AMPK Alchemy: Therapeutic Potentials in Allergy, Aging, and Cancer.

All cells are equipped with intricate signaling networks to meet the energy demands and respond to the nutrient availability in the body. AMP-activated protein kinase (AMPK) is among the most potent regulators of cellular energy balance. Under ATP -deprived conditions, AMPK phosphorylates substrates and affects various biological processes, such as lipid/glucose metabolism and protein synthesis. These actions further affect the cell growth, death, and functions, altering the cellular outcomes in energy-restricted environments. AMPK plays vital roles in maintaining good health. AMPK dysfunction is observed in various chronic diseases, making it a promising target for preventing and alleviating such diseases. Herein, we highlight the different AMPK functions, especially in allergy, aging, and cancer, to facilitate the development of new therapeutic approaches in the future.

Methionine consumption by cancer cells drives a progressive upregulation of PD-1 expression in CD4 T cells.

Programmed cell death protein 1 (PD-1), expressed on tumor-infiltrating T cells, is a T cell exhaustion marker. The mechanisms underlying PD-1 upregulation in CD4 T cells remain unknown. Here we develop nutrient-deprived media and a conditional knockout female mouse model to study the mechanism underlying PD-1 upregulation. Reduced methionine increases PD-1 expression on CD4 T cells. The genetic ablation of SLC43A2 in cancer cells restores methionine metabolism in CD4 T cells, increasing the intracellular levels of S-adenosylmethionine and yielding H3K79me2. Reduced H3K79me2 due to methionine deprivation downregulates AMPK, upregulates PD-1 expression and impairs antitumor immunity in CD4 T cells. Methionine supplementation restores H3K79 methylation and AMPK expression, lowering PD-1 levels. AMPK-deficient CD4 T cells exhibit increased endoplasmic reticulum stress and Xbp1s transcript levels. Our results demonstrate that AMPK is a methionine-dependent regulator of the epigenetic control of PD-1 expression in CD4 T cells, a metabolic checkpoint for CD4 T cell exhaustion.

T-cell engaging poly(lactic-co-glycolic acid) nanoparticles as a modular platform to induce a potent cytotoxic immunogenic response against PD-L1 overexpressing cancer.

Bispecific nanoparticles (NPs) are conjugated with two antibodies that enhance T cell cytotoxicity by sequentially targeting CD3 and tumor-specific proteins. This interaction redirects T cells to specific tumor antigens and activates them to lyse tumor cells by blocking two different signaling pathways simultaneously. This study developed NP-based bispecific T-cell engagers (nanoBiTEs), which are R848-loaded bispecific poly(lactic-co-glycolic acid) NPs decorated with anti-CD3 antibody targeting T cells and anti-PD-L1 antibody targeting PD-L1 ligands (bis-R848-PLGA-NPs). Bis-R848-PLGA-NPs enhance the immunogenic response in destroying cancer cells by restoring the T cell effector functions. These interactions allow T cells to come in close proximity to the tumor cells. Finally, the release of R848 from PLGA-NPs activates dendritic cells, enhancing T cell activation. In vitro results show maximum internalization of bis-R848-PLGA-NPs in SK-OV3 and B16F10 cell lines, attributed to high PD-L1 expression in both cells. Furthermore, bis-R848-PLGA-NPs-treated CD8+ T cells exhibit a significantly increased total amount of CD8+/CD25+, CD8+/CD69+, and cytokine expression that leads to the robust inhibition of PD-L1 expressed cancer cells. Additionally, tumor growth is significantly inhibited by bis-R848-PLGA-NPs in the B16F10 xenograft mouse model and significantly enhanced intratumoral infiltration of CD4+ and CD8+ T cells, as well as tumor-infiltrated cytokines.

AMPK Amplifies IL2-STAT5 Signaling to Maintain Stability of Regulatory T Cells in Aged Mice.

AMP-activated protein kinase (AMPK), an important regulator of the aging process, is expressed in various immune cells. However, its role in regulatory T cell (Treg) stability during aging is poorly understood. Here, we addressed the role of AMPK in Treg function and stability during aging by generating Treg-specific AMPKα1 knockout mice. In this study, we found that AMPKα1-deficient Tregs failed to control inflammation as effectively as normal Tregs did during aging. AMPK knockout from Tregs reduces STAT5 phosphorylation in response to interleukin (IL)-2 stimulation, thereby destabilizing Tregs by decreasing CD25 expression. Thus, our study addressed the role of AMPK in Tregs in sensing IL-2 signaling to amplify STAT5 phosphorylation, which, in turn, supports Treg stability by maintaining CD25 expression and controlling inflamm-aging.

AMPK promotes antitumor immunity by downregulating PD-1 in regulatory T cells via the HMGCR/p38 signaling pathway.

BACKGROUND: AMP-activated protein kinase (AMPK) is a metabolic sensor that maintains energy homeostasis. AMPK functions as a tumor suppressor in different cancers; however, its role in regulating antitumor immunity, particularly the function of regulatory T cells (Tregs), is poorly defined. METHODS: AMPKα1fl/flFoxp3YFP-Cre, Foxp3YFP-Cre, Rag1-/-, and C57BL/6 J mice were used for our research. Flow cytometry and cell sorting, western blotting, immuno-precipitation, immuno-fluorescence, glycolysis assay, and qRT-PCR were used to investigate the role of AMPK in suppressing programmed cell death 1 (PD-1) expression and for mechanistic investigation. RESULTS: The deletion of the AMPKα1 subunit in Tregs accelerates tumor growth by increasing the expression of PD-1. Metabolically, loss of AMPK in Tregs promotes glycolysis and the expression of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), a key enzyme of the mevalonate pathway. Mechanistically, AMPK activates the p38 mitogen-activated protein kinase (MAPK) that phosphorylates glycogen synthase kinase-3ß (GSK-3ß), inhibiting the expression of PD-1 in Tregs. CONCLUSION: Our study identified an AMPK regulatory mechanism of PD-1 expression via the HMGCR/p38 MAPK/GSK3ß signaling pathway. We propose that the AMPK activator can display synergic antitumor effect in murine tumor models, supporting their potential clinical use when combined with anti-PD-1 antibody, anti-CTLA-4 antibody, or a HMGCR inhibitor.

Co-existence of beta-lactamases in clinical isolates of Escherichia coli from Kathmandu, Nepal.

BACKGROUND: The trend of extended-spectrum beta-lactamases producing Escherichia coli (ESBL-EC) is increasing in Nepal. Limited studies have been reported investigating ESBL types and carbapenemases in E. coli. METHODS: A cross sectional study was conducted between June 2012 to January 2013 in Kathmandu Medical College and Teaching Hospital, Nepal. Non-repetitive clinical samples from out-patient department (OPD) and Intensive Care Units (ICU) were processed for bacteriological culture and identification of E. coli. Antibiotic susceptibility test, screening and phenotypic confirmation for ESBLs and carbapenemases and PCR (blaCTX-M, blaSHV and blaTEM-type ESBLs, blaVIM, blaIMP and blaNDM-1-type carbapenemases, and class 1 integron element integrase gene) were performed. Clones were resolved by PCR-Randomly Amplified Polymorphic DNA. RESULTS: Out of 332 non-repetitive clinical specimens processed for culture and identification 160 (48.2%) were culture positive. Of which, 93 (58.1%) were E. coli. Of these, 24 (25.8%) were phenotypically confirmed as ESBL-EC and 3 (12.50%) of 24 ESBL-EC were carbapenemase producers. blaCTX-M-type ESBL was most common (23, 95.8%) followed by blaTEM (7, 29.2%) and blaSHV (3, 12.5%). blaVIM, blaIMP and blaNDM-1 were present in 3, 2 and 2 ESBL-EC, respectively. Class 1 integron element was present in 18 (75.0%) ESBL-EC. Nine isolates possessed more than one type of beta-lactamases. Interestingly, all carbapenemase producers were isolated form ICU and co-existence of blaCTX-M, blaSHV, blaTEM, blaIMP, blaVIM and blaNDM-1 beta-lactamases was documented in one ESBL-EC (EC104). All most all isolates had different RAPD patterns. CONCLUSIONS: For the first time in Nepal, high prevalence of blaCTX-M-type ESBL and co-existence of ESBLs and carbapenemases has been described. Continuous monitoring and surveillance and proper infection control and prevention practices will limit the further spread of these super-bugs within this hospital and beyond.