Procyanidin trimer C1 derived from Theobroma cacao reactivates latent human immunodeficiency virus type 1 provirus.

Despite remarkable advances in combination antiretroviral therapy (cART), human immunodeficiency virus type 1 (HIV-1) infection remains incurable due to the incomplete elimination of the replication-competent virus, which persists in latent reservoirs. Strategies for targeting HIV reservoirs for eradication that involves reactivation of latent proviruses while protecting uninfected cells by cART are urgently needed for cure of HIV infection. We screened medicinal plant extracts for compounds that could reactivate the latent HIV-1 provirus and identified a procyanidin trimer C1 derived from Theobroma cacao as a potent activator of the provirus in human T cells latently infected with HIV-1. This reactivation largely depends on the NF-κB and MAPK signaling pathways because either overexpression of a super-repressor form of IκBα or pretreatment with a MEK inhibitor U0126 diminished provirus reactivation by C1. A pan-PKC inhibitor significantly blocked the phorbol ester-induced but not the C1-induced HIV-1 reactivation. Although C1-induced viral gene expression persisted for as long as 48 h post-stimulation, NF-κB-dependent transcription peaked at 12 h post-stimulation and then quickly declined, suggesting Tat-mediated self-sustainment of HIV-1 expression. These results suggest that procyanidin C1 trimer is a potential compound for reactivation of latent HIV-1 reservoirs.

Potential Medicinal Efficacy of Alkaline Extract of Pine Seed Shell: Anti-UVC Activity and Macrophage Activation.

BACKGROUND/AIM: Alkaline extracts of several plants which contain lignin degradation products have several unique biological activities. In order to search for new biological activities of alkaline extracts of pine seed shell (APs), their anti-ultraviolet C (UVC) and macrophage stimulation activity were investigated. MATERIALS AND METHODS: Anti-UVC activity was determined by the ratio of the 50% cytotoxic concentration against human melanoma cell line COLO679 to the 50% UVC-protective concentration. Extracellularly secreted nitrite (NO2 -) by unstimulated and lipopolysaccharide (LPS)-stimulated mouse macrophage-like cells RAW264.7 was determined by Griess method. RESULTS: APs showed significantly higher anti-UVC activity than previously reported hot-water extracts of medical herbs. Anti-UVC activity of AP and vanillic acid was maintained for much longer than that of sodium ascorbate and vanillin. APs enhanced the production of NO2 - to the level induced by LPS. Simultaneous addition of AP and LPS did not further increase NO2 - production, suggesting their mechanisms of action overlap. CONCLUSION: The present study suggests the possible application of APs as UVC protectors and immunopotentiators via macrophage activation.

Potentiation of Anticancer Activity of G2/M Blockers by Mild Hyperthermia.

BACKGROUND/AIM: Hyperthermia (HT), combined with chemotherapy, has been used to treat various types of cancer. This study aimed to investigate the HT-sensitivity of malignant and non-malignant cells, and then evaluate the combination effect of docetaxel (DTX) and a newly synthesized chromone derivative (compound A) with HT. MATERIALS AND METHODS: The number of viable cells was determined using the MTT method. Cell cycle distribution was analyzed using a cell sorter, and DNA fragmentation pattern was detected using agarose gel electrophoresis. RESULTS: Among 12 cultured cells, oral squamous cell carcinoma (OSCC), especially Ca9-22 cells, and myelogenous leukemia cells showed higher sensitivity to HT than lung carcinoma and glioblastoma cell lines, while normal oral cells were the most resistant. Cytotoxicity of DTX on Ca9-22 cells was maximum at 41-42°C and 45~60 min exposure to HT. DXT, compound A, and HT induced G2/M arrest of Ca-22 cells. Mild HT enhanced the DTX- and compound A-induced subG1 arrest, in a synergistic fashion. CONCLUSION: The combination G2/M blockers and mild-HT can potentially be used for the treatment of OSCC.

UVC-Protective Activity of Lemongrass Among 12 Fat-soluble Herbal Extracts: Rapid Decay Due to Cytotoxicity.

BACKGROUND/AIM: The COVID-19 pandemic led to the rapid spread of the use of ultraviolet C (UVC) sterilizers in many public facilities. Considering the harmful effects of prolonged exposure to UVC, manufacturing of safe skin care products is an important countermeasure. In continuation of our recent study of water-soluble herbal extracts, the present study aimed at searching for anti-UVC components from fat-soluble herbal extracts. MATERIALS AND METHODS: Human dermal fibroblast and melanoma cells were exposed to UVC (1.193 W/m2) for 3 min. Viable cell number was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell-cycle analysis was performed using a cell sorter. UVC-protective activity was quantified by the selective index (SI), i.e., the ratio of the 50% cytotoxic concentration for unirradiated cells to the concentration that restored viability of UVC-treated cells by 50%. RESULTS: Only lemongrass extract, among 12 fat-soluble herbal extracts, showed significant anti-UVC activity, comparable to that of lignified materials and tannins, but exceeding that of N-acetyl-L-cysteine and resveratrol. Lemongrass extract was highly cytotoxic, producing a subG1 cell population. During prolonged incubation in culture medium, the anti-UVC activity of lemongrass extract, sodium ascorbate and vanillic acid declined with an approximate half-life of <0.7, 5.4-21.6, and 27.8-87.0 h, respectively. CONCLUSION: Removal of cytotoxic principle(s) from lemongrass extract is crucial to producing long-lasting UVC-protective effects.

Comprehensive Study of Anti-UVC Activity and Cytotoxicity of Hot-water Soluble Herb Extracts.

BACKGROUND/AIM: COVID-19 pandemic caused the rapid dissemination of ultraviolet C (UVC) sterilization apparatuses. Prolonged exposure to UVC, however, may exert harmful effects on the human body. The aim of the present study was to comprehensively investigate the anti-UVC activity of a total of 108 hot-water soluble herb extracts, using human dermal fibroblast and melanoma cell lines, for the future development of skin care products. MATERIALS AND METHODS: Exposure time to UVC was set to 3 min, and cell viability was determined using the MTT assay. Anti-UVC activity was determined using the selective index (SI), a ratio of 50% cytotoxic concentration for unirradiated cells to 50% effective concentration that restored half of the UVC-induced decrease of viability. RESULTS: Dermal fibroblasts at any population doubling level were more resistant to UVC irradiation than melanoma cells. Both 49 herb extracts recommended by Japan Medical Herb Association (JAMHA) and 59 additional herb extracts showed comparable anti-UVC activity. SI values of selected herbs (Butterbur, Cloves, Curry Tree, Evening Primrose, Rooibos, Stevia, Willow) were several-fold lower than those of vitamin C and vanillin. Their potent anti-UVC activity was maintained for at least 6 h post irradiation, but declined thereafter to the basal level, possibly due to cytotoxic ingredients. CONCLUSION: UVC sensitivity may be related to the growth potential of target cells. Removal of cytotoxic ingredients of herb extracts may further potentiate and prolong their anti-UVC activity.

A Comparative Study of Tumor-Specificity and Neurotoxicity between 3-Styrylchromones and Anti-Cancer Drugs.

Background. Many anti-cancer drugs used in clinical practice cause adverse events such as oral mucositis, neurotoxicity, and extravascular leakage. We have reported that two 3-styrylchromone derivatives, 7-methoxy-3-[(1E)-2-phenylethenyl]-4H-1-benzopyran-4-one (Compound A) and 3-[(1E)-2-(4-hydroxyphenyl)ethenyl]-7-methoxy-4H-1-benzopyran-4-one (Compound B), showed the highest tumor-specificity against human oral squamous cell carcinoma (OSCC) cell lines among 291 related compounds. After confirming their superiority by comparing their tumor specificity with newly synthesized 65 derivatives, we investigated the neurotoxicity of these compounds in comparison with four popular anti-cancer drugs. Methods: Tumor-specificity (TSM, TSE, TSN) was evaluated as the ratio of mean CC50 for human normal oral mesenchymal (gingival fibroblast, pulp cell), oral epithelial cells (gingival epithelial progenitor), and neuronal cells (PC-12, SH-SY5Y, LY-PPB6, differentiated PC-12) to OSCC cells (Ca9-22, HSC-2), respectively. Results: Compounds A and B showed one order of magnitude higher TSM than newly synthesized derivatives, confirming its prominent tumor-specificity. Docetaxel showed one order of magnitude higher TSM, but two orders of magnitude lower TSE than Compounds A and B. Compounds A and B showed higher TSM, TSE, and TSN values than doxorubicin, 5-FU, and cisplatin, damaging OSCC cells at concentrations that do not affect the viability of normal epithelial and neuronal cells. QSAR prediction based on the Tox21 database suggested that Compounds A and B may inhibit the signaling pathway of estrogen-related receptors.

An IκB kinase 2 inhibitor IMD-0354 suppresses the survival of adult T-cell leukemia cells.

Adult T-cell leukemia (ATL) is a fatal T-cell malignancy associated with human T-cell leukemia virus type I infection. The aberrant expression of nuclear factor-κB (NF-κB) is considered to contribute to the malignant phenotype and chemo-resistance of ATL cells. Because of the poor prognosis of ATL, the development of new therapeutic strategies is direly needed. In the present study, we show that an IκB kinase 2 (IKK2) inhibitor, IMD-0354, efficiently inhibits the survival of CD4(+) CD25(+) primary ATL cells and prevents the growth of or induces apoptosis of patient-derived ATL cell lines. Assays of transcription with integrated forms of reporter genes revealed that IMD-0354 suppresses NF-κB-dependent transcriptional activity. Moreover, the daily administration of IMD-0354 prevents the growth of tumors in mice inoculated with ATL cells. Our results suggest that targeting IKK2 with a small molecule inhibitor, such as IMD-0354, is an attractive strategy for the treatment of ATL.

Prominent Anti-UVC Activity of Lignin Degradation Products.

BACKGROUND/AIM: The rapid spread of COVID-19 resulted in the revision of the value of ultraviolet C (UVC) sterilization in working spaces. This study aimed at re-evaluating the anti-UVC activity of four groups of natural products against human melanoma COLO679 and human normal dermal fibroblast (HDFa) cells, based on chemotherapeutic index. MATERIALS AND METHODS: Various cell lines were exposed to UVC for 3 min in the presence of increasing concentrations of test compounds and viable cell numbers were determined with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The anti-UVC activity was quantified by the ratio of the 50% cytotoxic concentration (determined without irradiation) to the 50% effective concentration (which abolished by 50% the UVC-induced loss of viability). Apoptosis was quantified as the subG1 population proportion following cell-cycle analysis. RESULTS: Among four groups of major natural products, six phenylpropanoids showed the highest anti-UVC activity, followed by the lignified products and alkaline products that contain lignin and its degradation products. On the other hand, tannins and flavonoids showed lower activity due to their higher cytotoxicity. UVC-sensitive COLO679 cells lack dectin-1 protein expression. CONCLUSION: These data suggest the prominent anti-UVC activity of lignin degradation products, and the possible involvement of dectin-1 expression in UVC-sensitivity.

Role for protein geranylgeranylation in adult T-cell leukemia cell survival.

Adult T-cell leukemia (ATL) is a fatal lymphoproliferative disease that develops in human T-cell leukemia virus type I (HTLV-I)-infected individuals. Despite the accumulating knowledge of the molecular biology of HTLV-I-infected cells, effective therapeutic strategies remain to be established. Recent reports showed that the hydroxyl-3-methylglutaryl (HMG)-CoA reductase inhibitor statins have anti-proliferative and apoptotic effects on certain tumor cells through inhibition of protein prenylation. Here, we report that statins hinder the survival of ATL cells and induce apoptotic cell death. Inhibition of protein geranylgeranylation is responsible for these effects, since simultaneous treatment with isoprenoid precursors, geranylgeranyl pyrophosphate or farnesyl pyrophosphate, but not a cholesterol precursor squalene, restored the viability of ATL cells. Simvastatin inhibited geranylgeranylation of small GTPases Rab5B and Rac1 in ATL cells, and a geranylgeranyl transferase inhibitor GGTI-298 reduced ATL cell viability more efficiently than a farnesyl transferase inhibitor FTI-277. These results not only unveil an important role for protein geranylgeranylation in ATL cell survival, but also implicate therapeutic potentials of statins in the treatment of ATL.

Inhibition of IkappaB kinase beta restrains oncogenic proliferation of pancreatic cancer cells.

PURPOSE: Pancreatic cancer is characterized by an extremely poor prognosis due to the aggressive disease course and lack of effective therapeutic intervention. IkappaB kinase (IKK), a central kinase for nuclear factor-kappaB (NF-kappaB) activation, is often constitutively activated in pancreatic cancer cells, playing a crucial role in the malignant phenotype and resistance to anti-cancer agents. This study explored how specific inhibition of IKKbeta suppresses oncogenic proliferation of pancreatic cancer cells. EXPERIMENTAL DESIGN: We employed two different approaches, RNA interference-mediated depletion of IKKbeta (IKKbetai) and use of a novel molecularly designed IKKbeta inhibitor IMD-0354 to investigate the effects on the in vitro and in vivo growth and apoptotic response of pancreatic cancer cells. RESULTS: IKKbetai and IMD-0354 efficiently suppressed constitutive NF-kappaB activity and the growth of pancreatic cancer cells in monolayer and soft agar. IMD-0354 induced Annexin V expression, a typical apoptotic cell response. Notably, daily administration of IMD-0354 significantly suppressed tumor growth in NOD/SCID/gamma c(null) (NOG) mice without any deleterious side effect. CONCLUSIONS: These results identify IKKbeta as an attractive molecular target for pancreatic cancer therapy.

Distribution of Human Papillomavirus Genotype in Anal Condyloma Acuminatum Among Japanese Men: The Higher Prevalence of High Risk Human Papillomavirus in Men Who Have Sex with Men with HIV Infection.

Human papillomavirus (HPV) infection is known to cause anal condyloma acuminatum (CA) and squamous cell carcinoma. Men who have sex with men (MSM) with HIV infection are frequently co-infected with HPV, especially high risk HPV (HR-HPV) that causes anal squamous cell carcinoma. However, there are few reports of HPV genotype studies in anal lesion of Japanese men. We tried to estimate the distribution of HPV genotypes in anal CA tissue specimens from the Japanese men to elucidate the risk of anal cancer. A total of 62 patients who had anal CA surgically excised were enrolled. They included 27 HIV-positive MSM, 18 HIV-negative MSM, 1 HIV-positive man who have sex with women (MSW), and 16 HIV-negative MSW. HPV genotypes in anal CA tissue were determined by the polymerase chain reaction technique with reverse line blot hybridization. HR-HPV was detected in 45.2% of the CA tissue specimens and high grade squamous intraepithelial lesion (HSIL) was observed in 15.3%. Moreover, the prevalence of HR-HPV in the HIV-positive MSM (70.4%) was higher than the HIV-negative MSM (33.3%, p = .0311) or the HIV-negative MSW (18.8%, p = .0016). The conditional logistic regression analysis suggested HIV positivity as the primary risk factor for the HR-HPV infection in CA. In addition, HSIL was detected in higher frequency in CA tissues from HIV-positive MSM (25.9%) than HIV-negative MSW (0.0%, p = .0346). HR-HPV and HSIL were frequently detected in anal CA tissues from Japanese MSM patients with HIV infection, suggesting the necessity of surveillance for this population.

Correction: EBV induces persistent NF-κB activation and contributes to survival of EBV-positive neoplastic T- or NK-cells.

[This corrects the article DOI: 10.1371/journal.pone.0174136.].

EBV induces persistent NF-κB activation and contributes to survival of EBV-positive neoplastic T- or NK-cells.

Epstein-Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-κB activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-κB activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-κB activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-κB activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-κB that suppresses NF-κB activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-κB-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells.

Overexpression of NF-κB inducing kinase underlies constitutive NF-κB activation in lung cancer cells.

The present study investigates roles for NF-κB inducing kinase (NIK) in constitutive NF-κB activation in lung cancer cells. A wealth of evidence showed that NF-κB is often constitutively activated in human cancer cells, including non-small cell lung cancer tissue specimens and cell lines, which may lead to deregulated apoptosis and enhanced resistance of tumor cells to chemotherapy. However, the mechanisms of NF-κB activation in lung cancer cells remain largely unknown. We report here that NF-κB inducing kinase (NIK) is aberrantly expressed at the pre-translational level in non-small cell lung cancer (NSCLC) cell lines. Depletion of NIK by RNA interference remarkably diminished nuclear NF-κB DNA binding activity and reporter gene expression. NIK depletion induced apoptosis in A549 cells, reduced the matrix metalloproteinase 9 (MMP-9) and survivin mRNA expression and affected efficiency of anchorage-independent H1299 cell growth, suggesting a role for NIK in the manifestation of oncogenic phenotype. These results indicate that NIK plays a key role in constitutive NF-κB activation in NSCLC cells and implicate NIK as a molecular target for lung cancer therapy.