COVID-19: Proposing a Ketone-Based Metabolic Therapy as a Treatment to Blunt the Cytokine Storm

Abstract

Human SARS-CoV-2 infection is characterized by a high mortality rate due to some patients developing a large innate immune response associated with a cytokine storm and acute respiratory distress syndrome (ARDS). This is characterized at the molecular level by decreased energy metabolism, altered redox state, oxidative damage, and cell death. Therapies that increase levels of (R)-beta-hydroxybutyrate (R-BHB), such as the ketogenic diet or consuming exogenous ketones, should restore altered energy metabolism and redox state. R-BHB activates anti-inflammatory GPR109A signaling and inhibits the NLRP3 inflammasome and histone deacetylases, while a ketogenic diet has been shown to protect mice from influenza virus infection through a protective γδ T cell response and by increasing electron transport chain gene expression to restore energy metabolism. During a virus-induced cytokine storm, metabolic flexibility is compromised due to increased levels of reactive oxygen species (ROS) and reactive nitrogen species (RNS) that damage, downregulate, or inactivate many enzymes of central metabolism including the pyruvate dehydrogenase complex (PDC). This leads to an energy and redox crisis that decreases B and T cell proliferation and results in increased cytokine production and cell death. It is hypothesized that a moderately high-fat diet together with exogenous ketone supplementation at the first signs of respiratory distress will increase mitochondrial metabolism by bypassing the block at PDC. R-BHB-mediated restoration of nucleotide coenzyme ratios and redox state should decrease ROS and RNS to blunt the innate immune response and the associated cytokine storm, allowing the proliferation of cells responsible for adaptive immunity. Limitations of the proposed therapy include the following: it is unknown if human immune and lung cell functions are enhanced by ketosis, the risk of ketoacidosis must be assessed prior to initiating treatment, and permissive dietary fat and carbohydrate levels for exogenous ketones to boost immune function are not yet established. The third limitation could be addressed by studies with influenza-infected mice. A clinical study is warranted where COVID-19 patients consume a permissive diet combined with ketone ester to raise blood ketone levels to 1 to 2 mM with measured outcomes of symptom severity, length of infection, and case fatality rate.

Figure 1

Mechanisms that lead to acute respiratory distress syndrome (ARDS) and mortality following SARS-CoV-2 infection are shown. The cells of the innate immune response secrete increasing amounts of cytokines. The cells that normally protect against a cytokine storm lose this ability leading to a runaway positive feedback loop of cytokine production. Abbreviations: 11β-HSD1 and 11β-HSD2: 11β-hydroxysteroid dehydrogenase types 1 and 2; ACE2: angiotensin-converting enzyme 2; AEC I and AEC II: alveolar epithelial cell types I and II; DCs: dendritic cells; FOXO1: forkhead box O1 transcription factor; FOXO3: forkhead box O3 transcription factor; HIF-1α: hypoxia-inducible factor 1 alpha; IFN: interferon; IL-6: interleukin-6; IRF3: IFN-regulatory factor 3; NAD(H): nicotinamide adenine dinucleotide; NADP(H): nicotinamide adenine dinucleotide phosphate; ONOO⁻: peroxynitrite; PGC1-α: PPARG coactivator 1-alpha; PDK1 and PDK4: pyruvate dehydrogenase kinases 1 and 4; RLR: retinoic acid-inducible gene I-like receptors; RNS: reactive nitrogen species; ROS: reactive oxygen species; SOD: superoxide dismutase; TGF-β: transforming growth factor-β; TNF-α: tumor necrosis factor-alpha.

Figure 2

Proposed mechanisms and time course of SARS-CoV-2 infection when using a ketone-based metabolic therapy. Abbreviations: HBP: hexosamine biosynthesis pathway.

Figure 3

The major pathways of central metabolism and reactions that alter the ratios of coenzyme couples are shown. The mitochondrial matrix and the cytoplasm have independent coenzyme couple ratios. ATP synthesized in the mitochondrial matrix is exported to the cytoplasm in exchange for ADP by the adenine nucleotide translocase present in the inner mitochondrial membrane. Acetyl-CoA synthesized in the mitochondrial matrix must also be exported to the cytoplasm to provide two carbon units for fatty acid synthesis and protein acetylation. However, acetyl-CoA cannot cross the mitochondrial inner membrane. Therefore, the transfer of acetyl units across the inner mitochondrial membrane is accomplished using the citrate-pyruvate shuttle or the citrate-malate shuttle. These shuttles alter coenzyme levels and use inner membrane carrier proteins for the transport of citrate, pyruvate, and malate. The net result of the citrate-pyruvate shuttle on coenzyme levels is the use of energy from ATP hydrolysis and NADH oxidation to reduce NADP⁺ to NADPH. There is also a citrate-alpha-ketoglutarate shuttle system that has the net effect of using ATP hydrolysis to increase NADPH in the cytoplasm instead of increasing NADH or NADPH in the mitochondrial matrix. Synthesizing fatty acids in the cytoplasm and reducing antioxidants require NADPH. The malate-aspartate shuttle transfers reducing equivalents from NADH between the cytoplasm and the mitochondrial matrix. The glycerol 3-phosphate shuttle transfers reducing equivalents from cytoplasmic NADH to the mitochondrial ETC. Glucose is catabolized by three pathways including the hexosamine biosynthesis pathway that synthesizes uridine diphosphate (UDP) N-acetylglucosamine, glycolysis that reduces NAD⁺ to NADH and synthesizes ATP from ADP and P_i, and the pentose phosphate pathway (PPP) that reduces NADP⁺ to NADPH and synthesizes ribose sugars for nucleotide synthesis. When the pyruvate dehydrogenase complex (PDC) is inhibited, lactate is synthesized from pyruvate to recycle NAD⁺ from NADH so glycolysis can continue. However, the lactate is exported from the cell and may contribute to lactic acidosis and multiorgan failure [20]. R-BHB decreases the reliance of cells on glycolysis leading to reduced cellular lactate export. Abbreviations: Glut1 and Glut3: glucose transporters 1 and 3; MCT: monocarboxylate transporter; NNT: nicotinamide nucleotide transhydrogenase; PDC: pyruvate dehydrogenase complex.

Figure 4

Phagocyte ROS and RNS metabolism. Most of the major forms of ROS and RNS are derived from superoxide or nitric oxide (NO).

Figure 5

The half-lives and diffusion limits of ROS/RNS.

Figure 6

Restoring PDC activity or metabolizing R-BHB and fatty acids to bypass the inhibited PDC activity that occurs following viral infection is central to beneficial energy reprogramming. Most transcription factor names are taken from the Encyclopedia of Signaling Molecules [97, 98]. Abbreviations: 1,3-BPG: 1,3-bisphosphoglyceric acid; C/EBPβ: CCAAT-enhancer-binding protein β; G3P: glyceraldehyde 3-phosphate; E2F1: E2F transcription factor 1; ERRα and ERRγ: estrogen-related receptor alpha and gamma; GR: glucocorticoid receptor; HNF4α: hepatic nuclear factor 4 alpha; ICER/CREM: inducible cAMP early repressor/cAMP-responsive element modulator; MPC: mitochondrial pyruvate carrier; PDC: pyruvate dehydrogenase complex; PGC1α: PPARG coactivator 1 alpha; PPARα and PPARγ: peroxisome proliferator-activated receptor alpha and gamma; RAR: retinoic acid receptor; RXR: retinoic x receptor; SIRT3: sirtuin 3; STAT5: signal transducer and activator of transcription 5; TR: thyroid hormone receptor.

Figure 7

Maintaining ATP levels during respiratory viral infection is the key to maintain proper ion distribution to avoid edema. The active and passive cotransporters and channels maintain the ion gradients between AEC and extracellular fluids. The apical ligand-gated channels function to maintain appropriate osmotic pressure to provide sufficient fluid in the airway without causing edema. AEC I express superoxide-activated ENaC Na⁺ channels and NOX2 that regulates them. Abbreviations: 3Na⁺/2K⁺-ATPase: 3 sodium 2 potassium ATPase; AE: anion exchange chloride bicarbonate exchanger; BCFTR: basolateral cystic fibrosis transmembrane conductance regulator-like channel; BHBDH: β-hydroxybutyrate dehydrogenase; BIRC: basolateral inward rectifying channel; BK_Ca: large-conductance Ca²⁺- and voltage-gated big K⁺ channel; BORC: basolateral outward rectifying channel; CACC/TMEM: calcium-activated chloride channels/transmembrane protein; CFTR: cystic fibrosis transmembrane conductance regulator; CNG: cyclic nucleotide-gated ion channel; ENaC: epithelial sodium channel; KCa3.1: calcium-activated potassium channel; Kv7.1: voltage-dependent potassium channel; NCX: sodium calcium exchanger; NHE: sodium-hydrogen exchanger; NKCC: sodium potassium chloride cotransporter; NOX2: NADPH oxidase 2; Pit1/2: sodium-dependent phosphate transporters 1 and 2; RYR: ryanodine receptor calcium-induced Ca²⁺ channel; SERCA: sarcoplasmic/endoplasmic reticulum Ca²⁺-ATPase; SK4: SK4 calcium-activated potassium channel.

Figure 8

Proinflammatory signaling that occurs during viral infection and mechanisms through which R-BHB inhibits this signaling. (a) The ACE system. (b) The mechanism by which R-BHB inhibits the NLRP3 inflammasome is unknown. Four possible mechanisms are shown. (c) The hexosamine biosynthesis pathway is required to initiate an effective antiviral innate immune response, but its increased activity can also stimulate a cytokine storm. Abbreviations: 6PGDL: 6-phosphonoglocono-D-lactone; 6PG: 6-phosphogluconate; ACE2: angiotensin-converting enzyme 2; ACE: angiotensin-converting enzyme; ANG (1-9): angiotensin (1-9); ANG (1-7): angiotensin (1-7), a vasodilator; ANG II: angiotensin II, a vasoconstrictor; ANG III: angiotensin III, a metabolite of ANG II; BRCC3: Lys-63-specific deubiquitinase BRCC36; CARD: caspase recruitment domain; CLIC: chloride intracellular channel protein; F1,6BP: fructose 1,6-bisphosphate; F6P: fructose 6-phosphate; FADD: fas-associated protein with death domain; GlcNAc: G,N-acetylglucosamine; G3P: glyceraldehyde 3-phosphate; G6P: glucose 6-phosphate; GlcN-6P: glucosamine-6-phosphate; GlcNAc-6P: N-acetyl glucosamine-6-phosphate; GlcNAc-1p: N-acetyl glucosamine-1-phosphate; HBP: hexosamine biosynthesis pathway; IKK_ε: IκB kinase ε; IL-1R: interleukin-1 receptor; IRAK-1 and IRAK-4: interleukin-1 receptor kinases 1 and 4; IRF3 and IRF5: IFN-regulatory factors 3 and 5; JNK1: c-Jun N-terminal protein kinase 1; K63-Ub: K63-linked polyubiquitin binding; MasR: Mas receptor; MAVS: mitochondrial antiviral signaling protein; MDA5: melanoma differentiation-associated gene 5; MyD88: myeloid differentiation primary response 88; NF-κB: nuclear factor kappa-light-chain-enhancer of activated B cells; NLRP3: NOD-, LRR-, and pyrin domain-containing protein 3; NOD1 and NOD2: nucleotide-binding oligomerization domain-containing proteins 1 and 2; OGT: O-linked N-acetylglucosamine (GlcNAc) transferase; P2X7: P2X purinoceptor 7; PDPK: phosphoinositide-dependent kinase-1; PPP: pentose phosphate pathway; PTMs: posttranslational modifications; R5P: ribose 5-phosphate; RIG-1: retinoic acid-inducible gene I; S7P: sedoheptulose 7-phosphate; SERCA: sarcoplasmic/endoplasmic reticulum Ca²⁺-ATPase; STAT3: signal transducer and activator of transcription 3; TBK-1: TANK-binding kinase 1; TLR: toll-like receptor; TNFR: tumor necrosis factor receptor; TRAF6: tumor necrosis factor receptor- (TNFR-) associated factor 6; TRIF: TIR domain-containing adapter-inducing interferon β; TRX: thioredoxin; TXNIP: thioredoxin-interacting protein; UDP-GlcNAc: uridine diphosphate N-acetylglucosamine.

Figure 9

The effects of increased R-BHB levels on the activity of the transcription factors FOXO3a, FOXO1, HIF1-α, Nrf2, and PGC-1α. (a) DNA wrapped around histones with deacetylated lysines blocks the access of transcription factors, so HDAC function represses transcription. (b) R-BHB, by inhibiting class I HDACs, allows increased histone acetylation, relaxed chromatin, and FOXO3a expression. (c) Chromatin is also relaxed by β-hydroxybutyrylation at promoters such as that of FOXO1 to increase transcription. (d) HIF-1α is activated by RNS. Peroxynitrite S-nitrosylates HIF-1α to inhibit its von Hippel-Landau factor-induced ubiquitination and proteasomal degradation. This stabilizes HIF-1α leading to increased PDK1 expression. This stabilization is likely inhibited by R-BHB, which increases NADPH to decrease ROS/RNS levels. (e) Nrf2 is activated by oxidized DJ-1, and this is regulated by the redox potential of NADP⁺/NADPH that controls the cellular antioxidant potential. The DJ-1 chaperone protein, which is activated by moderate levels of hydrogen peroxide, stimulates the release of Nrf2 from KEAP1 allowing Nrf2 to enter the nucleus and induce antioxidant response element gene expression. (f) PGC-1α is the master regulator of mitochondrial biogenesis. It is a coactivator of ERR-α, FOXO1, FOXO3a, PPARs, and nuclear respiratory factor 1 (NRF1). Abbreviations: Ac: acetate; ARE: antioxidant response element; CA9: carbonic anhydrase 9; CCR7: C-C chemokine receptor type 7; ERR-α: estrogen-related receptor alpha; ETC: electron transport chain; K: lysine; HDAC: histone deacetylase; FIH-1: factor inhibiting HIF-1; FOXO1: forkhead box O1; FOXO3: forkhead box O3; FOXP3: forkhead box P3; G6PC: glucose-6-phosphatase; G6PDH: glucose 6-phosphate dehydrogenase; γGCLM: glutamate cysteine ligase modifier subunit; Glut1: glucose transporter 1; GSRX: glutathione reductase; HIF-1α and HIF-1β: hypoxia-inducible factor 1 alpha and beta; HRE: hypoxia response element; IFNG: interferon gamma gene; IL7R: interleukin-7 receptor; iNOS: inducible nitric oxide synthase; IRF7: interferon regulatory factor 7; KEAP1: Kelch-like ECH-associated protein 1; MAF: musculoaponeurotic fibrosarcoma; Nrf1: nuclear respiratory factor 1; Nrf2: nuclear factor erythroid 2-related factor 2 (NFE2L2); RAG1 and RAG2: recombination activating genes 1 and 2; ONOO⁻: peroxynitrite; ONOOH: peroxynitrous acid; P300/CBP: binding protein 300/CREB-binding protein; PDK1 and PDK4: pyruvate dehydrogenase kinases 1 and 4; PGC1-α: PPARG coactivator 1 alpha; PHD2: prolyl-hydroxylase domain protein 2; pVHL: von Hippel-Landau tumor suppressor protein; SCOT/OXCT1: succinyl-CoA-3-oxaloacid CoA transferase also known as 3-oxoacid CoA-transferase 1; SELL: selectin L; SOD2: superoxide dismutase 2; Ub: ubiquitin; VEGF: vascular endothelial growth factor.