Development of a Novel, Potent, and Selective Sialyltransferase Inhibitor for Suppressing Cancer Metastasis.

Sialyltransferase-catalyzed membrane protein and lipid glycosylation plays a vital role as one of the most abundant post-translational modifications and diversification reactions in eukaryotes. However, aberrant sialylation has been associated with cancer malignancy and metastasis. Sialyltransferases thus represent emerging targets for the development of small molecule cancer drugs. Herein, we report the inhibitory effects of a recently discovered lithocholic acid derivative FCW393 on sialyltransferase catalytic activity, integrin sialyation, cancer-associated signal transduction, MDA-MB-231 and B16F10 cell migration and invasion, and in in vivo studies, on tumor growth, metastasis, and angiogenesis. FCW393 showed effective and selective inhibition of the sialyltransferases ST6GAL1 (IC50 = 7.8 µM) and ST3GAL3 (IC50 = 9.45 µM) relative to ST3GAL1 (IC50 > 400 µM) and ST8SIA4 (IC50 > 100 µM). FCW393 reduced integrin sialylation in breast cancer and melanoma cells dose-dependently and downregulated proteins associated with the integrin-regulated FAK/paxillin and GEF/Rho/ROCK pathways, and with the VEGF-regulated Akt/NFκB/HIF-1α pathway. FCW393 inhibited cell migration (IC50 = 2.6 µM) and invasion in in vitro experiments, and in in vivo studies of tumor-bearing mice, FCW393 reduced tumor size, angiogenesis, and metastatic potential. Based on its demonstrated selectivity, cell permeability, relatively low cytotoxicity (IC50 = 55 µM), and high efficacy, FCW393 shows promising potential as a small molecule experimental tool compound and a lead for further development of a novel cancer therapeutic.

Prediction of clinically significant prostate cancer through urine metabolomic signatures: A large-scale validated study.

PURPOSE: Currently, there are no accurate markers for predicting potentially lethal prostate cancer (PC) before biopsy. This study aimed to develop urine tests to predict clinically significant PC (sPC) in men at risk. METHODS: Urine samples from 928 men, namely, 660 PC patients and 268 benign subjects, were analyzed by gas chromatography/quadrupole time-of-flight mass spectrophotometry (GC/Q-TOF MS) metabolomic profiling to construct four predictive models. Model I discriminated between PC and benign cases. Models II, III, and GS, respectively, predicted sPC in those classified as having favorable intermediate risk or higher, unfavorable intermediate risk or higher (according to the National Comprehensive Cancer Network risk groupings), and a Gleason sum (GS) of ≥ 7. Multivariable logistic regression was used to evaluate the area under the receiver operating characteristic curves (AUC). RESULTS: In Models I, II, III, and GS, the best AUCs (0.94, 0.85, 0.82, and 0.80, respectively; training cohort, N = 603) involved 26, 24, 26, and 22 metabolites, respectively. The addition of five clinical risk factors (serum prostate-specific antigen, patient age, previous negative biopsy, digital rectal examination, and family history) significantly improved the AUCs of the models (0.95, 0.92, 0.92, and 0.87, respectively). At 90% sensitivity, 48%, 47%, 50%, and 36% of unnecessary biopsies could be avoided. These models were successfully validated against an independent validation cohort (N = 325). Decision curve analysis showed a significant clinical net benefit with each combined model at low threshold probabilities. Models II and III were more robust and clinically relevant than Model GS. CONCLUSION: This urine test, which combines urine metabolic markers and clinical factors, may be used to predict sPC and thereby inform the necessity of biopsy in men with an elevated PC risk.

Increase in Akkermansiaceae in Gut Microbiota of Prostate Cancer-Bearing Mice.

Gut microbiota are reported to be associated with many diseases, including cancers. Several bacterial taxa have been shown to be associated with cancer development or response to treatment. However, longitudinal microbiota alterations during the development of cancers are relatively unexplored. To better understand how microbiota changes, we profiled the gut microbiota composition from prostate cancer-bearing mice and control mice at five different time points. Distinct gut microbiota differences were found between cancer-bearing mice and control mice. Akkermansiaceae was found to be significantly higher in the first three weeks in cancer-bearing mice, which implies its role in the early stage of cancer colonization. We also found that Bifidobacteriaceae and Enterococcaceae were more abundant in the second and last sampling week, respectively. The increments of Akkermansiaceae, Bifidobacteriaceae and Enterococcaceae were previously found to be associated with responses to immunotherapy, which suggests links between these bacteria families and cancers. Additionally, our function analysis showed that the bacterial taxa carrying steroid biosynthesis and butirosin and neomycin biosynthesis were increased, whereas those carrying naphthalene degradation decreased in cancer-bearing mice. Our work identified the bacteria taxa altered during prostate cancer progression and provided a resource of longitudinal microbiota profiles during cancer development in a mouse model.

Sesquiterpene Lactone Deoxyelephantopin Isolated from Elephantopus scaber and Its Derivative DETD-35 Suppress BRAFV600E Mutant Melanoma Lung Metastasis in Mice.

Melanoma is a highly metastatic disease with an increasing rate of incidence worldwide. It is treatment refractory and has poor clinical prognosis; therefore, the development of new therapeutic agents for metastatic melanoma are urgently required. In this study, we created a lung-seeking A375LM5IF4g/Luc BRAFV600E mutant melanoma cell clone and investigated the bioefficacy of a plant sesquiterpene lactone deoxyelephantopin (DET) and its novel semi-synthetic derivative, DETD-35, in suppressing metastatic A375LM5IF4g/Luc melanoma growth in vitro and in a xenograft mouse model. DET and DETD-35 treatment inhibited A375LM5IF4g/Luc cell proliferation, and induced G2/M cell-cycle arrest and apoptosis. Furthermore, A375LM5IF4g/Luc exhibited clonogenic, metastatic and invasive abilities, and several A375LM5IF4g/Luc metastasis markers, N-cadherin, MMP2, vimentin and integrin α4 were significantly suppressed by treatment with either compound. Interestingly, DET- and DETD-35-induced Reactive Oxygen Species (ROS) generation and glutathione (GSH) depletion were found to be upstream events important for the in vitro activities, because exogenous GSH supplementation blunted DET and DETD-35 effects on A375LM5IF4g/Luc cells. DET and DETD-35 also induced mitochondrial DNA mutation, superoxide production, mitochondrial bioenergetics dysfunction, and mitochondrial protein deregulation. Most importantly, DET and DETD-35 inhibited lung metastasis of A375LM5IF4g/Luc in NOD/SCID mice through inhibiting pulmonary vascular permeability and melanoma cell (Mel-A+) proliferation, angiogenesis (VEGF+, CD31+) and EMT (N-cadherin) in the tumor microenvironment in the lungs. These findings indicate that DET and DETD-35 may be useful in the intervention of lung metastatic BRAFV600E mutant melanoma.

O-Glycosylation-mediated signaling circuit drives metastatic castration-resistant prostate cancer.

Disseminated castration-resistant prostate cancer (CRPC) is a common disease in men that is characterized by limited survival and resistance to androgen-deprivation therapy. The increase in human epidermal growth factor receptor 2 (HER2) signaling contributes to androgen receptor activity in a subset of patients with CRPC; however, enigmatically, HER2-targeted therapies have demonstrated a lack of efficacy in patients with CRPC. Aberrant glycosylation is a hallmark of cancer and involves key processes that support cancer progression. Using transcriptomic analysis of prostate cancer data sets, histopathologic examination of clinical specimens, and in vivo experiments of xenograft models, we reveal in this study a coordinated increase in glycan-binding protein, galectin-4, specific glycosyltransferases of core 1 synthase, glycoprotein- N-acetylgalactosamine 3-ß-galactosyltransferase 1 (C1GALT1) and ST3 beta-galactoside α-2,3-sialyltransferase 1 (ST3GAL1), and resulting mucin-type O-glycans during the progression of CRPC. Furthermore, galectin-4 engaged with C1GALT1-dependent O-glycans to promote castration resistance and metastasis by activating receptor tyrosine kinase signaling and cancer cell stemness properties mediated by SRY-box 9 (SOX9). This galectin-glycan interaction up-regulated the MYC-dependent expression of C1GALT1 and ST3GAL1, which altered cellular mucin-type O-glycosylation to allow for galectin-4 binding. In clinical prostate cancer, high-level expression of C1GALT1 and galectin-4 together predict poor overall survival compared with low-level expression of C1GALT1 and galectin-4. In summary, MYC regulates abnormal O-glycosylation, thus priming cells for binding to galectin-4 and downstream signaling, which promotes castration resistance and metastasis.-Tzeng, S.-F., Tsai, C.-H., Chao, T.-K., Chou, Y.-C., Yang, Y.-C., Tsai, M.-H., Cha, T.-L., Hsiao, P.-W. O-Glycosylation-mediated signaling circuit drives metastatic castration-resistant prostate cancer.

Development of single-chain variable fragments (scFv) against influenza virus targeting hemagglutinin subunit 2 (HA2).

Influenza A viruses (IAV) are widespread in birds and domestic poultry, occasionally causing severe epidemics in humans and posing health threats. Hence, the need to develop a strategy for prophylaxis or therapy, such as a broadly neutralizing antibody against IAV, is urgent. In this study, single-chain variable fragment (scFv) phage display technology was used to select scFv fragments recognizing influenza envelope proteins. The Tomlinson I and J scFv phage display libraries were screened against the recombinant HA2 protein (rHA2) for three rounds. Only the third-round elution sample of the Tomlinson J library showed high binding affinity to rHA2, from which three clones (3JA18, 3JA62, and 3JA78) were chosen for preparative-scale production as soluble antibody by E. coli. The clone 3JA18 was selected for further tests due to its broad affinity for influenza H1N1, H3N2 and H5N1. Simulations of the scFv 3JA18-HA trimer complex revealed that the complementarity-determining region of the variable heavy chain (VH-CDR2) bound the stem region of HA. Neutralization assays using a peptide derived from VH-CDR2 also supported the simulation model. Both the selected antibody and its derived peptide were shown to suppress infection with H5N1 and H1N1 viruses, but not H3N2 viruses. The results also suggested that the scFvs selected from rHA2 could have neutralizing activity by interfering with the function of the HA stem region during virus entry into target cells.

Shikonin-enhanced cell immunogenicity of tumor vaccine is mediated by the differential effects of DAMP components.

BACKGROUND: The tumor cell lysate-pulsed, dendritic cell (DC)-based cancer vaccine approaches are being actively evaluated for application to cancer immunotherapy, hopefully at a personalized medicine base. There is apparently an emerging technical problem however, the lack of highly efficacious potency in activation of patient's DCs for T-cell priming and the associated process for presenting tumor immunogenicity. METHODS: One strategy to address this is to consider the manipulation of the tumor immunogenic cells death (ICD) complex ex-vivo for maximal activation of DC efficacy. In our previous study we showed that phytochemical shikonin (SK) can drastically enhance ICD activity in mouse tumor cells treated ex-vivo, and the resultant tumor cell lysate (TCL) can effectively augment such SK-TCL pulsed DC vaccine activity in vivo in anti-tumor activities. In this study, we investigated the specifics and the multi-functional effects of various damaged associated molecular pattern (DAMP) components of the ICD complex for their participation, roles and potential cross talks in activating DCs, as measured by five different functional assays. RESULTS: Among three DAMPs tested, HSP70 and CRT mediate a key role in SK-TCL-induced DC immunity for both CD4(+) and CD8(+) T cell proliferations in vitro. HSP70 is the most important component, followed by CRT, then HMGB1 in facilitating DC immunity on suppressing metastasis of mouse 4 T1 mammary tumors and prolonging survival in test mice. Only HSP70, but not CRT or HMGB1, is effective for the suppression of both granulocytic and monocytic MDSC populations in vivo. Both HSP70 and HMGB1, but not CRT, are essential in activating the expression of three key ICD molecules-associated receptors on test DCs. Each of the three test ICD proteins can exhibit a distinguishable pattern in stimulating the expression of four key chemokines in test DCs. CONCLUSION: Our findings on the differential roles or effect of various ICD components in activating vaccinated DCs may help formulate new strategies for future cancer vaccine designs.

Assessment of Anti-Prostate Cancer Activity among Four Seaweeds, with Focus on Caulerpa lentillifera J.Agardh.

In response to a global shift towards health-conscious and environmentally sustainable food choices, seaweed has emerged as a focus for researchers due to its large-scale cultivation potential and the development of bioactive substances. This research explores the potential anticancer properties of seaweed extracts, focusing on analyzing the impact of four common edible seaweeds in Taiwan on prostate cancer (PCa) cells' activity. The study used bioassay-guided fractionation to extract Cl80 from various seaweeds with androgen receptor (AR)-inhibitory activity. Cl80 demonstrated effective suppression of 5α-dihydrotestosterone (DHT)-induced AR activity in 103E cells and attenuated the growth and prostate-specific antigen (PSA) protein expression in LNCaP and 22Rv1 cells. Additionally, Cl80 exhibited differential effects on various PCa cell lines. Concentrations above 5 µg/mL significantly inhibited LNCaP cell proliferation, while 22Rv1 cells were more resistant to Cl80. PC-3 cell proliferation was inhibited at 5 µg/mL but not completely at 50 µg/mL. A clonogenic assay showed that at a concentration of 0.5 µg/mL, the colony formation in LNCaP and PC-3 cells was significantly reduced, with a dose-dependent effect. Cl80 induced apoptosis in all PCa cell types, especially in LNCaP cells, with increased apoptotic cells observed at higher concentrations. Cl80 also decreased the mitochondrial membrane potential (ΔΨm) in a dose-dependent manner in all PCa cell lines. Furthermore, Cl80 suppressed the migration ability of PCa cells, with significant reductions observed in LNCaP, 22Rv1, and PC-3 cells at various concentrations. These compelling findings highlight the promising therapeutic potential of C. lentillifera J.Agardh and its isolated compound Cl80 in the treatment of PCa.

Predicting Clinically Significant Prostate Cancer Using Urine Metabolomics via Liquid Chromatography Mass Spectrometry.

PURPOSE: Biomarkers predicting clinically significant prostate cancer (sPC) before biopsy are currently lacking. This study aimed to develop a non-invasive urine test to predict sPC in at-risk men using urinary metabolomic profiles. MATERIALS AND METHODS: Urine samples from 934 at-risk subjects and 268 treatment-naïve PC patients were subjected to liquid chromatography/mass spectrophotometry (LC-MS)-based metabolomics profiling using both C18 and hydrophilic interaction liquid chromatography (HILIC) column analyses. Four models were constructed (training cohort [n=647]) and validated (validation cohort [n=344]) for different purposes. Model I differentiates PC from benign cases. Models II, III, and a Gleason score model (model GS) predict sPC that is defined as National Comprehensive Cancer Network (NCCN)-categorized favorable-intermediate risk group or higher (Model II), unfavorable-intermediate risk group or higher (Model III), and GS ≥7 PC (model GS), respectively. The metabolomic panels and predicting models were constructed using logistic regression and Akaike information criterion. RESULTS: The best metabolomic panels from the HILIC column include 25, 27, 28 and 26 metabolites in Models I, II, III, and GS, respectively, with area under the curve (AUC) values ranging between 0.82 and 0.91 in the training cohort and between 0.77 and 0.86 in the validation cohort. The combination of the metabolomic panels and five baseline clinical factors that include serum prostate-specific antigen, age, family history of PC, previously negative biopsy, and abnormal digital rectal examination results significantly increased AUCs (range 0.88-0.91). At 90% sensitivity (validation cohort), 33%, 34%, 41%, and 36% of unnecessary biopsies were avoided in Models I, II, III, and GS, respectively. The above results were successfully validated using LC-MS with the C18 column. CONCLUSIONS: Urinary metabolomic profiles with baseline clinical factors may accurately predict sPC in men with elevated risk before biopsy.

Long-term hematopoietic transfer of the anti-cancer and lifespan-extending capabilities of a genetically engineered blood system by transplantation of bone marrow mononuclear cells.

A causal relationship exists among the aging process, organ decay and disfunction, and the occurrence of various diseases including cancer. A genetically engineered mouse model, termed Klf1K74R/K74R or Klf1(K74R), carrying mutation on the well-conserved sumoylation site of the hematopoietic transcription factor KLF1/EKLF has been generated that possesses extended lifespan and healthy characteristics, including cancer resistance. We show that the healthy longevity characteristics of the Klf1(K74R) mice, as exemplified by their higher anti-cancer capability, are likely gender-, age-, and genetic background-independent. Significantly, the anti-cancer capability, in particular that against melanoma as well as hepatocellular carcinoma, and lifespan-extending property of Klf1(K74R) mice, could be transferred to wild-type mice via transplantation of their bone marrow mononuclear cells at a young age of the latter. Furthermore, NK(K74R) cells carry higher in vitro cancer cell-killing ability than wild-type NK cells. Targeted/global gene expression profiling analysis has identified changes in the expression of specific proteins, including the immune checkpoint factors PDCD and CD274, and cellular pathways in the leukocytes of the Klf1(K74R) that are in the directions of anti-cancer and/or anti-aging. This study demonstrates the feasibility of developing a transferable hematopoietic/blood system for long-term anti-cancer and, potentially, for anti-aging.

Monoglycosylated SARS-CoV-2 receptor binding domain fused with HAstem-scaffolded protein vaccine confers broad protective immunity against SARS-CoV-2 and influenza viruses.

The SARS-CoV-2 and influenza pandemics have posed a devastating threat to global public health. The best strategy for preventing the further spread of these respiratory viruses worldwide is to administer a vaccine capable of targeting both viruses. Here, we show that a novel monoglycosylated vaccine designed based on the influenza virus HAstem conserved domain fused with the SARS-CoV-2 spike-RBD domain (HSSRmg) can present proper antigenicity that elicits sufficient neutralization efficacy against various SARS-CoV-2 variants while simultaneously providing broad protection against H1N1 viruses in mice. Compared with the fully glycosylated HSSR (HSSRfg), HSSRmg induced higher ELISA titers targeting HAstem and spike-RBD and exhibited significantly enhanced neutralization activity against the Wuhan pseudovirus. The enhanced immune responses raised by JR300-adjuvanted HSSRmg compared to HSSRmg alone include more anti-HAstem and anti-spike-RBD antibodies that provide cross-protection against H1N1 challenges and cross-neutralization of SARS-CoV-2 pseudoviruses. Furthermore, the enhanced immune response raised by JR300-adjuvanted-HSSRmg skews toward a more balanced Th1/Th2 response than that raised by HSSRmg alone. Notably, HSSRmg elicited more plasma B cells and memory B cells, and higher IL-4 and IFN-γ cytokine immune responses than spike (S-2P) in mice with preexisting influenza-specific immunity, suggesting that B-cell activation most likely occurs through CD4+ T-cell stimulation. This study demonstrated that HSSRmg produced using a monoglycosylation process and combined with the JR300 adjuvant elicits superior cross-strain immune responses against SARS-CoV-2 and influenza viruses in mice compared with S-2P. JR300-adjuvanted HSSRmg has great potential as a coronavirus-influenza vaccine that provides dual protection against SARS-CoV-2 and influenza infections.

Pilot-scale production of inactivated monoglycosylated split H1N1 influenza virus vaccine provides cross-strain protection against influenza viruses.

Influenza epidemics and pandemics caused by newly emerging virus strains highlight an urgent need to develop a universal vaccine against viruses. Previously, a monoglycosylated X-181mg vaccine demonstrated that the HA possessing a single N-acetylglucosamine at each N-glycosylation site is superior to confer broader protection in mice than conventional vaccines. However, the greatest challenge in conducting clinical trials is the need to develop robust manufacturing processes capable of producing vaccines at the pilot scale with the desired stability, potency, and efficacy. Whether the monoglycosylated virus vaccine platform can be applied to the new vaccine strain in a timely manner and whether the mass-produced vaccine has the proper immunogenicity to induce cross-protective immunity remains unclear. Here, we show that a pilot-scale manufacturing process produced a monoglycosylated A/Brisbane/02/2018(H1N1) virus vaccine (IVR-190mg) with a single glycan at each glycosylation site of HA and NA. Compared with the fully glycosylated virus vaccine (IVR-190fg), the IVR-190mg provided broader cross-protection in mice against a wide range of H1N1 variants. The enhanced antibody responses induced by IVR-190mg immunization include higher hemagglutination-inhibition titers, higher neutralization activity, more anti-HA head domain, more anti-HA stem antibodies, higher neuraminidase activity inhibition titers, and notably, higher antibody-dependent cellular cytotoxicity. Additionally, the IVR-190mg also induced a more balanced Th1/Th2 response and elicited broader splenic CD4+ and CD8+ T-cell responses than IVR-190fg. This study demonstrated that IVR-190mg produced using a pilot-scale manufacturing process elicits comprehensive cross-strain immune responses that have great potential to substantially mitigate the need for yearly reformulation of strain-specific inactivated vaccines.

Immunoediting instructs tumor metabolic reprogramming to support immune evasion.

Immunoediting sculpts immunogenicity and thwarts host anti-tumor responses in tumor cells during tumorigenesis; however, it remains unknown whether metabolic programming of tumor cells can be guided by immunosurveillance. Here, we report that T cell-mediated immunosurveillance in early-stage tumorigenesis instructs c-Myc upregulation and metabolic reprogramming in tumor cells. This previously unexplored tumor-immune interaction is controlled by non-canonical interferon gamma (IFNγ)-STAT3 signaling and supports tumor immune evasion. Our findings uncover that immunoediting instructs deregulated bioenergetic programs in tumor cells to empower them to disarm the T cell-mediated immunosurveillance by imposing metabolic tug-of-war between tumor and infiltrating T cells and forming the suppressive tumor microenvironment.

Long noncoding RNA Smyca coactivates TGF-ß/Smad and Myc pathways to drive tumor progression.

BACKGROUND: Metastasis and chemoresistance are major culprits of cancer mortality, but factors contributing to these processes are incompletely understood. METHODS: Bioinformatics methods were used to identify the relations of Smyca expression to clinicopathological features of human cancers. RNA-sequencing analysis was used to reveal Smyca-regulated transcriptome. RNA pull-down and RNA immunoprecipitation were used to examine the binding of Smyca to Smad3/4 and c-Myc/Max. Chromatin immunoprecipitation and chromatin isolation by RNA purification were used to determine the binding of transcription factors and Smyca to various gene loci, respectively. Real-time RT-PCR and luciferase assay were used to examine gene expression levels and promoter activities, respectively. Xenograft mouse models were performed to evaluate the effects of Smyca on metastasis and chemoresistance. Nanoparticle-assisted gapmer antisense oligonucleotides delivery was used to target Smyca in vivo. RESULTS: We identify lncRNA Smyca for its association with poor prognosis of many cancer types. Smyca potentiates metabolic reprogramming, migration, invasion, cancer stemness, metastasis and chemoresistance. Mechanistically, Smyca enhances TGF-ß/Smad signaling by acting as a scaffold for promoting Smad3/Smad4 association and further serves as a Smad target to amplify/prolong TGF-ß signaling. Additionally, Smyca potentiates c-Myc-mediated transcription by enhancing the recruitment of c-Myc/Max complex to a set of target promoters and c-Myc binding to TRRAP. Through potentiating TGF-ß and c-Myc pathways, Smyca synergizes the Warburg effect elicited by both pathways but evades the anti-proliferative effect of TGF-ß. Targeting Smyca prevents metastasis and overcomes chemoresistance. CONCLUSIONS: This study uncovers a lncRNA that coordinates tumor-relevant pathways to orchestra a pro-tumor program and establishes the clinical values of Smyca in cancer prognosis and therapy.

Matriptase-2/NR4A3 axis switches TGF-ß action toward suppression of prostate cancer cell invasion, tumor growth, and metastasis.

Dysregulation of pericellular proteolysis is strongly implicated in cancer metastasis through alteration of cell invasion and the microenvironment. Matriptase-2 (MT-2) is a membrane-anchored serine protease which can suppress prostate cancer (PCa) cell invasion. In this study, we showed that MT-2 was down-regulated in PCa and could suppress PCa cell motility, tumor growth, and metastasis. Using microarray and biochemical analysis, we found that MT-2 shifted TGF-ß action towards its tumor suppressor function by repressing epithelial-to-mesenchymal transition (EMT) and promoting Smad2 phosphorylation and nuclear accumulation to upregulate two TGF-ß1 downstream effectors (p21 and PAI-1), culminating in hindrance of PCa cell motility and malignant growth. Mechanistically, MT-2 could dramatically up-regulate the expression of nuclear receptor NR4A3 via iron metabolism in PCa cells. MT-2-induced NR4A3 further coactivated Smad2 to activate p21 and PAI-1 expression. In addition, NR4A3 functioned as a suppressor of PCa and mediated MT-2 signaling to inhibit PCa tumorigenesis and metastasis. These results together indicate that NR4A3 sustains MT-2 signaling to suppress PCa cell invasion, tumor growth, and metastasis, and serves as a contextual factor for the TGF-ß/Smad2 signaling pathway in favor of tumor suppression via promoting p21 and PAI-1 expression.

Sialyltransferase Inhibitors Suppress Breast Cancer Metastasis.

We report the synthesis and evaluation of a series of cell-permeable and N- versus O-selective sialyltransferase inhibitors. Inhibitor design entailed the functionalization of lithocholic acid at C(3) and at the cyclopentane ring side chain. Among the series, FCW34 and FCW66 were shown to inhibit MDA-MB-231 cell migration as effectively as ST3GALIII-gene knockdown did. FCW34 was shown to inhibit tumor growth, reduce angiogenesis, and delay cancer cell metastasis in animal models. Furthermore, FCW34 inhibited vessel development and suppressed angiogenic activity in transgenic zebrafish models. Our results provide clear evidence that FCW34-induced sialyltransferase inhibition reduces cancer cell metastasis by decreasing N-glycan sialylation, thus altering the regulation of talin/integrin/FAK/paxillin and integrin/NFκB signaling pathways.

Comparison of Chicken Immune Responses after Inoculation with H5 Avian Influenza Virus-like Particles Produced by Insect Cells or Pupae.

INTRODUCTION: Novel clade 2.3.4.4 H5 highly pathogenic avian influenza virus (HPAIV) outbreaks have occurred since early 2015 in Taiwan and impacted the island economically, like they have many countries. This research investigates the immunogenicity of two HPAIV-like particles to assess their promise as vaccine candidates. MATERIAL AND METHODS: The haemagglutinin (HA) gene derived from clade 2.3.4.4 H5 HPAIV and matrix protein 1 (M1) gene were cloned into the pFastBac Dual baculovirus vector. The resulting recombinant viruses were expressed in Spodoptera frugiperda moth (Sf)21 cells and silkworm pupae to generate Sf21 virus-like particles (VLP) and silkworm pupa VLP. Two-week-old specific pathogen-free chickens were immunised and their humoral and cellular immune responses were analysed. RESULTS: The silkworm pupa VLP had higher haemagglutination competence. Both VLP types elicited haemagglutination inhibition antibodies, anti-HA antibodies, splenic interferon gamma (IFN-γ) and interleukin 4 (IL-4) mRNA expression, and CD4+/CD8+ ratio elevation. However, chickens receiving silkworm pupa VLP exhibited a significantly higher anti-HA antibody titre in ELISA after vaccination. Although Sf21 VLP recipients expressed more IFN-γ and IL-4, the increase in IFN-γ did not significantly raise the CD4+/CD8+ ratio and the increase in IL-4 did not promote anti-HA antibodies. CONCLUSION: Both VLP systems possess desirable immunogenicity in vivo. However, in respect of immunogenic efficacy and the production cost, pupa VLP may be the superior vaccine candidate against clade 2.3.4.4 H5 HPAIV infection.

Arginine is an epigenetic regulator targeting TEAD4 to modulate OXPHOS in prostate cancer cells.

Arginine plays diverse roles in cellular physiology. As a semi-essential amino acid, arginine deprivation has been used to target cancers with arginine synthesis deficiency. Arginine-deprived cancer cells exhibit mitochondrial dysfunction, transcriptional reprogramming and eventual cell death. In this study, we show in prostate cancer cells that arginine acts as an epigenetic regulator to modulate histone acetylation, leading to global upregulation of nuclear-encoded oxidative phosphorylation (OXPHOS) genes. TEAD4 is retained in the nucleus by arginine, enhancing its recruitment to the promoter/enhancer regions of OXPHOS genes and mediating coordinated upregulation in a YAP1-independent but mTOR-dependent manner. Arginine also activates the expression of lysine acetyl-transferases and increases overall levels of acetylated histones and acetyl-CoA, facilitating TEAD4 recruitment. Silencing of TEAD4 suppresses OXPHOS functions and prostate cancer cell growth in vitro and in vivo. Given the strong correlation of TEAD4 expression and prostate carcinogenesis, targeting TEAD4 may be beneficially used to enhance arginine-deprivation therapy and prostate cancer therapy.

CD28 engagement inhibits CD73-mediated regulatory activity of CD8+ T cells.

CD28 is required for T cell activation as well as the generation of CD4+Foxp3+ Treg. It is unclear, however, how CD28 costimulation affects the development of CD8+ T cell suppressive function. Here, by use of Hepa1.6.gp33 in vitro killing assay and B16.gp33 tumor mouse model we demonstrate that CD28 engagement during TCR ligation prevents CD8+ T cells from becoming suppressive. Interestingly, our results showed that ectonucleotidase CD73 expression on CD8+ T cells is upregulated in the absence of CD28 costimulation. In both murine and human tumor-bearing hosts, CD73 is upregulated on CD28-CD8+ T cells that infiltrate the solid tumor. UPLC-MS/MS analysis revealed that CD8+ T cells activation without CD28 costimulation produces elevated levels of adenosine and that CD73 mediates its production. Adenosine receptor antagonists block CD73-mediated suppression. Our data support the notion that CD28 costimulation inhibits CD73 upregulation and thereby prevents CD8+ T cells from becoming suppressive. This study uncovers a previously unidentified role for CD28 costimulation in CD8+ T cell activation and suggests that the CD28 costimulatory pathway can be a potential target for cancer immunotherapy.

Dual degradation of aurora A and B kinases by the histone deacetylase inhibitor LBH589 induces G2-M arrest and apoptosis of renal cancer cells.

PURPOSE: This study is aimed at investigating antineoplastic efficacy of histone deacetylase inhibitor (HDACI) LBH589 on renal cell carcinoma (RCC) and elucidating the novel molecular mechanisms involved in growth arrest and apoptosis by targeting the important nonhistone molecules. EXPERIMENTAL DESIGN: We analyzed the growth-inhibitory effect of LBH589 on RCC by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay in vitro and antitumor efficacy by xenograft experiments in vivo. To verify the associated molecular mechanisms involved in LBH589-mediated cell death and cell cycle progression by Western blotting and fluorescence-activated cell sorting analysis. RESULTS: HDACI LBH589 induced degradation of both Aurora A and B kinases through a proteasome-mediated pathway by targeting HDAC3 and HDAC6. The dual degradation of Aurora A and B kinases mediated by LBH589 resulted in inducing G2-M arrest and apoptosis of renal cancer cell lines and our results also showed that LBH589 potently inhibited renal cancer cell growth in vitro and suppressed tumor formation in vivo. The Aurora A and B kinases and HDAC3 are overexpressed in the human RCC tumor tissues examined, which make them perfect targets for HDACI LBH589 treatment. CONCLUSIONS: Our in vitro and in vivo data showed that LBH589 has potent anticancer effect of renal cancer cells. LBH589 and other HDACI treatment resulted in inducing G2-M arrest and apoptosis of renal cancer cells through degradation of Aurora A and B kinases by inhibition of HDAC3 and HDAC6. The clinical efficacy of LBH589 in the treatment of patients with metastatic RCC, especially those with high Aurora kinase and HDAC expression, is worthy of further investigation.