Iminosugar Glucosidase Inhibitors Reduce Hepatic Inflammation in Hepatitis A Virus-Infected Ifnar1 ^−/− Mice

Abstract

Iminosugar compounds are monosaccharide mimetics with broad but generally weak antiviral activities related to inhibition of enzymes involved in glycobiology. Miglustat (N-butyl-1-deoxynojirimycin), which is approved for treatment of lipid storage diseases in humans, and UV-4 (N-(9-methoxynonyl)-1-deoxynojirimycin), inhibit replication of hepatitis A virus (HAV) in cell culture (IC50 32.13 μM and 8.05 μM, respectively) by blocking the synthesis of gangliosides essential for HAV cell entry. We used a murine model of hepatitis A and targeted mass spectrometry to assess the capacity of these compounds to deplete hepatic gangliosides and modify the course of HAV infection in vivo. Miglustat, given by gavage to Ifnar1-/- mice (4800 mg/kg/day) depleted hepatic gangliosides by 69-75%, but caused substantial gastrointestinal toxicity and failed to prevent viral infection. UV-4, similarly administered in high doses (400 mg/kg/day), was well tolerated, but depleted hepatic gangliosides by only 20% after 14 days. UV-4 depletion of gangliosides varied by class. Several GM2 species were paradoxically increased, likely due to inhibition of β-glucosidases that degrade gangliosides. Both compounds enhanced, rather than reduced, virus replication. Nonetheless, both iminosugars had surprising anti-inflammatory effects, blocking the accumulation of inflammatory cells within the liver. UV-4 treatment also resulted in a decrease in serum alanine aminotransferase (ALT) elevations associated with acute hepatitis A. These anti-inflammatory effects may result from iminosugar inhibition of cellular α-glucosidases, leading to impaired maturation of glycan moieties of chemokine and cytokine receptors, and point to the potential importance of paracrine signaling in the pathogenesis of acute hepatitis A. IMPORTANCE Hepatitis A virus (HAV) is a common cause of viral hepatitis. Iminosugar compounds block its replication in cultured cells by inhibiting synthesis of gangliosides required for HAV cell entry, but have not been tested for their ability to prevent or treat hepatitis A in vivo. We show that high doses of the iminosugars miglustat and UV-4 fail to deplete gangliosides sufficiently to block HAV infection in mice lacking a key interferon receptor. These compounds nonetheless have striking anti-inflammatory effects on the HAV-infected liver, reducing the severity of hepatitis despite enhancing chemokine and cytokine expression resulting from hepatocyte-intrinsic antiviral responses. We propose that iminosugar inhibition of cellular α-glucosidases impairs maturation of glycan moieties of chemokine and cytokine receptors required for effective signaling. These data highlight the potential importance of paracrine signaling pathways in the inflammatory response to HAV, and add to our understanding of HAV pathogenesis in mice.

Mutant p53-ENTPD5 control of the calnexin/calreticulin cycle: a druggable target for inhibiting integrin-α5-driven metastasis

“…We hypothesized that α-glucosidase inhibitors could reduce the binding of unfolded ITGA5 precursors to the chaperones and, in turn, mitigate the consequences of increased CANX/CALR cycle activity. To test our hypothesis, we evaluated two chemically distinct classes of inhibitors: acarbose, a widely used antidiabetic drug that inhibits intestinal α-glucosidases [ 50 ], and UV-4, an antiviral iminosugar that inhibits endoplasmic α-glucosidases required for the maturation of viral glycoproteins [ 51 – 53 ]. Treating the tumor cells with either of the drugs resulted in a dose-dependent reduction of ITGA5 protein expression, but not mRNA expression (Fig.…”

IRF3-mediated pathogenicity in a murine model of human hepatitis A

“…Ifnar1 -/mice recapitulate many of the cardinal features of hepatitis A in humans, including hepatocyte apoptosis and elevated serum ALT levels [3,7]. Apoptotic hepatocytes are surrounded by inflammatory cells, and CD4+ and CD8+ T cells, NK cells, and F4/80+CD11 + macrophages are all increased in number in the liver [3,33]. However, antibody depletion of CD4+ or CD8+ T cells or NK/NKT cells, or clodronate-depletion of phagocytic cells prior to virus challenge does not prevent hepatocellular apoptosis or liver inflammation [3].…”

Recent advances in hepatitis A virus research and clinical practice guidelines for hepatitis A virus infection in Japan

“…21,23,24,227 Interferon and cytokine signaling play a role in inhibition of HAV replication. 211,216,[228][229][230][231][232] Ultrastructural alterations were observed in HAV-infected cells, such as large polyribosomes, swelling of the perinuclear space and the endoplasmic reticulum, and dilatation of Golgi cisternae. 233 Glucose-regulated protein 78, which is a chaperone protein and orchestrates the unfolded protein response in the endoplasmic reticulum, is involved in regulation of HAV replication.…”

“…Cleavage of mitochondrial antiviral signaling by HAV 3ABC protease boosts viral replication and promotes disease pathogenesis, and HAV protease inhibitors suppress these reactions, as well as cleaving HAV polyproteins, resulting in inhibition of HAV replication 21,23,24,227 . Interferon and cytokine signaling play a role in inhibition of HAV replication 211,216,228–232 . Ultrastructural alterations were observed in HAV‐infected cells, such as large polyribosomes, swelling of the perinuclear space and the endoplasmic reticulum, and dilatation of Golgi cisternae 233 .…”