Prostate Carcinoma Cells Research Articles

Shikonin Reduces Growth of Docetaxel-Resistant Prostate Cancer Cells Mainly through Necroptosis.

Simple SummaryProstate carcinoma (PCa) is the most common tumor in men with an increasing age-associated risk. Several therapy strategies, one of which is docetaxel (DX) chemotherapy, have been established. However, due to the development of therapy resistance, in which chemotherapy no longer effectively combats the cancer, advanced, metastasized PCa with a poor prognosis may become manifested and therapy inevitably fails. Thus, new treatment options are urgently needed. Shikonin (SHI), from Traditional Chinese Medicine, has revealed promising antitumor activity in several tumor entities. In the current study, the impact of SHI on four therapy-sensitive and four respective DX-resistant PCa cell lines was determined. SHI induced growth inhibition mainly by necroptosis, a type of cell death, in all the tested therapy-sensitive, but more importantly, DX-resistant PCa cell lines. Corresponding molecular alterations contributing to growth inhibition after SHI exposure were found. SHI could, therefore, be a promising additive in treating advanced PCa.The prognosis for advanced prostate carcinoma (PCa) remains poor due to development of therapy resistance, and new treatment options are needed. Shikonin (SHI) from Traditional Chinese Medicine has induced antitumor effects in diverse tumor entities, but data related to PCa are scarce. Therefore, the parental (=sensitive) and docetaxel (DX)-resistant PCa cell lines, PC3, DU145, LNCaP, and 22Rv1 were exposed to SHI [0.1–1.5 μM], and tumor cell growth, proliferation, cell cycling, cell death (apoptosis, necrosis, and necroptosis), and metabolic activity were evaluated. Correspondingly, the expression of regulating proteins was assessed. Exposure to SHI time- and dose-dependently inhibited tumor cell growth and proliferation in parental and DX-resistant PCa cells, accompanied by cell cycle arrest in the G2/M or S phase and modulation of cell cycle regulating proteins. SHI induced apoptosis and more dominantly necroptosis in both parental and DX-resistant PCa cells. This was shown by enhanced pRIP1 and pRIP3 expression and returned growth if applying the necroptosis inhibitor necrostatin-1. No SHI-induced alteration in metabolic activity of the PCa cells was detected. The significant antitumor effects induced by SHI to parental and DX-resistant PCa cells make the addition of SHI to standard therapy a promising treatment strategy for patients with advanced PCa.

Salvage chemotherapy with cisplatin, ifosfamide, and paclitaxel in metastatic castration-resistant prostate cancer.

123 Background: Recent developments in the treatment of metastatic castration resistant prostate cancer (mCRPC) lead to a revival of platinum-based chemotherapy demonstrating increased activity in patients with aggressive variants of disease. Here, we report on the results of a combinational salvage therapy with cisplatin, ifosfamide and paclitaxel (TIP) in mCRPC. Methods: We retrospectively analyzed patients with mCRPC treated with TIP at the University Hospital Hamburg-Eppendorf between November 2013 and September 2020. Accompanying in vitro analyses were performed using human prostate carcinoma cell lines harboring different levels of drug resistance including the docetaxel-resistant sublines PC3-DR and DU45-DR. Results: In total, 17 mCRPC patients treated with TIP were eligible for efficacy analyses with a median age of 65 yrs. At baseline, liver metastases were present in 88%, metastases of other visceral sides (lung, adrenal gland, brain) in 47% and bone metastases in 76% of the patients. Median hemoglobin was 9.8mg/dl, LDH 903 U/l and AP 205 U/l. Median PSA value was 77 ng/ml covering a wide range including three patients with a PSA-value below 1ng/ml. NSE was evaluated in 83% of the patients (median 38,5 U/l). Patients were extensively pretreated with a median of three treatment lines before TIP (100% docetaxel, 82% abirateron and/or enzalutamide, 47% cabazitaxel, 41% others). A median of 3,5 cycles of TIP were applied with 29% of the patients receiving the maximum of 6 cycles. Four patients discontinued treatment due to side effects (PNP, infection, ifosfamide induced psychosis). At interim analyses, 59 % of the patients showed a radiological response or stable disease with only one patient progressing till the end of treatment. Median PFS was 2.5 months, median OS 6 months. A decrease of PSA > 30% and LDH > 50% was observed in 41% and 35% of the patients, respectively. In vitro experiments revealed additive effects of TIP in 22Rv1, LNCaP and DU45 cells and synergistic effects in neuroendocrine LASCPC-01 cells. In PC3 cells, TIP induced antagonistic effects at lower doses, whereas dose-independent additive effects were observed in docetaxel-resistant PC3-DR. Surprisingly, preliminary data of combined therapies with different drug pairs suggest an antagonistic effect of paclitaxel in the combination with both, cisplatin and ifosfamide. Conclusions: Combinational therapy with cisplatin, ifosfamide and paclitaxel showed promising activity in some patients with aggressive mCRPC. Preclinical data suggest that the drug combination of cisplatin and ifosfamide rule the efficacy of TIP in mCRPC.

Cytotoxic and apoptotic potential of some coumarin and 2-amino-3-carbonitrile selenophene derivatives in prostate cancer.

3-acetyl coumarin derivatives (1a-d) are formed as a result of condensation of salicylaldehyde derivatives and ethyl acetoacetate and were converted into coumarin-selenophene hybrid compounds (2a-d) in the basic medium by modified Gewald reaction in the presence of malononitrile and selenium. Products are characterized by nuclear magnetic resonance (NMR). The prepared compounds are screened for their anticancer activity against DU-145 cell line. In addition, selected target compounds are evaluated for apoptosis and oxidative stress on DU-145 (prostate carcinoma) cell lines.

Increased Expression of AKT3 in Neuroendocrine Differentiated Prostate Cancer Cells Alters the Response Towards Anti-Androgen Treatment.

Simple SummaryMetastatic castration-resistant prostate cancer is still untreatable, and patients have a very short life expectancy after diagnosis. One factor that makes metastatic castration-resistant prostate cancer so aggressive and difficult to treat is neuroendocrine differentiation of prostate carcinoma cells. We already showed that neuroendocrine differentiation of LNCaP cells results in increased AKT3 expression. The aim of this study was to demonstrate the role of AKT3 in neuroendocrine differentiation. Therefore, the previously obtained in vitro data were validated and extended to tissue from patient with neuroendocrine prostate cancer, where we show the presence of AKT3 in neuroendocrine cells. Furthermore, we demonstrate the oncogenic consequences of an increased AKT3 expression including inactivation of apoptotic proteins and a potential role of some miR-17 family members, negatively regulating AKT3, in neuroendocrine differentiation.Patients with advanced prostate carcinoma are often treated with an androgen deprivation therapy but long-term treatment can result in a metastatic castration-resistant prostate cancer. This is a more aggressive, untreatable tumor recurrence often containing areas of neuroendocrine differentiated prostate cancer cells. Using an in vitro model of NE-like cancer cells, it could previously be shown that neuroendocrine differentiation of LNCaP cells leads to a strong deregulation of mRNA and miRNA expression. We observe elevated RNA and protein levels of AKT Serine/Threonine Kinase 3 (AKT3) in neuroendocrine-like LNCaP cells. We used prostate resections from patients with neuroendocrine prostate cancer to validate these results and detect a co-localization of neuroendocrine marker genes with AKT3. Analysis of downstream target genes FOXO3A and GSK3 strengthens the assumption AKT3 may play a role in neuroendocrine differentiation. Overexpression of AKT3 shows an increased survival rate of LNCaP cells after apoptosis induction, which in turn reflects the significance in vivo or for treatment. Furthermore, miR-17, −20b and −106b, which are decreased in neuroendocrine-like LNCaP cells, negatively regulate AKT3 biosynthesis. Our findings demonstrate AKT3 as a potential therapeutic target and diagnostic tool in advanced neuroendocrine prostate cancer and a new mRNA–miRNA interaction with a potential role in neuroendocrine differentiation of prostate cancer.

Bluetongue Viruses Act as Novel Oncolytic Viruses to Effectively Inhibit Human Renal Cancer Cell Growth In Vitro and In Vivo.

BackgroundThe bluetongue virus (BTV) is the prototype virus in the genus Orbivirus within the family Reoviridae. Recent studies indicate that BTVs are capable of infecting and selectively lysing human hepatic carcinoma cells (Hep-3B) and prostate carcinoma cells (pc-3). This study was designed to evaluate the oncolytic potential of BTV in experimental models of human renal cancer in vitro and in vivo.Material/MethodsFive human renal cancer cell lines, ACHN, CAKI-1, OS-RC-2, 786-O, and A498, were used in this study to analyze BTV replication. These cells were lysed by oncolysis compared to normal control. Xenograft models were used to assess the efficacy and toxicity of BTVs in vivo. Data were analyzed by one-way ANOVA or two-sided unpaired t tests.ResultsThe results showed HPTEC cells to be relatively resistant to cytotoxic effects of BTVs and exhibited normal growth rate even at high dose of BTVs. Nonetheless, the renal cancer cells showed a remarkably higher sensitivity to BTVs. Moreover, the ultramicroscopic subcellular changes were also detected in the renal cells. The viral particles were observed in all the RCC cell lines, but not in HPTEC cells. Intratumoral injections of BTVs significantly decreased the tumor volume as compared to animals that received no virus treatment. Infection with BTVs significantly increased the percentage of apoptotic renal cancer cells but not the HPTEC cells. Moreover, BTV triggered apoptosis in renal cancer cells via a mitochondria-mediated pathway.ConclusionsThis study for the first time demonstrated the oncolytic potential of BTV in experimental models of human renal cancer. BTV exhibits the potential to inhibit human renal cancer cell growth in vitro and in vivo.

Phytochemical Characterization, In Vitro Anti-Inflammatory, Anti-Diabetic, and Cytotoxic Activities of the Edible Aromatic Plant; Pulicaria jaubertii.

Pulicaria jaubertii is a medicinal herb that alleviates inflammations and fever. Chromatographic separation, phytochemical characterization, and in vitro biological activities of the plant n-hexane extract were conducted for the first time in this study. Six compounds were isolated for the first time from the n-hexane fraction of Pulicaria jaubertii aerial parts and were identified on the bases of NMR and MS analyses as pseudo-taraxaterol (1), pseudo-taraxasterol acetate (2), 3β-acetoxytaraxaster-20-en-30-aldehyde (3), calenduladiol-3-O-palmitate (4), stigmasterol (5), and α-tocospiro B (6). Compound (6) was a rare tocopherol-related compound and was isolated for the first time from family Asteraceae, while compound (3) was isolated for the first time from genus Pulicaria. The total alcoholic extract and n-hexane fraction were tested for their anti-inflammatory, antidiabetic, and cytotoxic activities. The n-hexane fraction has dose dependent red blood cells (RBCs) membrane stabilization and inhibition of histamine release activities with IC50: 60.8 and 72.9 µg/mL, respectively. As antidiabetic activity, the alcoholic extract exerted the most inhibition on the activity of yeast α-glucosidase, with an IC50: 76.8 µg/mL. The n-hexane fraction showed cytotoxic activity against hepatocarcinoma (HepG-2), breast carcinoma (MCF-7), and prostate carcinoma (PC-3) cell lines with IC50: 51.8, 90.8 and 62.2 µg/mL, respectively. In conclusion, the anti-inflammatory effect of Pulicaria jaubertii might be attributed to the triterpenoid constituents of the n-hexane extract of the plant.

Functional Impact of Acidosis-Regulated MicroRNAs on the Migration and Adhesion of Tumor Cells.

Tumor tissue shows special features in metabolism in contrast to healthy tissue. Besides a distinctive oxygen deficiency, tumors often show a reduced extracellular pH (acidosis) resulting from an intensified glycolysis not only under hypoxic but also under normoxic conditions (Warburg effect). As shown in previous studies, cell migration is increased in AT1 prostate carcinoma cells after incubation at pH6.6, and this leads to an increased number of lung metastases in vivo. However, the signaling pathway causing these functional changes is still unknown. Possible mediators could be acidosis-regulated microRNAs (miR-7, miR-183, miR-203, miR-215). The aim of the study was therefore to analyze whether a change in the expression of these microRNAs has an impact on the tumor cell migration and adhesion. Studies were performed with AT1 rat prostate cancer cells which were incubated for 24h at pH7.4 or 6.6. Keeping AT1 tumor cells at low pH increased the migratory capacity by about 100%. But also the decrease of miR-203 and miR-215 expression (at normal pH) led to an increase in migration velocity by 50%. In contrast, cell adhesion was increased by about 75% at low pH. However, an increase in miR-215 expression at pH6.6 reduced the adhesion by trend. These results clearly indicated that the extracellular pH has an impact on migration and adhesion of tumor cells. In this mechanism, pH-regulated microRNAs could play a role since changes in the expression of these microRNAs (especially miR-203) are also able to modulate the migratory behavior.

Effects of Schisandrae Fructus and Corni Fructus Extracts on the Proliferation and Expression of Prostatic Hyperplasia-inducing Factors in Dihydrotestosterone-stimulated LNCaP Human Prostate Carcinoma Cells

Effects of Schisandrae Fructus and Corni Fructus Extracts on the Proliferation and Expression of Prostatic Hyperplasia-inducing Factors in Dihydrotestosterone-stimulated LNCaP Human Prostate Carcinoma Cells

Co-localisation of intra-nuclear membrane type-1 matrix metalloproteinase and hypoxia inducible factor-2α in osteosarcoma and prostate carcinoma cells.

Increased membrane type-1 matrix metalloproteinase (MT1-MMP) expression in osteosarcoma is predictive of poor prognosis and directs bone metastasis in prostate carcinoma. MT1-MMP subcellular localisation varies with oxygen tension, and, therefore, the aim of the present study was to assess protein interactions between MT1-MMP and the hypoxia inducible factors (HIF-1α and HIF-2α). MT1-MMP protein expression was investigated across a panel of cancer cell lines, including a positive and negative control. The hypoxia-induced alteration in subcellular location of MT1-MMP, HIF-1α and HIF-2α in the U2OS osteosarcoma cell line was assessed using subcellular fractionation. A proximity ligation assay was utilised to assess protein to protein interactions in the osteosarcoma U2OS and prostate carcinoma PC3 cell lines. U2OS and PC3 cells exhibited a significantly increased intra-nuclear interaction between MT1-MMP and HIF-2α in response to hypoxia. The role of this warrants further investigation as it may unveil novel opportunities to target MT1-MMP, which is of particular significance for osteosarcoma since current treatment options are limited.

An IgG‐based bispecific antibody for improved dual targeting in PSMA‐positive cancer

The prostate‐specific membrane antigen (PSMA) has been demonstrated in numerous studies to be expressed specifically on prostate carcinoma cells and on the neovasculature of several other cancer entities. However, the simultaneous expression of PSMA on both, tumor cells as well as tumor vessels remains unclear, even if such “dual” expression would constitute an important asset to facilitate sufficient influx of effector cells to a given tumor site. We report here on the generation of a PSMA antibody, termed 10B3, which exerts superior dual reactivity on sections of prostate carcinoma and squamous cell carcinoma of the lung. 10B3 was used for the construction of T‐cell recruiting bispecific PSMAxCD3 antibodies in Fab‐ and IgG‐based formats, designated Fabsc and IgGsc, respectively. In vitro, both molecules exhibited comparable activity. In contrast, only the larger IgGsc molecule induced complete and durable elimination of established tumors in humanized mice due to favorable pharmacokinetic properties. Upon treatment of three patients with metastasized prostate carcinoma with the IgGsc reagent, marked activation of T cells and rapid reduction of elevated PSA levels were observed.

Genomic GLO1 deletion modulates TXNIP expression, glucose metabolism, and redox homeostasis while accelerating human A375 malignant melanoma tumor growth

Glyoxalase 1 (encoded by GLO1) is a glutathione-dependent enzyme detoxifying the glycolytic byproduct methylglyoxal (MG), an oncometabolite involved in metabolic reprogramming. Recently, we have demonstrated that GLO1 is overexpressed in human malignant melanoma cells and patient tumors and substantiated a novel role of GLO1 as a molecular determinant of invasion and metastasis in melanoma. Here, employing NanoString™ gene expression profiling (nCounter™ ‘PanCancer Progression Panel’), we report that CRISPR/Cas 9-based GLO1 deletion from human A375 malignant melanoma cells alters glucose metabolism and redox homeostasis, observable together with acceleration of tumorigenesis. Nanostring™ analysis identified TXNIP (encoding thioredoxin-interacting protein), a master regulator of cellular energy metabolism and redox homeostasis, displaying the most pronounced expression change in response to GLO1 elimination, confirmed by RT-qPCR and immunoblot analysis. TXNIP was also upregulated in CRISPR/Cas9-engineered DU145 prostate carcinoma cells lacking GLO1, and treatment with MG or a pharmacological GLO1 inhibitor (TLSC702) mimicked GLO1_KO status, suggesting that GLO1 controls TXNIP expression through regulation of MG. GLO1_KO status was characterized by (i) altered oxidative stress response gene expression, (ii) attenuation of glucose uptake and metabolism with downregulation of gene expression (GLUT1, GFAT1, GFAT2, LDHA) and depletion of related key metabolites (glucose-6-phosphate, UDP-N-acetylglucosamine), and (iii) immune checkpoint modulation (PDL1). While confirming our earlier finding that GLO1 deletion limits invasion and metastasis with modulation of EMT-related genes (e.g. TGFBI, MMP9, ANGPTL4, TLR4, SERPINF1), we observed that GLO1_KO melanoma cells displayed a shortened population doubling time, cell cycle alteration with increased M-phase population, and enhanced anchorage-independent growth, a phenotype supported by expression analysis (CXCL8, CD24, IL1A, CDKN1A). Concordantly, an accelerated growth rate of GLO1_KO tumors, accompanied by TXNIP overexpression and metabolic reprogramming, was observable in a SCID mouse melanoma xenograft model, demonstrating that A375 melanoma tumor growth and metastasis can be dysregulated in opposing ways as a consequence of GLO1 elimination.

Selective cytotoxic and anti-metastatic activity in DU-145 prostate cancer cells induced by Annona muricata L. bark extract and phytochemical, annonacin

BackgroundAnnona muricata L. was identified as a popular medicinal plant in treatment regimens among cancer patients in Jamaica by a previously conducted structured questionnaire. Ethnomedically used plant parts, were examined in this study against human prostate cancer cells for the first time and mechanisms of action elucidated for the most potent of them, along with the active phytochemical, annonacin.MethodsNine extracts of varying polarity from the leaves and bark of A. muricata were assessed initially for cytotoxicity using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay on PC-3 prostate cancer cells and the ethyl acetate bark (EAB) extract was identified as the most potent. EAB extract was then standardized for annonacin content using High-performance Liquid Chromatography - Mass Spectrometry (HPLC-MS) and shown to be effective against a second prostate cancer cell line (DU-145) also. The mode of cell death in DU-145 cells were assessed via several apoptotic assays including induction of increased reactive oxygen species (ROS) production, reduction of mitochondrial membrane potential, activation of caspases and annexin V externalization combined with morphological observations using confocal microscopy. In addition, the potential to prevent metastasis was examined via inhibition of cell migration, vascular endothelial growth factor (VEGF) and angiogenesis using the chorioallantoic membrane assay (CAM).ResultsAnnonacin and EAB extract displayed selective and potent cytotoxicity against the DU-145 prostate carcinoma cells with IC50 values of 0.1 ± 0.07 μM and 55.501 ± 0.55 μg/mL respectively, without impacting RWPE-1 normal prostate cells, in stark contrast to chemotherapeutic docetaxel which lacked such selectivity. Docetaxel’s impact on the cancerous DU-145 was improved by 50% when used in combination with EAB extract. Insignificant levels of intracellular ROS content, depolarization of mitochondrial membrane, Caspase 3/7 activation, annexin V content, along with stained morphological evaluations, pointed to a non-apoptotic mode of cell death. The extract at 50 μg/mL deterred cell migration in the wound-healing assay, while inhibition of angiogenesis was displayed in the CAM and VEGF inhibition assays for both EAB (100 μg /mL) and annonacin (0.5 μM).ConclusionsTaken together, the standardized EAB extract and annonacin appear to induce selective and potent cell death via a necrotic pathway in DU-145 cells, while also preventing cell migration and angiogenesis, which warrant further examinations for mechanistic insights and validity in-vivo.

Effect of myeloperoxidase on the anticoagulant activity of low molecular weight heparin and rivaroxaban in an in vitro tumor model

BackgroundInflammation with leukocyte activation is a hallmark of cancer‐associated thrombosis (CAT), and elevated leukocytes predict venous thromboembolism in cancer outpatients. In a recent trial, rivaroxaban was more efficacious than dalteparin in preventing CAT recurrence. ObjectivesIn a proof‐of‐concept study, we aimed to provide a mechanistic basis for improved efficacy of rivaroxaban compared to low molecular weight heparin in CAT treatment. MethodsWe studied the effects of rivaroxaban, dalteparin, and tinzaparin at peak and trough levels on tumor cell–induced procoagulant activity and platelet aggregation in the presence or absence of the cationic leukocyte‐derived enzyme, myeloperoxidase (MPO). Furthermore, pro‐inflammatory conditions were generated by stimulating whole blood with lipopolysaccharide (LPS) or phorbol‐myristate‐acetate (PMA), before measuring thrombin generation in plasma supernatants. ResultsAll three anticoagulants inhibited thrombin generation, fibrin clot formation, and platelet aggregation induced by the tissue factor‐expressing prostate carcinoma cell line, 22Rv1. Pre‐incubation with MPO partially attenuated the anticoagulant activity of dalteparin and tinzaparin, but not rivaroxaban, at trough levels. The effect of MPO did not involve the enzyme's catalytic properties, but required its structural integrity, as indicated by heat denaturation. In plasma obtained from LPS‐ or PMA‐stimulated whole blood, elevated MPO antigen levels inversely correlated with the ability of tinzaparin to inhibit 22Rv1‐induced thrombin generation. ConclusionsMyeloperoxidase release may partially attenuate the anticoagulant activity of trough levels of dalteparin and tinzaparin in the context of paraneoplastic leukocyte activation. However, this effect is likely not sufficient to explain the improved efficacy of rivaroxaban, and possibly other oral factor Xa inhibitors, in CAT treatment.

Dual EGFR/VEGFR2 inhibitors and apoptosis inducers: Synthesis and antitumor activity of novel pyrazoline derivatives.

Novel derivatives of the pyrazoline scaffold were synthesized and investigated for their cytotoxicity against prostate (PC-3), hepatocellular (HepG2), and breast (MDA-MB-231) carcinoma cells. The most active compounds,4a, 4b, 5b, and 7c, revealed significant and broad-spectrum anticancer activities with IC50 values of 1.30-7.18 μM in comparison with doxorubicin (IC50 = 5.12-7.33 μM). Additionally, they exhibited lower cytotoxicity against normal WI-38 cells, indicating their high safety profiles. Aiming to enlighten the inhibitory potential on receptor tyrosine kinases (RTKs), compounds 4a, 4b, 5b, and 7c were assessed for their activities against four different RTKs (EGFR, FGFR2, HER2, and VEGFR2) and their apoptotic potencies on PC-3 cells. The results revealed that compounds 5b and 7c are potent dual EGFR and VEGFR2 inhibitors (IC50 = 0.21 and 0.23 μM, respectively, against EGFR; 0.22 and 0.21 μM, respectively, against VEGFR2), whereas they displayed moderate inhibitory activities against HER2 and FGFR2. Besides, compounds 4a, 4b, 5b, and 7c prompted apoptosis via theupregulation of Bax, p53, and caspase-3, together with the downregulation of the levels of Bcl-2. Also, it was found that compounds 5b and 7c are more potent as apoptosis inducers than the other tested derivatives. Furthermore, molecular docking analyses of compounds 4a, 4b, 5b, and 7c in the EGFR and VEGFR ATP binding sites were performed, to confirm the in vitro assays.

2-Amino-nicotinamide induces apoptosis of prostate cancer cells via inhibition of PI3K/AKT and phosphorylation of STA3/JAK2

Purpose: To study the cytotoxicity of 2-amino-nicotinamide against prostate cancer (PCa) cells, and the underlying molecular mechanism.Methods: The effect of 2-amino-nicotinamide on cell viability and apoptosis was determined by 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) and flow cytometry, respectively, while its effect on cellular production of fluorescent-oxidized product from DCFH-DA was measured using flow cytometry. Apoptosis-related protein expressions were evaluated by western blot assay.Results: 2-Amino-nicotinamide produced cytotoxicity against MCF-7, SGC7901, PCa 22Rv1 and LNCaP cancer cell lines (p < 0.05). Mechanistic data revealed that 2-amino-nicotinamide activated apoptosis, and enhanced cleavage of PARP and caspase-3 in PCa 22Rv1 and LNCaP cells. In PCa 22Rv1 and LNCaP cell lines, cytochrome C and Bax levels were enhanced by treatment with 2-aminonicotinamide, while Bcl-2 protein level was suppressed (p < 0.05). Activated expressions of PI3K, Akt and ERK in PCa 22Rv1 and LNCaP cells were down-regulated, while p38 expression was increased.Moreover, 2-amino-nicotinamide suppressed the activation of JAK2 and STAT3, but did not alter total JAK2 and STAT3 levels in PCa 22Rv1 and LNCaP cells (p < 0.05).Conclusion: 2-Amino-nicotinamide exerts cytotoxic effects on prostate carcinoma cells via activation of apoptosis and down-regulation of PI3K/AKT and STA3/JAK2. Thus, 2-amino nicotinamide is a potential bioactive agent for prostate cancer management.
 Keywords: 2-Amino-nicotinamide, Apoptosis, Fluorescent-oxidized, Cytotoxicity

8-Epi-xanthatin induces the apoptosis of DU145 prostate carcinoma cells through signal transducer and activator of transcription 3 inhibition and reactive oxygen species generation.

Signal transducer and activator of transcription 3 (STAT3) is aberrantly activated in many human cancers. We tried to find STAT3 inhibitors from natural sources and found that Xanthium fruit extracts decreased phosphorylation of STAT3-Y705. 8-Epi-xanthatin (EXT) was isolated from the extracts. When DU145 cancer cells were treated with EXT, p-STAT3-Y705 was decreased with an IC50 of 3.2 μM. EXT decreased the expression of STAT3 target genes, such as cyclin A, cyclin D1, and BCL-2, and induced PARP cleavage, indicating apoptotic cell death. Downregulation of EXT-induced p-STAT3-Y705 was rescued by pretreating DU145 cells with antioxidants, such as N-acetyl-L-cysteine (NAC), indicating that reactive oxygen species (ROS) were involved in the EXT-induced inhibition of STAT3 activation. Furthermore, we proved the association of EXT with STAT3 protein by using a drug affinity responsive target stability (DARTS) assay and a cellular thermal shift assay (CETSA). EXT inhibited proliferation of DU145 cells with a GI50 of 6 μM and reduced tumor growth in mice xenografted with DU145 cells. Immunoblotting showed that phosphorylation of STAT3-Y705 was lower in EXT-treated tumor tissue than in control tissues. Collectively, we found that EXT binds to, and inhibits, STAT3 activation and could be a lead compound for anticancer therapy.

Fluorescent Products of Anthocyanidin and Anthocyanin Oxidation.

We found that the oxidation of cyanidin with hydrogen peroxide, tert-butyl hydroperoxide, 2,2'-azobis(2-methylpropionamidine), 3-morpholinosydnonimine hydrochloride, sodium hypochlorite, Fe3+, and Fe2+ induced the appearance of a new fluorescence band, centered at 525-540 nm when excited at 470-490 nm. The intensity of this fluorescence was related to the oxidant concentration. The same fluorescence was induced by the oxidation of other anthocyanidins, an anthocyanin (cyanidin-3-O-glucoside) as well as anthocyanin-rich red cabbage extract, and blackcurrant and elderberry juices. Peroxidized blood plasma also induced the appearance of cyanidin fluorescence. We also showed that the formation of the fluorescent product of pelargonidin can be observed in a culture of the MRC-5 human fetal lung fibroblast cell line and the DU-145 human prostate carcinoma cell line. Our results suggest that this new anthocyanidin/anthocyanin fluorescence may be an indicator of oxidation, especially of food products, where these compounds are present or added as colorants, and can also be useful to detect oxidation in biomedical experiments.

Corilagin Represses Epithelial to Mesenchymal Transition Process Through Modulating Wnt/β-Catenin Signaling Cascade.

Corilagin (CLG), a major component of several medicinal plants, can exhibit diverse pharmacological properties including those of anti-cancer, anti-inflammatory, and hepatoprotective qualities. However, there are no prior studies on its potential impact on the epithelial-to-mesenchymal transition (EMT) process. EMT can lead to dissemination of tumor cells into other organs and promote cancer progression. Hence, we aimed to investigate the effect of CLG on EMT and its mechanism(s) of action in tumor cells. We noted that CLG reduced the expression of various epithelial markers and up-regulated the expression of Occludin and E-cadherin in both basal and TGFβ-stimulated tumor cells. CLG treatment also abrogated cellular invasion and migration in colon and prostate carcinoma cells. In addition, CLG effectively attenuated the Wnt/β-catenin signaling cascade in TGFβ-stimulated cells. Overall, our study suggests that CLG may function as and effective modulator of EMT and metastasis in neoplastic cells.

Imaging of Cell Structures Using Optimized Soft X-ray Contact Microscopy

This work is to study the relationship between the exposure conditions and the quality of cell imaging with soft X-ray contact microscopy (SXCM). It is a crucial step in the efficient visualization of cell structures. Three different human cell lines: DU145 prostate carcinoma cells, HCC38 breast cancer cells, and Poietics mesenchymal stem cells were used to establish the optimal exposure conditions in SXCM. The image quality depended on the soft X-ray (SXR) absorbed energy and photoresist development conditions. At lower SXR energy (200 or 400 SXR pulses), sharp cell edges, membrane projections, and cell–cell connections were visible. In contrast, higher energy (600 or 800 SXR pulses) allowed observation of the cytoskeleton and the nucleus in a cell type-dependent manner (the influence of cell thickness and internal complexity was noted).

Differential anti-proliferative and apoptotic effects of lichen species on human prostate carcinoma cells.

Lichens are stable symbiotic associations between fungus and algae and/or cyanobacteria that have different biological activities. Around 60% of anti-cancer drugs are derived from natural resources including plants, fungi, sea creatures, and lichens. This project aims to identify the apoptotic effects and proliferative properties of extracts of Bryoria capillaris (Ach.) Brodo & D.Hawksw, Cladonia fimbriata (L.) Fr., Evernia divaricata (L.) Ach., Hypogymnia tubulosa (Schaer.) Hav., Lobaria pulmonaria (L.) Hoffm., and Usnea florida (L.) Weber ex Wigg. lichen species on prostate cancer cells. Lichen extracts were performed by ethanol, methanol, and acetone separately by using the Soxhlet apparatus and the effects of the extracts on cell viability, proliferation, and apoptosis were measured with the utilization of MTT, LDH assay, Annexin V assay, and Western Blot. Findings of our study revealed a positive correlation between the elevation of cell sensitivity and the increase in the treatment doses of the extract in that higher doses applied reverberate to higher cell sensitivity. A similar correlation was also identified between cell sensitivity elevation and the duration of the treatment. Evidence in our study have shown the existence of an anti-proliferative effect in the extracts of Bryoria capillaris, Evernia divaricata (L.) Ach., Hypogymnia tubulosa (Schaer.) Hav., Lobaria pulmonaria (L.) Hoffm., and Usnea florida (L.) Weber ex Wigg., while a similar effect was not observed in the extracts of Cladonia fimbriata. Evernia divaricata induced anti-proliferative and apoptotic effects in PC-3 cells, which induced apoptotic cell death by both extrinsic and intrinsic pathways. Hypogymnia tubulosa has been shown to have anti-proliferative and apoptotic effects in all extractions methods and our findings identified that both the percentage of the apoptotic cells and apoptotic protein expressions recorded an increase at lower treatment concentrations. Although Lobaria pulmonaria is known to have significant cytotoxic effects, we did not observe a decrease in cell proliferation. Indeed, proliferation marker proliferating cell nuclear antigen (PCNA) protein expression levels have shown an increase in all extracts, while Usnea florida exhibited apoptosis induction and slight proliferation reduction in extract treatments with lower concentrations. We tested 18 extracts of six lichen species during our study. Of these, Evernia divaricata and Hypogymnia tubulosa demonstrated significant apoptotic activity on prostate cancer cells including at low concentrations, which implies that it is worth pursuing the biologically active lead compounds of these extracts on prostate cancer in vitro. Further corroboratory studies are needed to validate the relative potential of these extracts as anti-metastatic and anti-tumorigenic agents.