[Effect of Spatholobi Caulis extract from ethyl acetate on immune killing function of NK cells].

This study aims to investigate the regulatory effect of the Spatholobi Caulis extract from ethyl acetate(SEA) on natural killer(NK) cells under physiological conditions and elucidate the underlying mechanism. The C57BL/6 mice were randomized into NC and SEA groups, and NK-92 cells were respectively treated with 0, 25, 50, and 100 µg·mL~(-1) SEA. The body weight and immune organ index of the mice were compared between groups. The lactate dehydrogenase(LDH) assay was employed to examine the cytotoxicity of NK-92 cells treated with SEA and the killing activity of mouse NK cells against YAC-1 cells. The cell-counting kit-8(CCK-8) was used to examine the impact of SEA on the proliferation of NK-92 cells. Flow cytometry was employed to measure the number of NK cells in the peripheral blood as well as the expression levels of natural killer group 2 member A(NKG2A) and natural killer group 2 member D(NKG2D). The enzyme-linked immunosorbent assay(ELISA) was performed to determine the interferon(IFN)-γ secretion in the serum. Semi-quantitative PCR was conducted to determine the mRNA levels of NKG2A, NKG2D, and IFN-γ in spleen cells. Western blot was employed to investigate the involvement of phosphoinositide 3-kinase(PI3K)/extracellular regulated protein kinase 1(ERK1) signaling pathway. The results showed that SEA exhibited no adverse effects on the body, while significantly enhance the number of NK cells and augment the cytotoxicity of NK-92 cells against YAC-1 cells. Moreover, it suppressed the expression of NKG2A, enhanced the expression of NKG2D, promoted IFN-γ secretion, and upregulated the protein levels of PI3K and ERK. The findings suggest that SEA has the potential to enhance the immune recognition and effector function of NK cells by increasing the cell number, modulating the expression of functional receptors, and promoting IFN-γ secretion via the PI3K/ERK signaling pathway.

A Double-Blind, Placebo-Controlled, Crossover Study of Magnesium Supplementation in Patients with XMEN Disease.

X-linked MAGT1 deficiency with increased susceptibility to Epstein-Barr virus (EBV) infection and N-linked glycosylation defect (XMEN) disease is an inborn error of immunity caused by loss-of-function mutations in the magnesium transporter 1 (MAGT1) gene. The original studies of XMEN patients focused on impaired magnesium regulation, leading to decreased EBV-cytotoxicity and the loss of surface expression of the activating receptor "natural killer group 2D" (NKG2D) on CD8+ T cells and NK cells. In vitro studies showed that supraphysiological supplementation of magnesium rescued these defects. Observational studies in 2 patients suggested oral magnesium supplementation could decrease EBV viremia. Hence, we performed a randomized, double-blind, placebo-controlled, crossover study in 2 parts. In part 1, patients received either oral magnesium L-threonate (MLT) or placebo for 12 weeks followed by 12 weeks of the other treatment. Part 2 began with 3 days of high-dose intravenous (IV) magnesium sulfate (MgSO4) followed by open-label MLT for 24 weeks. One EBV-infected and 3 EBV-naïve patients completed part 1. One EBV-naïve patient was removed from part 2 of the study due to asymptomatic elevation of liver enzymes during IV MgSO4. No change in EBV or NKG2D status was observed. In vitro magnesium supplementation experiments in cells from 14 XMEN patients failed to significantly rescue NKG2D expression and the clinical trial was stopped. Although small, this study indicates magnesium supplementation is unlikely to be an effective therapeutic option in XMEN disease.

Traditional Thai Massage Promoted Immunity in the Elderly via Attenuation of Senescent CD4+ T Cell Subsets: A Randomized Crossover Study.

The beneficial physiological effects of traditional Thai massage (TTM) have been previously documented. However, its effect on immune status, particularly in the elderly, has not been explored. This study aimed to investigate the effects of multiple rounds of TTM on senescent CD4+ T cell subsets in the elderly. The study recruited 12 volunteers (61-75 years), with senescent CD4+ T cell subsets, who received six weekly 1-h TTM sessions or rest, using a randomized controlled crossover study with a 30-day washout period. Flow cytometry analysis of surface markers and intracellular cytokine staining was performed. TTM could attenuate the senescent CD4+ T cell subsets, especially in CD4+28null NKG2D+ T cells (n = 12; p < 0.001). The participants were allocated into two groups (low < 2.75% or high ≥ 2.75%) depending on the number of CD4+28null NKG2D+ T cells. After receiving TTM over 6 sessions, the cell population of the high group had significantly decreased (p < 0.001), but the low group had no significant changes. In conclusion, multiple rounds of TTM may promote immunity through the attenuation of aberrant CD4+ T subsets. TTM may be provided as a complementary therapy to improve the immune system in elderly populations.

An Update on XMEN Disease.

"X-linked immunodeficiency with magnesium defect, Epstein-Barr virus (EBV) infection, and neoplasia" (XMEN) disease is an inborn error of glycosylation and immunity caused by loss of function mutations in the magnesium transporter 1 (MAGT1) gene. It is a multisystem disease that strongly affects certain immune cells. MAGT1 is now confirmed as a non-catalytic subunit of the oligosaccharyltransferase complex and facilitates Asparagine (N)-linked glycosylation of specific substrates, making XMEN a congenital disorder of glycosylation manifesting as a combined immune deficiency. The clinical disease has variable expressivity, and impaired glycosylation of key MAGT1-dependent glycoproteins in addition to Mg2+ abnormalities can explain some of the immune manifestations. NKG2D, an activating receptor critical for cytotoxic function against EBV, is poorly glycosylated and invariably decreased on CD8+ T cells and natural killer (NK) cells from XMEN patients. It is the best biomarker of the disease. The characterization of EBV-naïve XMEN patients has clarified features of the genetic disease that were previously attributed to EBV infection. Extra-immune manifestations, including hepatic and neurological abnormalities, have recently been reported. EBV-associated lymphomas remain the main cause of severe morbidity. Unfortunately, treatment options to address the underlying mechanism of disease remain limited and Mg2+ supplementation has not proven successful. Here, we review the expanding clinical phenotype and recent advances in glycobiology that have increased our understanding of XMEN disease. We also propose updating XMEN to "X-linked MAGT1 deficiency with increased susceptibility to EBV-infection and N-linked glycosylation defect" in light of these novel findings.

Targeting ADAM10 in Cancer and Autoimmunity.

Generating inhibitors for A Disintegrin And Metalloproteinase 10 (ADAM10), a zinc-dependent protease, was heavily invested in by the pharmaceutical industry starting over 20 years ago. There has been much enthusiasm in basic research for these inhibitors, with a multitude of studies generating significant data, yet the clinical trials have not replicated the same results. ADAM10 is ubiquitously expressed and cleaves many important substrates such as Notch, PD-L1, EGFR/HER ligands, ICOS-L, TACI, and the "stress related molecules" MIC-A, MIC-B and ULBPs. This review goes through the most recent pre-clinical data with inhibitors as well as clinical data supporting the use of ADAM10 inhibitor use in cancer and autoimmunity. It additionally addresses how ADAM10 inhibitor therapy can be improved and if inhibitor therapy can be paired with other drug treatments to maximize effectiveness in various disease states. Finally, it examines the ADAM10 substrates that are important to each disease state and if any of these substrates or ADAM10 itself is a potential biomarker for disease.

Chlorin-Based Photoactivable Galectin-3-Inhibitor Nanoliposome for Enhanced Photodynamic Therapy and NK Cell-Related Immunity in Melanoma.

Photodynamic therapy (PDT) is an encouraging alternative therapy for melanoma treatment and Ce6-mediated PDT has shown some exciting results in clinical trials. However, PDT in melanoma treatment is still hampered by some melanoma's protective mechanisms like antiapoptosis mechanisms and treatment escape pathways. Combined therapy and enhancing immune stimulation were proposed as effective strategies to overcome this resistance. In this paper, a Chlorin-based photoactivable Galectin-3-inhibitor nanoliposome (PGIL) was designed for enhanced Melanoma PDT and immune activation of Natural Killer (NK) cells. PGIL were synthesized by encapsulating the photosensitizer chlorin e6 and low molecular citrus pectin in the nanoliposome to realize NIR-triggered PDT and low molecular citrus pectin (LCP) release into the cytoplasm. The intracellular release of LCP inhibits the activity of galectin-3, which increases the apoptosis, inhibits the invade ability, and enhances the recognition ability of Natural Killer (NK) cells to tumor cells in melanoma cells after PDT. These effects of PGIL were tested in cells and nude mice, and the mechanisms during the in vivo treatment were preliminarily studied. The results showed that PGIL can be an effective prodrug for melanoma therapy.

Long-term survival of a patient with recurrent gallbladder carcinoma, treated with chemotherapy, immunotherapy, and surgery: a case report.

BACKGROUND: Gallbladder cancer (GBC) is one of the refractory diseases. Multidisciplinary approach including immunotherapy for such cancers has received much attention in recent years. CASE PRESENTATION: A 59-year-old man underwent an extended cholecystectomy for GBC (pathological stage II, T2 N0 M0, [per UICC 7th edition]) that was incidentally found during cholelithiasis surgery, and was then treated with adjuvant gemcitabine (GEM). Three months later, when a recurrence-suspected lesion was detected in segment 5 (S5) of his liver, we started adoptive immunotherapies with cytokine-activated killer (CAK) cell infusions, combined with chemotherapy. After a year of adjuvant immunochemotherapy, the S5 lesion disappeared on imaging, but lesions suspected metastatic recurrence again appeared in S7 and S8 at 4 years and 6 months post-surgery, for which GEM and cisplatin (CDDP) were administered as second-line chemotherapy. Immunochemotherapy produced stable disease (per RECIST) for 9 months, when tumor growth was detected; open microwave coagulo-necrotic therapy (MCN) was performed for these lesions. Three years after MCN, a solitary liver metastasis was detected in S4. MCN was conducted again, and peritoneal dissemination was found intraoperatively. A month after the second MCN, the patient's carcinoembryonic antigen (CEA) level had increased. Therefore, GEM and tegafur-gimeracil-oteracil potassium (TS-1) were administered as third-line chemotherapy. We also switched the adoptive immunotherapy for tumor-associated antigen-pulsed dendritic cell-activated killer (DAK) cell immunotherapy. After nine courses of GEM and TS-1 administration, CEA had decreased to a normal level. At the time of reporting, 9 years and 6 months have passed since the initial surgery, and 18 months have passed since the peritoneal metastasis was detected. GEM and CDDP are currently administered as fourth-line chemotherapy because of re-increased CEA. Although an undeniable metastasis was found in his para-aortic lymph node, this patient visits our clinic regularly for immunotherapy. CONCLUSION: We here report a rare case of long-term survival of recurrent GBC well controlled by multidisciplinary therapy. Immunotherapy may be a promising modality among multidisciplinary methods for advanced cancer.

Exploiting natural killer group 2D receptors for CAR T-cell therapy.

Chimeric antigen receptors (CARs) are genetically engineered proteins that combine an extracellular antigen-specific recognition domain with one or several intracellular T-cell signaling domains. When expressed in T cells, these CARs specifically trigger T-cell activation upon antigen recognition. While the clinical proof of principle of CAR T-cell therapy has been established in hematological cancers, CAR T cells are only at the early stages of being explored to tackle solid cancers. This special report discusses the concept of exploiting natural killer cell receptors as an approach that could broaden the specificity of CAR T cells and potentially enhance the efficacy of this therapy against solid tumors. New data demonstrating feasibility of this approach in humans and supporting the ongoing clinical trial are also presented.

Effect of total flavonoids from the seeds of Astragali complanati on natural killer cell function.

ETHNOPHARMACOLOGICAL RELEVANCE: The seeds of Astragalus complanatus R.Br. (AC), a traditional Chinese medicine, have been extensively used for clinical treatment of liver and kidney complaints and tumors for more than a thousand years. It was believed that AC is warm and sweet in the most authoritative medical book of Ancient China "Compendium of Materia Medica". Our previous studies showed that the flavonoid component from the seeds of Astragalus complanatus (FAC) is mainly an active constituent and has the hepatoprotective effect, anti-liver fibrosis, and anti-tumor and immune enhancement. AIM: The aim of this study was to investigate in vitro the regulation effect of FAC on NK cells function and possible mechanism of action. METHODS: The effect of FAC on the proliferation ability of NK-92 cells and the cytolysis of NK-92 cells to K562 and SMMC-7721 were measured by MTT assay and lactase dehydrogenase (LDH)-releasing assay, respectively. The phenotypical characterization (CD3, CD16 and CD56) and activation markers (CD25, CD69 and CD95) of NK-92 cell were detected by flow cytometry analysis. IFN-γ production of NK-92 cells stimulating by K562 cells was quantitated using ELISA. To explore the mechanism of action, mRNA and protein expressions of activating receptors (NKp30, NKp44, NKp46 and NKG2D) in NK-92 cells were detected by quantitative real-time PCR and flow cytometry analysis. RESULTS: Our results demonstrated that FAC significantly promoted the proliferation and the cytotoxicity of NK-92 cells in a dose-dependent manner by enhancing IFN-γ and increasing the expression of the activation markers CD25 and CD69. In addition, FAC had not changed the NK-92 cells phenotypical characterization, but markedly enhanced the expression intensity of CD56. Furthermore, FAC significantly enhanced mRNA and protein expressions lever of NKp44 and NKG2D in NK-92 cells. CONCLUSION: These results suggest that FAC upregulate the expressions of NKG2D, NKp44, which in turn influence NK-92 cells activation. FAC may serve as an immunostimulatory of NK cells for tumor treatment.

Ganoderma lucidum stimulates NK cell cytotoxicity by inducing NKG2D/NCR activation and secretion of perforin and granulysin.

Ganoderma lucidum (G. lucidum) is a medicinal mushroom long used in Asia as a folk remedy to promote health and longevity. Recent studies indicate that G. lucidum activates NK cells, but the molecular mechanism underlying this effect has not been studied so far. To address this question, we prepared a water extract of G. lucidum and examined its effect on NK cells. We observed that G. lucidum treatment increases NK cell cytotoxicity by stimulating secretion of perforin and granulysin. The mechanism of activation involves an increased expression of NKG2D and natural cytotoxicity receptors (NCRs), as well as increased phosphorylation of intracellular MAPKs. Our results indicate that G. lucidum induces NK cell cytotoxicity against various cancer cell lines by activating NKG2D/NCR receptors and MAPK signaling pathways, which together culminate in exocytosis of perforin and granulysin. These observations provide a cellular and molecular mechanism to account for the reported anticancer effects of G. lucidum extracts in humans.