Genetic predisposition similarities between NASH and ASH: Identification of new therapeutic targets

Abstract

Fatty liver disease can be triggered by a combination of excess alcohol, dysmetabolism and other environmental cues, which can lead to steatohepatitis and can evolve to acute/chronic liver failure and hepatocellular carcinoma, especially in the presence of shared inherited determinants. The recent identification of the genetic causes of steatohepatitis is revealing new avenues for more effective risk stratification. Discovery of the mechanisms underpinning the detrimental effect of causal mutations has led to some breakthroughs in the comprehension of the pathophysiology of steatohepatitis. Thanks to this approach, hepatocellular fat accumulation, altered lipid droplet remodelling and lipotoxicity have now taken centre stage, while the role of adiposity and gut-liver axis alterations have been independently validated. This process could ignite a virtuous research cycle that, starting from human genomics, through omics approaches, molecular genetics and disease models, may lead to the development of new therapeutics targeted to patients at higher risk. Herein, we also review how this knowledge has been applied to: a) the study of the main PNPLA3 I148M risk variant, up to the stage of the first in-human therapeutic trials; b) highlight a role of MBOAT7 downregulation and lysophosphatidyl-inositol in steatohepatitis; c) identify IL-32 as a candidate mediator linking lipotoxicity to inflammation and liver disease. Although this precision medicine drug discovery pipeline is mainly being applied to non-alcoholic steatohepatitis, there is hope that successful products could be repurposed to treat alcohol-related liver disease as well.

Keywords: AA, arachidonic acid; ASH, alcoholic steatohepatitis; DAG, diacylglycerol; DNL, de novo lipogenesis; ER, endoplasmic reticulum; FFAs, free fatty acids; FGF19, fibroblast growth factor 19; FLD, fatty liver disease; FXR, farnesoid X receptor; GCKR, glucokinase regulator; GPR55, G protein-coupled receptor 55; HCC, hepatocellular carcinoma; HFE, homeostatic iron regulator; HSC, hepatic stellate cells; HSD17B13, hydroxysteroid 17-beta dehydrogenase 13; IL-, interleukin-; IL32; LDs, lipid droplets; LPI, lysophosphatidyl-inositol; MARC1, mitochondrial amidoxime reducing component 1; MBOAT7; MBOAT7, membrane bound O-acyltransferase domain-containing 7; NASH, non-alcoholic steatohepatitis; PNPLA3; PNPLA3, patatin like phospholipase domain containing 3; PPAR, peroxisome proliferator-activated receptor; PRS, polygenic risk score; PUFAs, polyunsaturated fatty acids; SREBP, sterol response element binding protein; TAG, triacylglycerol; TNF-α, tumour necrosis factor-α; alcoholic liver disease; cirrhosis; fatty liver disease; genetics; interleukin-32; non-alcoholic fatty liver disease; precision medicine; steatohepatitis; therapy.

Fig. 1

Ethanol metabolism and related mechanisms promoting hepatic lipids accumulation. During alcohol consumption, ethanol is oxidised to acetaldehyde (by the constitutive pathway involving the NAD-dependent ADH) or metabolised at the level of the microsomal system through an inducible NADPH-dependent pathway involving cytochrome P450 (MEOS). Both lead to the formation of acetaldehyde, which is subsequently metabolised to acetic acid by the 2-ADH. ADH, alcohol dehydrogenase; 2-ADH, 2-alcohol dehydrogenase; AMPK, AMP-activated kinase; FFAs, free fatty acids; PPAR-α, peroxisome proliferator-activated receptor a; SREBP-1c, sterol regulatory element-binding protein 1c.

Fig. 2

Alcohol-related and non-alcoholic steatohepatitis share major genetic determinants. Among the main genetic determinants which predispose to the development of progressive FLD, it is possible to distinguish a central core of genes that include modulators of fatty acid metabolism, lipid storage and secretion. These genes highlight the common genetic background, shared between AFLD and NAFLD. Conversely, other genes are specifically associated with only 1 type of liver injury such as GCKR and APOB in NAFLD/MAFLD. AFLD, alcoholic fatty liver disease; APOB, apolipoprotein B; FLD, fatty liver disease; GCKR, glucokinase regulator; HSD17B13, hydroxysteroid 17-beta dehydrogenase 13; MAFLD, metabolic dysfunction-associated fatty liver disease; MARC1, mitochondrial amidoxime reducing component 1; MBOAT7, membrane bound O-acyltransferase domain containing 7; NAFLD, non-alcoholic fatty liver disease; PCSK7, proprotein convertase subtilisin/kexin Type 7; PNPLA3, patatin like phospholipase domain containing 3; TM6SF2, transmembrane 6 superfamily member 2.

Fig. 3

The new discovery paradigm: From human to molecular genetics and into the clinic. FLD risk stratification results from a cyclic interplay between clinical studies in at risk individuals and the manipulation of the associated pathways to improve liver damage. The direct target discovery approach – based on human genome level data – aims to identify high-impact FLD variants. This strategy may lead to a progressive improvement in risk stratification, coupling the information carried by the characterisation of risk variants with information derived from several bioinformatic “omics” approaches. The following characterisation of the disease mechanisms in experimental models leads to novel therapeutics. Among all the efforts devoted to the pursuit of a personalised medicine approach, this represents an optimal strategy for the modulation of disease pathways during pre-clinical studies and then in clinical trials. The example of PNPLA3 I148M variant is illustrated in the outer circle of the figure (in green). The modulation of genes involved in lipid droplet remodelling and lipotoxicity employing ASOs, as in the case of PNPLA3 risk variant, may be a successful blueprint. ASO, anti-sense oligonucleotides; FLD, fatty liver disease; HSD17B13, hydroxysteroid 17-beta dehydrogenase 13; PNPLA3, patatin like phospholipase domain containing 3; PRS, polygenic risk score.

Fig. 4

MBOAT7 downregulation and LPI accumulation leads to steatohepatitis. MBOAT7 localises to the endoplasmic reticulum, where it catalyses the transfer of AA to LPI resulting in AA-PI, that is incorporated into cell membranes. In carriers of the rs641738 risk variant or during hyperinsulinemia, MBOAT7 is downregulated, resulting in a reduction of AA-PI and accumulation of LPI that is converted into TAG by the alternative synthesis pathway through the intermediate DAG, a process promoted by SREBP1c. Deficiency of AA-PI upregulates CDS, enhancing LPI synthesis. In HSCs, LPI binds to GPR55 triggering inflammatory and profibrotic gene expression and ECM deposition. AA, arachidonic acid; AA-PI, AA-containing PI; CDS, CDP-diacylglycerol-synthase; DAG, diacylglycerol; ECM, extracellular matrix; HSCs, hepatic stellate cells; LPI, lysophosphatidyl-inositol; MBOAT7, membrane bound O-acyltransferase domain containing 7; PI, phosphatidylinositol; SREBP-1c, sterol regulatory element-binding protein 1c; TAG, triacylglycerol.

Fig. 5

Potential mechanisms linking IL-32 with liver damage in fatty liver disease and steatohepatitis. IL-32 levels are elevated in obese and diabetic patients, suggesting a link between this cytokine and low-grade chronic inflammation and insulin resistance. More importantly, IL-32 is highly induced during liver damage, and its levels correlate with disease severity, progression to NASH and carriage of the PNPLA3 I148M variant. Acting as a master regulator of other pro-inflammatory cytokines, IL-32 promotes inflammation and liver damage by increasing the secretion of IL-1β, IL-6 and IL-10, and may also drive hepatic fibrogenesis by promoting the transcription of ECM-remodelling genes. Moreover, the IL-32 promoter is targeted by fatty acid-responsive transcription factors and thus steatosis promotes non-canonical IL-32β secretion in the liver. Both IL-32 and carriage of PNPLA3 I148M upregulate STAT3, leading to lipotoxicity and ultimately liver damage. ECM, extracellular matrix; IL-interleukin-; NASH, non-alcoholic steatohepatitis; PNPLA3, patatin like phospholipase domain containing 3.

Fig. 6

Human genetics supports the involvement of the gut-liver axis in NASH. NASH is associated to microbiota dysbiosis, and the over representation of pathogenic bacterial species leads to impairment in the gut-vascular barrier and bile acid management. Reduction in secondary bile acids results in hampered FXR signalling, facilitating intestinal inflammation and further impairment of the gut-blood barrier, which can be rescued by FXR agonists. FXR activation results in the release of FGF19, an enterokine that regulates hepatic bile acid metabolism by interacting with FGFR4 receptor and KLB co-receptor. Reduction of FXR in steatohepatitis negatively affects bile acid metabolism, and therefore bile acids accumulate in the liver promoting inflammation and carcinogenesis. Notably, rs35724 falling in the NR1H4 locus is protective against liver damage, but increases cardiovascular risk by increasing cholesterol synthesis; on the other hand, a KLB variant reducing protein levels promotes liver damage in obese patients. Pathways altered during NASH are indicated by red arrows, with potential therapeutic approaches in green. BAs, bile acids; CHOL, cholesterol; FGF, fibroblast growth factor; FGFR, FGF receptor; FMT, faecal microbiota transplant; FXR, farnesoid X receptor; HCC, hepatocellular carcinoma; KLB, klotho beta; NASH, non-alcoholic steatohepatitis; SCFAs, short-chain fatty acids.