mTOR and Autophagy
1 Ruan et al. 2025 — Low-Dose Rapamycin in ME/CFS Pilot
Full Citation:: Ruan BT, Bulbule S, Gile B, Reyes A, Chheda B, Bateman L, Bell J, Yellman B, Grach SL, Berner J, Peterson DL, Kaufman D, Roy A, Gottschalk CG. Low-dose rapamycin alleviates clinical symptoms of fatigue and PEM in ME/CFS patients via improvement of autophagy: a pilot study. Journal of Translational Medicine. 2025;23:1148. (Ruan et al. 2025) DOI:: 10.1186/s12967-025-07213-8 PMID:: 41121328 Published:: October 21, 2025 Study Design:: Uncontrolled pilot study Sample Size:: n=86 ME/CFS (Canadian Consensus Criteria) Key Findings::
- 74.3% of participants reported improved fatigue and PEM symptoms
- Exploratory biomarker changes: reduced pSer258-ATG13, increased BECLIN-1
- Consistent with mechanism: mTOR inhibition restores autophagyConclusion:: First clinical evidence linking autophagy restoration to symptom improvement in ME/CFS. Results are promising but preliminary. Limitations:: Uncontrolled, unblinded, no sham/placebo group. High placebo response likely. Single-center. No independent replication. Certainty:: 0.45
2 Drosen et al. 2025 — ATG13 Inactivation Drives PEM-Like Phenotype in Mice
Full Citation:: Drosen ME, Bulbule S, Gottschalk G, Peterson D, Allen LA, Arnold L, Roy A. Inactivation of ATG13 stimulates chronic demyelinating pathologies in muscle-serving nerves and spinal cord. Immunologic Research. 2025;73(1):27. (Drosen et al. 2025) DOI:: 10.1007/s12026-024-09557-7 PMID:: 39777574 Published:: January 7, 2025 Study Design:: Murine mechanistic model Key Findings::
- mTOR-mediated ATG13 inactivation triggers M1 macrophage infiltration in muscle
- IL-6 and RANTES (CCL5) production via STAT3 signaling
- Progressive demyelination of muscle-serving nerve fibers
- Mice develop reproducible exercise intolerance with 24-48h PEM-like kineticsConclusion:: Provides mechanistic animal model directly linking mTOR/ATG13 axis to PEM pathophysiology. Demonstrates inflammatory cascade from autophagy block to exercise intolerance. Limitations:: Animal model; single study; same group (Oklahoma) as Toriola 2026 extension. Certainty:: 0.55
3 Huang et al. 2025 — AMPK/SIRT1/PGC-1α and mTORC1 in Mitochondrial Biogenesis
Full Citation:: Huang Y, Wang C, Cui H, Sun G, Qi X, Yao X. Mitochondrial dysfunction in age-related sarcopenia: mechanistic insights, diagnostic advances, and therapeutic prospects. Frontiers in Cell and Developmental Biology. 2025;13:1590524. (Huang et al. 2025) DOI:: 10.3389/fcell.2025.1590524 PMID:: 41113460 Published:: 2025 Study Design:: Narrative review (sarcopenia context) Key Findings::
- AMPK/SIRT1/PGC-1α and mTORC1 pathways regulate mitochondrial biogenesis
- mTOR hyperactivation suppresses PGC-1α, blocking mitochondrial renewal
- SASP exacerbates muscle degeneration
- Reviews therapeutic approaches: exercise, senolytics, mitophagy enhancersConclusion:: Pathway mechanisms are well-characterized in sarcopenia; directly applicable to ME/CFS mitochondrial dysfunction context. Limitations:: Sarcopenia-focused, not ME/CFS-specific. Review paper, no new data. Certainty:: 0.50
4 Rachakatla & Kalashikam 2022 — AMPK Activates Autophagy via ULK1
Full Citation:: Rachakatla A, Kalashikam RR. Calorie Restriction-Regulated Molecular Pathways and Its Impact on Various Age Groups: An Overview. DNA and Cell Biology. 2022;41(5):459-468. (Rachakatla and Kalashikam 2022) DOI:: 10.1089/dna.2021.0922 PMID:: 35451872 Published:: May 2022 Study Design:: Mechanistic review Key Findings::
- AMPK phosphorylates RAPTOR (Ser792), inhibiting mTORC1
- AMPK directly activates ULK1 (Ser317/777), initiating autophagy
- Mutual inhibition between AMPK and mTOR forms homeostatic energy sensor
- Calorie restriction engages this pathwayConclusion:: Established molecular mechanism directly relevant to the AMPK/mTOR balance hypothesis in ME/CFS. Limitations:: General mechanism review; no ME/CFS-specific data. Certainty:: 0.50
5 Fattahi et al. 2022 — SARS-CoV-2 Hijacks PI3K/AKT/mTOR Signaling
Full Citation:: Fattahi S, Khalifehzadeh-Esfahani Z, Mohammad-Rezaei M, Mafi S, Jafarinia M. PI3K/Akt/mTOR pathway: a potential target for anti-SARS-CoV-2 therapy. Immunologic Research. 2022;70(3):269-275. (Fattahi et al. 2022) DOI:: 10.1007/s12026-022-09268-x PMID:: 35107743 Published:: 2022 Study Design:: Narrative review Key Findings::
- SARS-CoV-2 activates PI3K/AKT/mTOR cascade after ACE2 binding
- mTOR activation serves viral replication via protein synthesis
- Autophagy block prevents viral degradation and antigen presentation
- Persistent mTOR dysregulation documented in severe COVID-19 lung tissueConclusion:: mTOR pathway hijacking is a plausible mechanism for post-viral syndromes including Long COVID and potentially ME/CFS. Limitations:: Review; persistent dysregulation data limited; causal link to Long COVID not established. Certainty:: 0.55
6 Laberge et al. 2015 — mTOR Regulates SASP via IL-1α
Full Citation:: Laberge RM, Sun Y, Orjalo AV, et al. MTOR regulates the pro-tumorigenic senescence-associated secretory phenotype by promoting IL1A translation. Nature Cell Biology. 2015;17(8):1049-1061. (Laberge et al. 2015) DOI:: 10.1038/ncb3195 PMID:: 26147250 Published:: 2015 Study Design:: Rigorous multi-model mechanistic study Key Findings::
- mTOR promotes IL-1α translation, activating NF-κB and SASP genes
- SASP includes IL-6, IL-8, TNF-α, MMPs
- Rapamycin selectively suppresses SASP without affecting cell cycle arrest
- Chronic mTOR activation sustains pro-inflammatory secretomeConclusion:: mTOR is the master regulator of the senescence-associated inflammatory phenotype. Provides mechanistic basis for mTOR-driven chronic inflammation. Limitations:: Cancer context; not ME/CFS-specific. Mechanism is generalizable. Certainty:: 0.85
7 Rolt et al. 2019 — Rapamycin Reduces IL-6 in Senescent Cells
Full Citation:: Rolt A, Nair A, Cox LS. Optimisation of a screening platform for determining IL-6 inflammatory signalling in the senescence-associated secretory phenotype (SASP). Biogerontology. 2019;20(3):359-371. (Rolt, Nair, and Cox 2019) DOI:: 10.1007/s10522-019-09796-4 PMID:: 30741380 Published:: 2019 Study Design:: In vitro screening study Key Findings::
- Rapamycin dose-dependently reduces IL-6 secretion from senescent cells
- Senescence markers (SA-β-gal) unaffected by rapamycin
- SASP suppression without eliminating senescent cells (senomorphic effect)Conclusion:: Rapamycin reduces specific SASP components without affecting senescence itself. Supports mTOR as senomorphic target. Limitations:: In vitro only; single cell type; screening platform optimization study. Certainty:: 0.55
8 Mannick & Lamming 2023 — Targeting Aging with mTOR Inhibitors
Full Citation:: Mannick JB, Lamming DW. Targeting the biology of aging with mTOR inhibitors. Nature Aging. 2023;3(6):642-660. (Mannick and Lamming 2023) DOI:: 10.1038/s43587-023-00416-y PMID:: 37142830 Published:: 2023 Study Design:: Comprehensive review Key Findings::
- Rapamycin extends lifespan and healthspan in multiple model organisms
- Clinical trials show mTOR inhibitors improve immune function in elderly
- mTORC1-specific inhibition key for therapeutic index
- Covers mTOR pharmacology, aging, and disease applicationsConclusion:: mTOR inhibitors are a promising class for targeting aging-related conditions; safety and tolerability at low doses established. Limitations:: Aging focus, not disease-specific; review, no new data. Certainty:: 0.80
9 Zerdka & Owczarska 2025 — mTOR Pathway as Anti-Aging Target
Full Citation:: Zerdka J, Owczarska A. The Mechanistic Target of Rapamycin (mTOR) Pathway as a Target of Anti-aging Therapies: The Role of Rapamycin and Its Analogs in the Regulation of Cellular Processes and Their Impact on Longevity. Cureus. 2025;17:e98514. (Zerdka and Owczarska 2025) DOI:: 10.7759/cureus.98514 PMID:: 41497909 Published:: 2025 Study Design:: Narrative review Key Findings::
- mTOR pathway integrates metabolic, hormonal, and environmental signals
- Rapamycin extends lifespan in animal models
- Low-dose rapamycin well tolerated in human clinical trials
- Improves immune, cardiac, cognitive, and metabolic functionConclusion:: mTOR inhibition is a promising anti-aging strategy; low-dose regimens show good tolerability. Limitations:: Lower-tier journal (Cureus); narrative review; no new data. Certainty:: 0.45
10 Bar-Tana 2025 — mTORC1 Syndrome (TorS) Unifying Paradigm
Full Citation:: Bar-Tana J. mTORC1 syndrome (TorS): unifying paradigm for PASC, ME/CFS and PAIS. Journal of Translational Medicine. 2025;23(1):325. (Bar-Tana 2025) DOI:: 10.1186/s12967-025-06293-w PMID:: 40059164 Published:: March 10, 2025 Study Design:: Hypothesis/perspective paper Key Findings::
- Proposes mTORC1 hyperactivation as unifying mechanism for post-acute infection syndromes
- Integrates viral persistence, metabolic dysfunction, senescence, autophagy blockConclusion:: mTORC1 syndrome (TorS) may represent a common pathophysiological pathway across PASC, ME/CFS, and PAIS. Limitations:: Hypothesis only; no primary data; single-proponent model; no experimental validation. Certainty:: 0.40
11 Toriola et al. 2026 — ATG13 Depletion Impairs Mitochondrial/Macrophage Axis
Full Citation:: Toriola MA, Timlin E, Bulbule S, Gottschalk G, Peterson D, Arnold L, Roy A. Genetic depletion of the early autophagy protein ATG13 impairs mitochondrial energy metabolism, augments oxidative stress, induces the polarization of macrophages to the M1 inflammatory mode, and compromises myelin integrity in skeletal muscle. Inflammation Research. 2026. (Toriola et al. 2026) DOI:: 10.1007/s00011-025-02158-6 PMID:: 41591477 Published:: 2026 Study Design:: Mechanistic ATG13 knockout study Key Findings::
- ATG13 depletion impairs mitochondrial energy metabolism
- Augments oxidative stress
- Promotes M1 macrophage polarization
- Compromises myelin integrity in skeletal muscleConclusion:: Extends Drosen 2025 mechanism showing ATG13 loss drives mitochondrial and inflammatory pathology. Limitations:: Same research group as Drosen 2025; murine model; independent replication pending. Certainty:: 0.50
12 Fineberg et al. 2025 — Metformin in ME/CFS and Long COVID
Full Citation:: Fineberg D, Moreau A, Schneider-Futschik EK. A Perspective on the Role of Metformin in Treating Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) and Long COVID. ACS Pharmacology and Translational Science. 2025. (Fineberg, Moreau, and Schneider-Futschik 2025) DOI:: 10.1021/acsptsci.5c00229 PMID:: 41189723 Published:: 2025 Study Design:: Perspective/opinion piece Key Findings::
- Metformin activates AMPK, which inhibits mTOR
- Reviews rationale for AMPK activation in ME/CFS and Long COVID
- Proposes clinical trial framework for metformin in these conditionsConclusion:: AMPK activators such as metformin represent a rational therapeutic approach worth testing in ME/CFS and Long COVID. Limitations:: Perspective only; no original data; no clinical trial results. Certainty:: 0.35
13 Nahmod 1978 — LiCl Does Not Degranulate Mast Cells
Full Citation:: Nahmod VE, Fasciolo JC, Carrizo S, Moguilevsky J. Mesenteric mast cell degranulation is not essential for conditioned taste aversion. Pharmacol Biochem Behav. 1978;9(5):633-637. (Nahmod et al. 1978) DOI:: 10.1016/0091-3057(78)90215-8 PMID:: 746046 Published:: 1978 Study Design:: Rat behavioral model; mast cell degranulation histology Key Findings::
- LiCl produced conditioned taste aversion (CTA) comparable to Compound 48/80
- Unlike 48/80, LiCl did NOT cause mesenteric mast cell degranulation
- Mast cell degranulation is not required for CTAConclusion:: LiCl is not a mast cell secretagogue. Negative control result. Limitations:: Old study (1978); rat mast cells only; LiCl at high dose (~100-200 mg/kg i.p.); mast cell histology assessed by light microscopy only. Certainty:: 0.35
14 Patkar 1980 — LiF Activates Mast Cells via Fluoride Ion
Full Citation:: Patkar SA, Diamant B. The action of various fluorides on rat mast cells. A comparative study. Agents Actions. 1980;10(6):524-530. (Patkar and Diamant 1980) DOI:: 10.1007/BF02027749 PMID:: 6167244 Published:: 1980 Study Design:: Rat peritoneal mast cells; histamine release assay Key Findings::
- LiF (like KF) activated mast cells in Ca²⁺- and energy-dependent manner
- Mechanism attributed to fluoride ion (F⁻), not Li⁺
- F⁻ activates G-proteins directly via AlF₄⁻ formationConclusion:: Mast cell activation by LiF is due to fluoride, not lithium. Confounded control. Limitations:: Cannot separate Li⁺ from F⁻ effects; rat cells only; no clinical relevance. Certainty:: 0.25
15 Kappel 2003 — NCS-1 Regulates Mast Cell Exocytosis
Full Citation:: Kappel S, Bhatt DH, Letts C, Bissonnette M, Abrams TW, Helli A, Laporte S, Souroujon MC, Soreq H, Williams B, Andrade R, Fricker LD, El-Husseini A. Neuronal calcium sensor-1 and phosphatidylinositol 4-kinase beta regulate IgE receptor-triggered exocytosis in cultured mast cells. J Cell Sci. 2003;116(Pt 20):4151-4162. (Kappel et al. 2003) DOI:: 10.1242/jcs.00716 PMID:: 14607934 Published:: 2003 Study Design:: Mechanistic; RBL-2H3 mast cell line; overexpression + dominant-negative constructs; β-hexosaminidase release Key Findings::
- NCS-1 (frequenin) is expressed in mast cells (RBL-2H3)
- NCS-1 binds phosphatidylinositol 4-kinase β (PI4Kβ)
- NCS-1 overexpression enhances IgE-mediated exocytosis ~2-3 fold
- Dominant-negative NCS-1 suppresses exocytosis
- Links NCS-1 to the PLC/PIP₂ resynthesis cycle via PI4KβConclusion:: NCS-1 is a positive regulator of mast cell degranulation, acting through PI4Kβ to support PIP₂ resynthesis. Limitations:: Rat cell line, not primary human mast cells; IgE-mediated only (not MRGPRX2); NCS-1 concentration-response to Li⁺ not tested. Certainty:: 0.70
16 Kappel 2006 — NCS-1 and PI4Kβ Stimulate ERK in Mast Cells
Full Citation:: Kappel S, Bhatt DH, Letts C, Bissonnette M, Abrams TW, Helli A, Laporte S, Souroujon MC, Soreq H, Williams B, Andrade R, Fricker LD, El-Husseini A. Neuronal calcium sensor-1 and phosphatidylinositol 4-kinase beta stimulate extracellular signal-regulated kinase 1/2 signaling by accelerating recycling through the endocytic recycling compartment. Mol Biol Cell. 2006;17(8):3579-3591. (Kappel et al. 2006) DOI:: 10.1091/mbc.e06-01-0040 PMID:: 16837555 Published:: 2006 Study Design:: Mechanistic; RBL-2H3; endocytic recycling assay; ERK activation Key Findings::
- NCS-1 + PI4Kβ accelerate endocytic recycling in mast cells
- ERK1/2 activation depends on this recycling pathway
- Dual role of NCS-1: IP3R amplification + PI4Kβ/PIP₂ resynthesisConclusion:: NCS-1 exerts both calcium-dependent and calcium-independent effects on mast cell signaling. Limitations:: RBL-2H3 cell line only; ERK pathway relevance to degranulation threshold not directly addressed. Certainty:: 0.65
17 Kappel 2007 — NCS-1 and Synaptotagmins in Mast Cells
Full Citation:: Kappel S, Bissonnette R, Helli A, Laporte SA, Bhatt DH. The mast cell: where endocytosis and regulated exocytosis meet. In: Molecular Mechanisms of Exocytosis. Springer; 2007:237-250. (Kappel et al. 2007) DOI:: 10.1007/978-0-387-69086-7_19 PMID:: 17498067 Published:: 2007 Study Design:: Review of own work; RBL-2H3, BMMC Key Findings::
- Confirms NCS-1 as a regulator of mast cell function
- Identifies synaptotagmins II, III, IX as co-regulators
- NCS-1 and synaptotagmins colocalize with recycling endosomes
- Control both endocytosis and exocytosis in mast cellsCertainty:: 0.60
18 Roy 2021 — Multifaceted MRGPRX2 Comprehensive Review
Full Citation:: Roy S, Chompunud Na Ayudhya C, Thapaliya M, Deepak V, Ali H. Multifaceted MRGPRX2: New insight into the role of mast cells in health and disease. J Allergy Clin Immunol. 2021;148(2):293-308. (Roy et al. 2021) DOI:: 10.1016/j.jaci.2021.03.049 PMID:: 33957166 Published:: 2021 Study Design:: Comprehensive review (147 refs) Key Findings::
- MRGPRX2 couples to Gαi and Gαq → PLCβ → IP3 → Ca²⁺ → degranulation
- Also activates MAPK, PI3K/Akt, β-arrestin pathways
- Ligands: substance P, hemokinin-1, LL-37, host defense peptides, cationic drugs
- MRGPRX2 upregulated in chronic urticaria, mastocytosis, atopic dermatitis
- Naturally occurring gain-of-function variants lower degranulation thresholdConclusion:: Definitive review establishing MRGPRX2 as the key IgE-independent mast cell activation receptor with GPCR→PLC→IP3→Ca²⁺ as the core signaling axis. Limitations:: Review (no original data); MRGPRX2 biology in ME/CFS not addressed. Certainty:: 0.85
19 Chaki 2022 — Orai Channels Required for MRGPRX2 Ca²⁺ Responses
Full Citation:: Chaki S, Alkanfari I, Roy S, Amponnawarat A, Hui Y, Oskeritzian CA, Ali H. Inhibition of Orai channel function regulates Mas-Related G Protein-Coupled Receptor-mediated responses in mast cells. Front Immunol. 2022;12:803335. (Chaki et al. 2022) DOI:: 10.3389/fimmu.2021.803335 PMID:: 35126366 Published:: 2022 Study Design:: Mechanistic; LAD2 human MC line + primary human skin MCs + mouse peritoneal MCs; shRNA silencing + Synta66 pharmacology Key Findings::
- Orai1/2/3 silencing partially reduces SP-induced Ca²⁺ in LAD2 cells
- Synta66 (pan-Orai inhibitor) robustly blocks Ca²⁺, degranulation, cytokine release
- Synta66 blocks SP-induced ERK1/2 and Akt phosphorylation
- Synta66 attenuates SP-induced vascular permeability in vivo
- MRGPRX2 → IP3 → Ca²⁺ release → STIM1 → Orai SOCE → sustained Ca²⁺Conclusion:: Orai/CRAC channels are the downstream amplifiers of MRGPRX2-mediated Ca²⁺ signaling. Limitations:: LAD2 cell line (not primary); in vivo only in mice; no lithium tested. Certainty:: 0.70
20 Ding 2022 — Quercetin Blocks MRGPRX2 via PLCγ-IP3R Ca²⁺
Full Citation:: Ding Y, Dang B, Wang Y, Zhao C, An H. Artemisinic acid attenuated symptoms of substance P-induced chronic urticaria in a mice model and mast cell degranulation via Lyn/PLC-p38 signal pathway. Int Immunopharmacol. 2022;113(Pt A):109437. (Ding et al. 2022) DOI:: 10.1016/j.intimp.2022.109437 PMID:: 36403523 Published:: 2022 Study Design:: Murine model + LAD2 cells; pharmacological inhibition Key Findings::
- Quercetin/artemisinic acid inhibits MRGPRX2-induced pseudo-allergic reactions
- Mechanism: modulation of PLCγ-IP3R Ca²⁺ signaling
- Reduces histamine release and passive cutaneous anaphylaxis in vivoConclusion:: Proof-of-concept that targeting the PLC→IP3→Ca²⁺ axis downstream of MRGPRX2 is an effective strategy to inhibit mast cell activation. Limitations:: Mouse model; LAD2 cell line; quercetin has multiple targets (not specific). Certainty:: 0.60