The folate cycle enzyme MTHFD2 induces cancer immune evasion through PD-L1 up-regulation.

Metabolic enzymes and metabolites display non-metabolic functions in immune cell signalling that modulate immune attack ability. However, whether and how a tumour's metabolic remodelling contributes to its immune resistance remain to be clarified. Here we perform a functional screen of metabolic genes that rescue tumour cells from effector T cell cytotoxicity, and identify the embryo- and tumour-specific folate cycle enzyme methylenetetrahydrofolate dehydrogenase 2 (MTHFD2). Mechanistically, MTHFD2 promotes basal and IFN-γ-stimulated PD-L1 expression, which is necessary for tumourigenesis in vivo. Moreover, IFN-γ stimulates MTHFD2 through the AKT-mTORC1 pathway. Meanwhile, MTHFD2 drives the folate cycle to sustain sufficient uridine-related metabolites including UDP-GlcNAc, which promotes the global O-GlcNAcylation of proteins including cMYC, resulting in increased cMYC stability and PD-L1 transcription. Consistently, the O-GlcNAcylation level positively correlates with MTHFD2 and PD-L1 in pancreatic cancer patients. These findings uncover a non-metabolic role for MTHFD2 in cell signalling and cancer biology.

Fluorescent AgInS/ZnS quantum dots microplate and lateral flow immunoassays for folic acid determination in juice samples.

A noninstrumental rapid test for folic acid (FA) detection with visual results evaluation utilizing bright water-stable AgInS/ZnS (AIS/ZnS) quantum dots (QDs) is reported . AIS/ZnS QDs are hydrophilic photostable nanocrystals with size < 7 nm and emission in the visible range. They were synthesized directly in the water phase by a simple method compared to the synthesis of other QDs and conjugated with monoclonal antibodies specific for FA via ligand carboxyl groups. The conjugate was used for the development of instrumental qualitative and rapid quantitative FA detection methods. The competitive fluorescent microplate immunosorbent assay provided a limit of detection of 0.1 ng/mL FA and half maximal inhibitory concentration (IC50) of 24 ng/mL FA. The analytical signal was measured at ʎex = 410 nm and ʎem=590 nm. The proposed method showed no cross-reaction with other group B vitamins. For FA screening in juice samples, the lateral flow immunoassay was developed with a visual cutoff level of 3 µg/mL. In our perception, the developed methods are convenient for proving the perception of the AIS/ZnS QDs application as a luminescent label for immunoassay and are effective for FA detection. Graphical abstract.

Immunodeficiency and inborn disorders of vitamin B12 and folate metabolism.

PURPOSE OF REVIEW: Immune dysfunction, including severe combined immunodeficiency, has been described in genetic disorders affecting the metabolism of the vitamins cobalamin (vitamin B12) and folate. We have reviewed reports of clinical findings in patients with a number of inborn errors of cobalamin or folate metabolism, specifically looking for immune problems. RECENT FINDINGS: There is little evidence that immune function is affected in most of the disorders. Exceptions are Imerslund-Gräsbeck syndrome and hereditary folate malabsorption (affecting intestinal absorption of cobalamin and folate, respectively), transcobalamin deficiency (affecting transport of cobalamin in blood and cellular cobalamin uptake), and methylenetetrahydrofolate dehydrogenase 1 deficiency (catalyzing cytoplasmic interconversion of reduced folate coenzyme derivatives). SUMMARY: Although some inborn errors of cobalamin or folate can be associated with immune dysfunction, the degree and type of immune dysfunction vary with no obvious pattern.

Evaluation of regulatory T cells frequency and FoxP3/GDF-15 gene expression in ß-thalassemia major patients with and without alloantibody; correlation with serum ferritin and folate levels.

ß-thalassemia major is one of the most common hematologic disorders in the world. It causes severe anemia and patients require regular blood transfusions, which causes different complications such as iron overload and alloimmunization. Regulatory T cells (Tregs) have an important role in regulation of immune responses. FoxP3 is the major marker of Tregs and its expression can be influenced by different factors. GDF-15 is another gene that plays a role in iron homeostasis and regulation of immune system in different diseases. The aim of this study was to assess the frequency of Tregs and FoxP3/GDF-15 gene expression in ß-thalassemia major patients with and without alloantibody as well as its correlation with different factors such as serum ferritin and folate levels. This study was conducted on 68 ß-thalassemia major patients with and without alloantibodies in comparison with 20 healthy individuals with matched age and sex as control group. Enzyme-linked immunosorbent assay (ELISA), flow cytometry, and real-time PCR were performed in order to evaluate serum ferritin and folate levels, frequency of Tregs, and the expression of FoxP3 and GDF-15 genes, respectively. The percentage and absolute count of Tregs were increased in patients compared with controls (P = 0.0003), but there was no difference between responders and non-responders (P > 0.05). The Tregs count correlated positively with serum ferritin. No correlation was observed between target genes and serum ferritin and folate, but there was a positive significant correlation between the expression of FoxP3 and GDF-15 genes, which shows the immunosuppressive role of GDF-15.

Folate-Modified Chitosan Nanoparticles Coated Interferon-Inducible Protein-10 Gene Enhance Cytotoxic T Lymphocytes' Responses to Hepatocellular Carcinoma.

Adoptive therapy using tumor antigen-specific cytotoxic T lymphocytes (CTLs) is a promising approach for treatment of human cancers. Due to immune suppression in cancer patients, it is difficult for tumor antigen-specific CTLs to arrive at tumor tissues. Interferon-inducible protein-10 (IP-10) is a powerful chemokine that effectively attracts CTLs to tumor tissues and improves their anti-tumor activity. Increase over expression of IP-10 in tumor tissues can efficiently promote efficacy of adoptive therapy. Folate-modified chitosan nanoparticles coating the human IP-10 gene (FA-CS-hIP-10) were therefore developed in this study. The FA-CS-hIP-10 nanoparticles were specifically bound to folate receptors on hepatoma cells and promoted the expression of IP-10, to improve the activity of pMAGE-A1(278-286) specific CTLs. Combination of the FA-CS-hIP-10 and pMAGE-A1(278-286) specific CD8+ CTLs efficiently increased secretion of IFN-γ, inhibited tumor growth and extended survival of nude mice with subcutaneously transplanted human hepatocellular carcinoma. Our results demonstrated that the mechanism behind this novel therapeutic approach involved inhibition of angiogenesis and proliferation, and also promoted apoptosis of tumor cells. Our study provides a potentially novel approach for treatment of human hepatocellular carcinoma by improving the activity of tumor antigen-specific CTLs.

Levels of folate receptor autoantibodies in maternal and cord blood and risk of neural tube defects in a Chinese population.

BACKGROUND: After years of periconceptional folic acid supplementation, the prevalence of neural tube defects (NTDs) remains stable following the remarkable reduction observed immediately after the fortification practice. There is accumulating evidence that folate receptor (FR) autoimmunity may play a role in the etiology of folate-sensitive NTDs. METHODS: From 2011 to 2013, 118 NTD cases and 242 healthy controls were recruited from a population-based birth defects surveillance system in Northern China. Enzyme-linked immunosorbent assay was used to measure FR autoantibodies in maternal and cord blood. Logistic regression models were used to estimate the odds ratios (OR) and 95% confidence intervals (95% CI). RESULTS: Plasma FR autoantibodies levels were significantly elevated in mothers of infants with NTDs compared with mothers of healthy controls. Using the lowest tertile as the referent group, 2.20-fold (95% CI, 0.71-6.80) and 5.53-fold increased odds (95% CI, 1.90-16.08) of NTDs were observed for the second and third tertile of immunoglobulin G (IgG), respectively, and the odds of NTDs for each successive tertile of IgM was 0.98 (95% CI, 0.35-2.75) and 3.49 (95% CI, 1.45-8.39), respectively. A dose-response relationship was found between FR autoantibodies levels and risk of NTDs (P < 0.001 for IgG, P = 0.002 for IgM). The same pattern was observed in both subtypes of spina bifida and anencephaly. No significant difference in levels of cord blood FR autoantibodies was observed. CONCLUSION: Higher levels of FR autoimmunity in maternal plasma are associated with elevated risk of NTDs in a dose-response manner. Birth Defects Research (Part A) 106:685-695, 2016. © 2016 Wiley Periodicals, Inc.

Optimization of Folate-Targeted Immunotherapy for the Treatment of Experimental Arthritis.

Folate-targeted immunotherapy constitutes a powerful method for the treatment of established arthritis in multiple animal models of the disease. The therapy involves immunization of the animal against a hapten to induce anti-hapten antibodies, followed by injection with a folate-hapten conjugate to decorate the surface of folate receptor-positive (activated) macrophages with the antigenic hapten. The hapten-marked macrophages are then recognized by the anti-hapten antibodies and eliminated by immune mechanisms, leading to attenuation of disease symptoms. In the following paper, we optimize the therapy for elimination of inflammatory macrophages and suppression of rheumatoid arthritis symptoms. We also demonstrate a tight correlation between folate receptor-positive macrophage abundance in the liver and inflammation of affected joints. The results suggest that therapies that reduce folate receptor-positive macrophage populations in the body should constitute effective treatments for rheumatoid arthritis.

Autoantibodies against homocysteinylated protein in a mouse model of folate deficiency-induced neural tube defects.

BACKGROUND: Periconceptional supplementation with folic acid results in a significant reduction in the incidence of neural tube defects (NTDs). Nonetheless, NTDs remain a leading cause of perinatal morbidity and mortality worldwide, and the mechanism(s) by which folate exerts its protective effects are unknown. Homocysteine is an amino acid that accumulates under conditions of folate-deficiency, and is suggested as a risk factor for NTDs. One proposed mechanism of homocysteine toxicity is its accumulation into proteins in a process termed homocysteinylation. METHODS & RESULTS: Herein, we used a folate-deficient diet in pregnant mice to demonstrate that there is: (i) a significant inverse correlation between maternal serum folate levels and serum homocysteine; (ii) a significant positive correlation between serum homocysteine levels and titers of autoantibodies against homocysteinylated protein; and (iii) a significant increase in congenital malformations and NTDs in mice deficient in serum folate. Furthermore, in mice administered the folate-deplete diet before conception, supplementation with folic acid during the gestational period completely rescued the embryos from congenital defects, and resulted in homocysteinylated protein titers at term that are comparable to that of mice administered a folate-replete diet throughout both the pre- and postconception period. These results demonstrate that a low-folate diet that induces NTDs also increases protein homocysteinylation and the subsequent generation of autoantibodies against homocysteinylated proteins. CONCLUSION: These data support the hypotheses that homocysteinylation results in neo-self antigen formation under conditions of maternal folate deficiency, and that this process is reversible with folic acid supplementation.

SERS encoded nanoparticle heterodimers for the ultrasensitive detection of folic acid.

In this paper, gold­silver nanoparticle (AuNP­AgNP) heterodimers were assembled with highly yield as an active SERS substrate, based on antigen­antibody immunoreaction. The developed SERS sensor has successful achieved the ultrasensitive detection of folic acid (FA) with the limit of detection (LOD) as 0.86 pg/mL. And the linear range was from 0.005 ng/mL to 1 ng/mL. The results also demonstrated that this developed method showed high specificity and excellent recovery for the human serum samples, indicating its promising potential in bio-diagnosis and the environmental monitoring.

Allergy workup for suspected folic acid hypersensitivity

No disponible

A Phase I/Ib study of folate immune (EC90 vaccine administered with GPI-0100 adjuvant followed by EC17) with interferon-α and interleukin-2 in patients with renal cell carcinoma.

Folate immune (EC90 vaccine with GPI-0100 adjuvant followed by EC17) is a novel folate-targeted hapten immunotherapy designed to exploit the overexpression of folate receptors on renal cell carcinoma (RCC) cells. In this open-label, phase I/II clinical study, we report the safety, pharmacokinetics, and antitumor activity of folate immune with concurrent interleukin-2 (IL-2) and interferon-α (IFN-α) in patients with recurrent or metastatic RCC. Twenty-four patients were enrolled. Following 2 phase I cohorts of 6 patients each, we extended the study to 12 additional patients: 18 received weekly vaccination of 1.2 mg of EC90 with 3.0 mg of GPI-0100 adjuvant for 4 weeks. Beginning on cycle 1, day 8, 0.3 mg/kg of EC17 was administered once daily, 5 days per week (Monday-Friday) for 4 consecutive weeks. Beginning on cycle 1, day 15, IL-2 and IFN-α were administered at doses of 12 and 3.0 MIU, respectively, after the EC17 dose, 3 times per week (Monday, Wednesday, and Friday) for 3 weeks. In cycle 2, IL-2 and IFN-α, doses of 7.0 and 3.0 MIU, respectively, were administered 3 days per week (Monday, Wednesday, and Friday) for 4 consecutive weeks. No dose-limiting toxicities were observed. Most adverse events reported were grade 1 or 2, with only twelve grade ≥3 toxicities reported. Sixteen patients had progressive disease, 7 patients were observed to have stable disease, and 1 patient achieved a partial response lasting 71 days. Overall, folate immune plus low-dose IFN-α and IL-2 was safe and well tolerated with some observed clinical activity.

Biomarkers of one-carbon metabolism are associated with biomarkers of inflammation in women.

Folate-mediated one-carbon metabolism is essential for DNA synthesis, repair, and methylation. Perturbations in one-carbon metabolism have been implicated in increased risk of some cancers and may also affect inflammatory processes. We investigated these interrelated pathways to understand their relation. The objective was to explore associations between inflammation and biomarkers of nutritional status and one-carbon metabolism. In a cross-sectional study in 1976 women selected from the Women's Health Initiative Observational Study, plasma vitamin B-6 [pyridoxal-5'-phosphate (PLP)], plasma vitamin B-12, plasma folate, and RBC folate were measured as nutritional biomarkers; serum C-reactive protein (CRP) and serum amyloid A (SAA) were measured as biomarkers of inflammation; and homocysteine and cysteine were measured as integrated biomarkers of one-carbon metabolism. Student's t, chi-square, and Spearman rank correlations, along with multiple linear regressions, were used to explore relations between biomarkers; additionally, we tested stratification by folic acid fortification period and multivitamin use. With the use of univariate analysis, plasma PLP was the only nutritional biomarker that was modestly significantly correlated with serum CRP and SAA (ρ = -0.22 and -0.12, respectively; P < 0.0001). Homocysteine (µmol/L) showed significant inverse correlations with all nutritional biomarkers (ranging from ρ = -0.30 to ρ = -0.46; all P < 0.0001). With the use of multiple linear regression, plasma PLP, RBC folate, homocysteine, and cysteine were identified as independent predictors of CRP; and PLP, vitamin B-12, RBC folate, and homocysteine were identified as predictors of SAA. When stratified by folic acid fortification period, nutrition-homocysteine correlations were generally weaker in the postfortification period, whereas associations between plasma PLP and serum CRP increased. Biomarkers of inflammation are associated with PLP, RBC folate, and homocysteine in women. The connection between the pathways needs to be further investigated and causality established. The trial is registered at clinicaltrials.gov as NCT00000611.

Natural killer cell cytotoxicity is not regulated by folic acid in vitro.

OBJECTIVES: Folate supplementation may be associated with an increased risk of developing several types of cancer and a derangement of immune function. Among the latter, Natural killer (NK) cells are involved in non-MHC-restricted natural immunity against malignant target cells. Abnormalities in NK cell number or function have been associated with a higher cancer risk. The aim of this study was to study in vitro the possible effect of different concentrations of 5-methyltetrahydrofolic acid (5-MTHF) or folic acid on NK cell cytotoxic function, and expression of the stimulatory and inhibitory receptors KIRDL4, KIRDL3, and NKG2D. METHODS: Volunteer-derived peripheral mononuclear cells (PBMC) and highly enriched NK cells (95% CD56+ CD16+) were grown in folic acid free-RPMI 1640, supplemented either with folic acid or 5-MTHF (15-100 nM) during 72 h to 96 h. RESULTS: No differences in the cytolytic activity of PBMC and enriched NK cells were observed. After 96 h of in vitro culture without folate or supplemented with FA or 5-MTHF (30 or 100 nM), there were no changes in the percentage of HPNK receptor-positive cells. CONCLUSIONS: Our data indicate that a high dose of 5-MTHF or folic acid does not influence NK cell function in vitro.

Francisella tularensis live vaccine strain folate metabolism and pseudouridine synthase gene mutants modulate macrophage caspase-1 activation.

Francisella tularensis is a Gram-negative bacterium and the causative agent of the disease tularemia. Escape of F. tularensis from the phagosome into the cytosol of the macrophage triggers the activation of the AIM2 inflammasome through a mechanism that is not well understood. Activation of the AIM2 inflammasome results in autocatalytic cleavage of caspase-1, resulting in the processing and secretion of interleukin-1ß (IL-1ß) and IL-18, which play a crucial role in innate immune responses to F. tularensis. We have identified the 5-formyltetrahydrofolate cycloligase gene (FTL_0724) as being important for F. tularensis live vaccine strain (LVS) virulence. Infection of mice in vivo with a F. tularensis LVS FTL_0724 mutant resulted in diminished mortality compared to infection of mice with wild-type LVS. The FTL_0724 mutant also induced increased inflammasome-dependent IL-1ß and IL-18 secretion and cytotoxicity in macrophages in vitro. In contrast, infection of macrophages with a F. tularensis LVS rluD pseudouridine synthase (FTL_0699) mutant resulted in diminished IL-1ß and IL-18 secretion from macrophages in vitro compared to infection of macrophages with wild-type LVS. In addition, the FTL_0699 mutant was not attenuated in vivo. These findings further illustrate that F. tularensis LVS possesses numerous genes that influence its ability to activate the inflammasome, which is a key host strategy to control infection with this pathogen in vivo.

MR1 presents microbial vitamin B metabolites to MAIT cells.

Antigen-presenting molecules, encoded by the major histocompatibility complex (MHC) and CD1 family, bind peptide- and lipid-based antigens, respectively, for recognition by T cells. Mucosal-associated invariant T (MAIT) cells are an abundant population of innate-like T cells in humans that are activated by an antigen(s) bound to the MHC class I-like molecule MR1. Although the identity of MR1-restricted antigen(s) is unknown, it is present in numerous bacteria and yeast. Here we show that the structure and chemistry within the antigen-binding cleft of MR1 is distinct from the MHC and CD1 families. MR1 is ideally suited to bind ligands originating from vitamin metabolites. The structure of MR1 in complex with 6-formyl pterin, a folic acid (vitamin B9) metabolite, shows the pterin ring sequestered within MR1. Furthermore, we characterize related MR1-restricted vitamin derivatives, originating from the bacterial riboflavin (vitamin B2) biosynthetic pathway, which specifically and potently activate MAIT cells. Accordingly, we show that metabolites of vitamin B represent a class of antigen that are presented by MR1 for MAIT-cell immunosurveillance. As many vitamin biosynthetic pathways are unique to bacteria and yeast, our data suggest that MAIT cells use these metabolites to detect microbial infection.

Frontiers in vitamin research: new antibodies, new data.

Since 2004, the anatomical distribution of vitamins in the monkey brain, studied using immunohistochemical techniques and new tools (specific antisera that discriminate different vitamins reasonably well), has been an ongoing research field. The visualization of immunoreactive structures containing vitamins (folic acid, riboflavin, thiamine, pyridoxal, and vitamin C) has recently been reported in the monkey brain (Macaca fascicularis), all these vitamins showing a restricted or very restricted distribution. Folic acid, thiamine, and riboflavin have only been observed in immunoreactive fibers, vitamin C has only been found in cell bodies (located in the primary somatosensory cortex), and pyridoxal has been found in both fibers and cell bodies. Perikarya containing pyridoxal have been observed in the paraventricular hypothalamic nucleus, the periventricular hypothalamic region, and in the supraoptic nucleus. The fibers containing vitamins are thick, smooth (without varicosities), and are of medium length or long, whereas immunoreactive cell bodies containing vitamins are round or triangular. At present, there are insufficient data to elucidate the roles played by vitamins in the brain, but the anatomical distribution of these compounds in the monkey brain provides a general idea (although imprecise and requiring much more study) about the possible functional implications of these molecules. In this sense, here the possible functional roles played by vitamins are discussed.