Metformin, a prescribed drug for type 2 diabetes, has been reported to have anti-cancer effects; however, the underlying mechanism is poorly understood. Here we show that this mechanism may be ...immune-mediated. Metformin enabled normal but not T-cell–deficient SCID mice to reject solid tumors. In addition, it increased the number of CD8 ⁺ tumor-infiltrating lymphocytes (TILs) and protected them from apoptosis and exhaustion characterized by decreased production of IL-2, TNFα, and IFNγ. CD8 ⁺ TILs capable of producing multiple cytokines were mainly PD-1 ⁻Tim-3 ⁺, an effector memory subset responsible for tumor rejection. Combined use of metformin and cancer vaccine improved CD8 ⁺ TIL multifunctionality. The adoptive transfer of antigen-specific CD8 ⁺ T cells treated with metformin concentrations as low as 10 μM showed efficient migration into tumors while maintaining multifunctionality in a manner sensitive to the AMP-activated protein kinase (AMPK) inhibitor compound C. Therefore, a direct effect of metformin on CD8 ⁺ T cells is critical for protection against the inevitable functional exhaustion in the tumor microenvironment.
Significance The multifunctional ability of CTLs is downregulated by interaction between immune-checkpoint molecules expressed on CTLs and their ligands expressed on cancer cells, referred to as immune exhaustion. The antibody-mediated, immune-checkpoint blockade turned out to a promising method for immunotherapy against advanced melanoma. Metformin, a drug prescribed for patients with type 2 diabetes, has been recognized to have anti-cancer effect. We found that CD8 ⁺ tumor infiltrating lymphocytes (TILs) is a target of metformin. CD8 ⁺ TILs inevitably undergo immune exhaustion, characterized by diminished production of multiple cytokines such as IL-2, TNFα, and IFNγ, followed by elimination with apoptosis. Metformin is able to counter the state. Along with conventional therapy, treatment of cancer patients with metformin may have a great advantage for cancer therapy.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
The emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic in 2019 has led to a global health crisis. Mutations of the SARS-CoV-2 genome have impeded the development ...of effective therapeutics and vaccines against SARS-CoV-2. Natural products are important for discovering therapeutics to treat the 2019 coronavirus disease (COVID-19). In the present study, we investigated the antiviral activity of herbal drug extracts from Polygala Root, Areca, and Quercus Bark and natural compounds derived from herbal drug such as baicalin and glabridin, with IC
50
values of 9.5 µg/ml, 1.2 µg/ml, 5.4 µg/ml, 8.8 µM, and 2.5 µM, respectively, against SARS CoV-2 infection in vitro. Certain herbal drug extracts and natural compounds were found to inhibit viral RNA levels and infectious titers of SARS-CoV-2 in a dose-dependent manner. Furthermore, viral protein analyses showed that herbal drug extracts and natural compounds effectively inhibited SARS-CoV-2 in the various entry treatments. Our study revealed that three herbal drugs are good candidates for further in vivo and clinical studies.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Delivery of messenger RNA (mRNA) using lipid nanoparticles (LNPs) is expected to be applied to various diseases following the successful clinical use of the mRNA COVID-19 vaccines. This study aimed ...to evaluate the effect of the cholesterol molar percentage of mRNA-LNPs on protein expression in hepatocellular carcinoma-derived cells and in the liver after intramuscular or subcutaneous administration of mRNA-LNPs in mice. For mRNA-LNPs with cholesterol molar percentages reduced to 10 mol% and 20 mol%, we formulated neutral charge particles with a diameter of approximately 100 nm and polydispersity index (PDI) <0.25. After the intramuscular or subcutaneous administration of mRNA-LNPs with different cholesterol molar percentages in mice, protein expression in the liver decreased as the cholesterol molar percentage in mRNA-LNPs decreased from 40 mol% to 20 mol% and 10 mol%, suggesting that reducing the cholesterol molar percentage in mRNA-LNPs decreases protein expression in the liver. Furthermore, in HepG2 cells, protein expression decreased as cholesterol in mRNA-LNPs was reduced by 40 mol%, 20 mol%, and 10 mol%. These results suggest that the downregulated expression of mRNA-LNPs with low cholesterol content in the liver involves degradation in systemic circulating blood and decreased protein expression after hepatocyte distribution.
In antigen (Ag) cross-presentation, dendritic cells (DCs) take up extracellular Ag and translocate them from the endosome to the cytosol for proteasomal degradation. The processed peptides can enter ...the conventional MHC I pathway. The molecules responsible for the translocation of Ag across the endosomal membrane into the cytosol are unknown. Here we demonstrate that heat shock protein 90 (HSP90) is critical for this step. Cross-presentation and -priming were decreased in both HSP90α-null DCs and mice. CD8α+ DC apoptosis mediated by translocation of exogenous cytochrome c to the cytosol was also eliminated in HSP90α-null mice. Ag translocation into the cytosol was diminished in HSP90α-null DCs and in DCs treated with an HSP90 inhibitor. Internalized Ag was associated with HSP90 and translocated to the cytosol, a process abrogated by the HSP90 inhibitor. Ag within purified phagosomes was released in an HSP90-dependent manner. These results demonstrate the important role of HSP90 in cross-presentation by pulling endosomal Ag out into the cytosol.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Two phenylpropanoid-conjugated iridoids, deglucosyl gaertneroside (
1
) and morindoidin (
2
), were isolated from the leaves of
Morinda morindoides
(Rubiaceae) by activity-guided fractionation using ...an anti-malarial activity assay. The known related iridoids molucidin (
3
) and prismatomerin (
4
), two lignans, abscisic acid, two megastigmanes, and two flavonol glycosides were also identified. The structures of isolated compounds were elucidated using spectroscopic analysis. The isolated compounds were evaluated for anti-malarial activity against the chloroquine/mefloquine-sensitive strains of
Plasmodium falciparum
together with cytotoxicity against adult mouse brain cells. Potent anti-malarial activity of
3
and
4
(IC
50
of 0.96 and 0.80 μM, CC
50
of 1.02 and 0.88 μM, and SI of 1.06 and 1.10, respectively) was shown, while new iridoids
1
and
2
and pinoresinol (
5
) displayed moderate activity (IC
50
of 40.9, 20.6, and 24.2 μM) without cytotoxicity (CC
50
> 50 μM). These results indicate that
1
–
5
may be promising lead compounds for anti-malarial drugs. In addition, our results imply the necessity of the quality control of the extract of
M. morindoides
leaves based on the contents of
1
–
5
in terms of the safety and efficacy.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Three phenylpropanoid-conjugated iridoid glucosides, acetylgaertneric acid (
1
), acetyldehydrogaertneroside (
2
), and dehydrogaertneric acid (
10
), together with nine known related iridoid ...glucosides (
3
–
9
,
11
, and
12
), two coumaroyl alkaloids, one benzenoid, and three flavonoid glucosides were isolated from leaves of
Morinda morindoides
(Rubiaceae). Structures of these isolated compounds were determined using spectroscopic analysis. Compounds
1
–
18
and previously isolated compounds (
19
–
29
) were evaluated for anti-trypanosomal activity against
Trypanosoma cruzi
Tulahuen strain (trypomastigote and amastigote) together with cytotoxicity against host cells, new-born mouse heart cells. Among them, molucidin (
21
) and prismatomerin (
22
) exhibited good anti-trypanosomal activity (IC
50
of 4.67 and 5.70 µM, respectively), together with cytotoxicity (CC
50
of 2.76 and 3.22 μM, respectively). Compounds
1
–
18
did not show anti-malarial activity against a chloroquine/mefloquine-sensitive strain of
Plasmodium falciparum
.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
The outbreak of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a serious threat to global public health. The emergence of SARS-CoV-2 ...variants is a significant concern regarding the continued effectiveness of vaccines and antiviral therapeutics. Thus, natural products such as foods, drinks, and other compounds should be investigated for their potential to treat COVID-19. Here, we examined the
in vitro
antiviral activity against SARS-CoV-2 of various polyethylene terephthalate (PET)-bottled green Japanese teas and tea compounds. Six types of PET-bottled green tea were shown to inhibit SARS-CoV-2 at half-maximal inhibitory concentrations (IC
50
) of 121- to 323-fold dilution. Our study revealed for the first time that a variety of PET-bottled Japanese green tea drinks inhibit SARS-CoV-2 infection in a dilution-dependent manner. The tea compounds epigallocatechin gallate (EGCG) and epicatechin gallate showed virucidal activity against SARS-CoV-2, with IC
50
values of 6.5 and 12.5 µM, respectively. The investigated teas and tea compounds inactivated SARS-CoV-2 in a dose-dependent manner, as demonstrated by the viral RNA levels and infectious titers. Furthermore, the green teas and EGCG showed significant inhibition at the entry and post-entry stages of the viral life cycle and inhibited the activity of the SARS-CoV-2 3CL-protease. These findings indicate that green tea drinks and tea compounds are potentially useful in prophylaxis and COVID-19 treatment.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Lipid nanoparticles (LNPs) are currently in the spotlight as delivery systems for mRNA therapeutics and have been used in the Pfizer/BioNTech and Moderna COVID-19 vaccines. mRNA-LNP formulations have ...been indicated to require strict control, including maintenance at fairly low temperatures during their transport and storage. Since it is a new pharmaceutical modality, there is a lack of information on the systematic investigation of how storage and handling conditions affect the physicochemical properties of mRNA-LNPs and their protein expression ability. In this study, using the mRNA-LNPs with standard composition, we evaluated the effects of temperature, cryoprotectants, vibration, light exposure, and syringe aspiration from the vials on the physicochemical properties of nanoparticles in relation to their in vitro/in vivo protein expression ability. Among these factors, storage at −80 °C without a cryoprotectant caused a decrease in protein expression, which may be attributed to particle aggregation. Exposure to vibration and light also caused similar changes under certain conditions. Exposure to these factors can occur during laboratory and hospital handling. It is essential to have sufficient knowledge of the stability of mRNA-LNPs in terms of their physical properties and protein expression ability at an early stage to ensure reproducible research and development and medical care.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Zika virus (ZIKV) is a re-emerging mosquito-borne flavivirus of African origin that is transmitted by Aedes mosquitoes. ZIKV was historically limited to Africa and Asia, where mild cases were ...reported. However, ZIKV has recently been responsible for major global outbreaks associated with a wide range of neurological complications. Since no antiviral therapy exists for ZIKV, drug discovery research for ZIKV is crucial. Intracellular lipids regulated by sterol regulatory element-binding proteins (SREBPs) are important in flavivirus pathogenesis. PF-429242 has been reported to inhibit the activity of site-1 protease (S1P), which regulates the expression of SREBP target genes. Our primary objective in this study is to elucidate the mechanism of the antiviral activity of PF-429242 against the African genotype (ZIKVMR-766) and Asian genotypes (ZIKV H/PF 2013 and ZIKV PRVABC59) using several primate-derived cell lines. The virus titer was determined via a focus-forming assay; we used flow cytometry to quantify intracellular lipids in ZIKV-infected and mock-treated cells. The PF-429242 molecule effectively suppressed ZIKV infection in neuronal cell lines; T98G, U-87MG, SK-N-SH and primary monocytes cell, indicating that PF-429242 molecule can be used therapeutically. A strong reduction in ZIKV replication was observed at 12 μM and 30 μM in in neuronal cell lines and primary monocytes, respectively. Interestingly, the inhibitory effects of the PF-429242 molecule were observed when it was tested on various ZIKV-lineage infections. Lipid quantification reveals that ZIKV increases lipogenesis in infected cells, while the exogenous addition of cholesterol effectively blocks ZIKV replication. Furthermore, the supplementation of oleic acid increases the ZIKV titer. Fenofibrate, an inhibitor of lipid droplet formation, reduces the ZIKV titer. Collectively, our results demonstrate that the development of antiviral drugs against ZIKV could be based on key regulators of lipid metabolism. In addition, this study reveals that the mechanism of the PF-429242-mediated suppression among flavivirus infections is not entirely identical. Our results warrant further evaluation of PF-429242 as a prospective antiviral drug, given the multiple advantageous properties of this compound, such as its limited toxicity, neuroprotective properties, and broad spectrum of capabilities.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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