 nm micro-Raman spectroscopy was used to determine variations in Si-Si, Si-Ge, and Ge-Ge bonds in BSG. XPS exhibited the binding energy evolution of Ge3d during different annealing processes at varied Ge ratios and B concentrations. RSHG revealed the polar Si-B and Ge-B bonds formed during annealing. Synchrotron XRD provided the structure and strain changes of BSG. Secondary-ion mass spectrometer profiles provided the species distribution, which was used to examine the results of multi-optical method. Furthermore, double-layered BSG (DBSG) with different B concentrations were analyzed using the multi-optical method. Results revealed that Ge aggregated in the homogeneous interface of DBSG, and that B dopants in BSG served as carrier providers that strongly influenced the BSG structure. However, BSG with excessive B concentration was unstable and increased the B content (SiB3) through metallization. For BSG with a suitable B concentration, the formation of Si-B and Ge-B bonds suppressed the diffusion of Ge from SiGe, thereby reducing the possibility of Ge loss and further B pipe-up in the heavily doped S/D region.

Discovery of 4-Anilinoquinolinylchalcone Derivatives as Potential NRF2 Activators.

Activation of nuclear factor erythroid-2-related factor 2 (NRF2) has been proven to be an effective means to prevent the development of cancer, and natural curcumin stands out as a potent NRF2 activator and cancer chemopreventive agent. In this study, we have synthesized a series of 4-anilinoquinolinylchalcone derivatives, and used a NRF2 promoter-driven firefly luciferase reporter stable cell line, the HaCaT/ARE cells, to screen a panel of these compounds. Among them, (E)-3-{4-[(4-acetylphenyl)amino]quinolin-2-yl}-1-(4-fluorophenyl)prop-2-en-1-one (13b) significantly increased NRF2 activity in the HaCaT cell with a half maximal effective concentration (EC50) value of 1.95 µM. Treatment of compound 13b upregulated HaCaT cell NRF2 expression at the protein level. Moreover, the mRNA level of NRF2 target genes, heme oxygenase-1 (HO-1), glutamate-cysteine ligase catalytic subunit (GCLC), and glucose-6-phosphate dehydrogenase (G6PD) were significantly increased in HaCaT cells upon the compound 13b treatment. The molecular docking results exhibited that the small molecule 13b is well accommodated by the bound region of Kelch-like ECH-associated protein 1 (Keap1)-Kelch and NRF2 through stable hydrogen bonds and hydrophobic interaction, which contributed to the enhancement of affinity and stability between the ligand and receptor. Compound 13b has been identified as the lead compound for further structural optimization.