Myricetin inhibits interferon-γ-induced programmed death ligand-1 and indoleamine 2,3-dioxygenase 1 expression in lung cancer cells

OBJECTIVE Programmed death ligand-1(PD-L1)and indoleamine 2,3-dioxygenase 1(IDO1)are immune checkpoints which can be induced by interferon-γ(IFN-γ)in the tumor microenvironment,leading to immune escape of tumors.Myricetin(MY)is a flavonoid distributed in many edible and medicinal plants.The aim of this study is to clarify the effect and the mechanism of MY on inhibiting IFN-γ-induced PD-L1 and IDO1 in lung cancer cells.METHODS Expressions of PD-L1 and major histocompatibility complex-I(MHC-I)were evaluated by flow cytometry and Western blotting,and the expression of IDO1 was measured by Western blotting.qRT-PCR was used to detect their mRNA levels.The function of T cells was evaluated using a co-culture system consist of lung cancer cells and the Jurkat-PD-1 T cell line that overexpressing PD-1.Molecular docking analysis,Western blotting and immunofluorescence were used for mechanism study.RESULTS MY potently inhibited IFN-γ-induced PD-L1 and IDO1 expression in human lung cancer cells,while didn't show obvious effect on the expression of MHC-I.In addition,MY restored the survival,proliferation,CD69 expression and interleukin-2(IL-2)secretion of Jurkat-PD-1 T cells suppressed by IFN-γ-treated lung cancer cells in the co-culture system.Mechanistically,IFN-γup-regulated PD-L1 and IDO1 at the transcriptional level through the JAK-STAT-IRF1 axis,which was targeted and inhibited by MY.CONCLUSION Our research revealed a new insight into the anti-tumor effects of MY which inhibited IFN-γ-induced PD-L1 and IDO1 expression,supporting the potential of MY in anti-tumor immunotherapy.

Indoleamine 2,3-dioxygenase in tumor induced tolerance

Objective To review the recent studies about the role of indoleamine 2,3-dioxygenase （IDO） in tumor induced tolerance. Data sources Published articles （1978-2009） on IDO and tumor induced tolerance were selected from Medline. Study selection Articles selected were relevant to development of IDO in tumor induced tolerance. Of all originally identified articles, 50 specially addressed the stated purpose. Results Recent work has revealed IDO at high levels in tumors and in tumor-draining lymph nodes and a close relationship between IDO activity and the regulatory T cells. Conclusion Up-regulation of IDO is proven to be a mechanism of acquired tolerance in tumors, in which the closely coupled positive feedback system between IDO and reclulatorv T cells may be considered to play an important role.

Combination of carboxyamidotriazole and 1-Methyl-L-tryptophan has synergistic inhibtory effects on programmed death 1 expression

OBJECTIVE To evaluate whether the IDO1 inhibitor 1-methyl-L-tryptophan(1-MT)combine calcium influx inhibitor carboxyamidotriazole(CAI)could further enhance the suppression of programmed death 1(PD-1)in CD8^+T cells and investigate the curative effect of the combined use.METHODS CD8^+T cells were isolated from normal mice spleen by negative selection using magnetic cell separation.The isolated CD8^+T cells were cultured in RPMI 1640 medium containing 10%FBS and 100 U·mL^(-1)IL-2 and activated by the addition of anti-CD3 and anti-CD28(1 g·L^(-1) each mabs).CD8^+T cells were pretreated for 48 h with drug and the fluo-3 as a marker of intracellular calcium concentration was detected by flow cytometry.The calcineurin(Ca N)levels were assayed with ELISA in CD8^+T cells after 48 h incubation with 10μm CAI.The nuclear translocations of NFAT and AHR were detected by immunofluorescent staining after 48 h of drug treatment.The expression of PD-1 in CD8^+T cells was analyzed by flow cytometry.RESULTS Intracellular fluorescent intensity was markedly debase due to CAI treatment(P<0.01).Meanwhile,the changes of CaN content had a resembled correlation(P<0.01).Immunofluorescence experiment showed that after combination therapy the transfer of NFAT and AHR in nuclear substantially reduced.Flow cytometry revealed that after the combination caused a significant decrease in PD-1 expression in CD8^+T cells.CONCLUSION CAI and 1-MT could inhibit markedly the expression of PD-1 in CD8^+T cells by inhibiting the nuclear translocation of NFAT and AHR,respectively and the combination of them has synergetic effect.

1-MT Enhances Potency of Tumor Cell Lysate-pulsed Dendritic Cells against Pancreatic Adenocarcinoma by Downregulating the Percentage of Tregs

This study examined whether 1-methyl-tryptophan [1-MT,an indoleamine 2,3-dioxygenase(IDO) inhibitor] could reduce CD4+CD25+ regulatory T cells(Tregs) proliferation and improve the anti-tumor efficacy of dendritic cells(DCs) pulsed with tumor cell lysate in the mice bearing pancreatic adenocarcinoma.The models of pancreatic adenocarcinoma were established in C57BL/6 mice by subcutaneous injection of Pan02 cells.Eight mice which were subcutaneously injected with PBS served as control.The expression of IDO was determined in tumor draining lymph nodes(TDLNs) and spleens of the murine pancreatic adenocarcinoma models.The prevalence of Tregs was measured in the TDLNs and spleens before and after 1-MT administration.The dendritic cells were pulsed with tumor cell lysate for preparing DC vaccine.The DC vaccine,as a single agent or in combination with 1-MT,was administered to pancreatic adenocarcinoma mice.The anti-tumor efficacy was determined after different treatments by regular observation of tumor size.The results showed that the levels of IDO mRNA and protein in tumor-bearing mice were significantly higher than those in the normal control mice.The percentage of Tregs in the spleen and TDLNs was also higer in tumor-bearing mice than in normal control mice(P<0.05).Foxp3 expression was significantly lower in the TDLNs and spleens of tumor-bearing mice administrated with 1-MT than that in normal control mice.Furthemore,in the mice that were administered 1-MT plus DC vaccine,the tumor was increased more slowly than in mice treated with DC vaccine or 1-MT alone,or PBS on day 36(P<0.01).Our results indicated that 1-MT may enhance anti-tumor efficacy of dendritic cells pulsed with tumor cell lysate by downregulating the percentage of Tregs.

Interactions of Indoleamine 2,3-dioxygenase-expressing LAMP3^(+) dendritic cells with CD4^(+) regulatory T cells and CD8^(+) exhausted T cells:synergistically remodeling of the immunosuppressive microenvironment in cervical cancer and therapeutic implications

Background Cervical cancer(CC)is the fourth most common cancer in women worldwide.Although immunotherapy has been applied in clinical practice,its therapeutic efficacy remains far from satisfactory,necessitating further investigation of the mechanism of CC immune remodeling and exploration of novel treatment targets.This study aimed to investigate the mechanism of CC immune remodeling and explore potential therapeutic targets.Methods We conducted single-cell RNA sequencing on a total of 17 clinical specimens,including normal cervical tissues,high-grade squamous intraepithelial lesions,and CC tissues.To validate our findings,we conducted multicolor immunohistochemical staining of CC tissues and constructed a subcutaneous tumorigenesis model in C57BL/6 mice using murine CC cell lines(TC1)to evaluate the effectiveness of combination therapy involving indoleamine 2,3-dioxygenase 1(IDO1)inhibition and immune checkpoint blockade(ICB).We used the unpaired two-tailed Student's t-test,Mann-Whitney test,or Kruskal-Wallis test to compare continuous data between two groups and one-way ANOVA with Tukey's post hoc test to compare data between multiple groups.Results Malignant cervical epithelial cells did not manifest noticeable signs of tumor escape,whereas lysosomal-associated membrane protein 3-positive(LAMP3^(+))dendritic cells(DCs)in a mature state with immunoregulatory roles were found to express IDO1 and affect tryptophan metabolism.These cells interacted with both tumor-reactive exhausted CD8^(+)T cells and CD4^(+) regulatory T cells,synergistically forming a vicious immunosuppressive cycle and mediating CC immune escape.Further validation through multicolor immunohistochemical staining showed co-localization of neoantigen-reactive T cells(CD3^(+),CD4^(+)/CD8^(+),and PD-1^(+))and LAMP3+DCs(CD80^(+) and PD-L1^(+)).Additionally,a combination of the IDO1 inhibitor with an ICB agent significantly reduced tumor volume in the mouse model of CC compared with an ICB agent alone.Conclusions Our study suggested that a combination treatment consisting of targeting IDO1 and ICB agent could improve the therapeutic efficacy of current CC immunotherapies.Additionally,our results provided crucial insights for designing drugs and conducting future clinical trials for CC.

Immunogenic-cell-killing and immunosuppression-inhibiting nanomedicine

Combining chemo-therapeutics with immune checkpoint inhibitors facilitates killing cancer cells and activating the immune system through inhibiting immune escape.However,their treatment effects remain limited due to the compromised accumulation of both drugs and inhibitors in certain tumor tissues.Herein,a new poly(acrylamide-co-acrylonitrile-co-vinylimidazole-co-bis(2-methacryloyl)oxyethyl disulfide)(PAAVB)polymer-based intelligent platform with controllable upper critical solution temperature(UCST)was used for the simultaneous delivery of paclitaxel(PTX)and curcumin(CUR).Additionally,a hyaluronic acid(HA)layer was coated on the surface of PAAVB NPs to target the CD44-overexpressed tumor cells.The proposed nanomedicine demonstrated a gratifying accumulation in tumor tissue and uptake by cancer cells.Then,the acidic microenvironment and high level of glutathione(GSH)in cancer cells could spontaneously decrease the UCST of polymer,leading to the disassembly of the NPs and rapid drug release at body temperature without extra-stimuli.Significantly,the released PTX and CUR could induce the immunogenic cell death(ICD)to promote adaptive anti-tumor immunogenicity and inhibit immunosuppression through suppressing the activity of indoleamine 2,3-dioxygenase 1(IDO1)enzyme respectively.Therefore,the synergism of this intelligent nanomedicine can suppress primary breast tumor growth and inhibit their lung metastasis.