Pharmacological inhibition of apical sodium‐dependent bile acid transporter changes bile composition and blocks progression of sclerosing cholangitis in multidrug resistance 2 knockout mice

Abstract

Deficiency for mdr2, a canalicular phospholipid floppase, leads to excretion of low phospholipid “toxic” bile causing progressive cholestasis. We hypothesize that pharmacological inhibition of the ileal apical sodium-dependent bile acid transporter (ASBT) blocks progression of sclerosing cholangitis in mdr2−/− mice. 30-day-old, female mdr2−/− mice were fed high-fat chow containing 0.006% SC-435, a minimally absorbed, potent inhibitor of ASBT, providing on average 11 mg/kg/day of compound. Bile acids (BA) and phospholipids were measured by mass spectrometry. Compared with untreated mdr2−/− mice, SC-435 treatment for 14 days increased fecal BA excretion by 8-fold, lowered total BA concentration in liver by 65%, reduced total BA and individual hydrophobic BA concentrations in serum by >98%, and decreased plasma ALT, total bilirubin, and serum alkaline phosphatase levels by 86, 93 and 55%, respectively. Liver histology of sclerosing cholangitis improved, and extent of fibrosis decreased concomitant with reduction of hepatic profibrogenic gene expression. Biliary BA concentrations significantly decreased and phospholipids remained low and unchanged with treatment. The phosphatidylcholine/BA ratio in treated mice corrected towards a ratio of 0.28 found in wild type mice, indicating decreased bile toxicity. Hepatic RNAseq studies revealed upregulation of putative anti-inflammatory and antifibrogenic genes, including Ppara and Igf1 and downregulation of several pro-inflammatory genes, including Ccl2 and Lcn2, implicated in leukocyte recruitment. Flow cytometric analysis revealed significant reduction of frequencies of hepatic CD11b+F4/80+ Kupffer cells and CD11b+Gr1+ neutrophils, accompanied by expansion of anti-inflammatory Ly6C− monocytes in treated mdr2−/− mice. Conclusion Inhibition of ASBT reduces BA pool size and retention of hydrophobic BA, favorably alters the biliary PC/BA ratio, profoundly changes the hepatic transcriptome, attenuates recruitment of leukocytes, and abrogates progression of murine sclerosing cholangitis.

Apical Sodium-Dependent Transporter Inhibitors in Primary Biliary Cholangitis and Primary Sclerosing Cholangitis

“…More recently, the effects of ASBTi in animal models of cholestasis have been reported. Miethke et al [21] treated mdr2 -/-mice (an established animal model for chronic cholestasis with some features of sclerosing cholangitis) with SC-435 for 14 days. They observed an eightfold increase in faecal BA excretion associated with 65, 98.9 and 98.8% decrease in hepatic, serum and biliary concentrations of BAs, respectively.…”

Therapy of Primary Sclerosing Cholangitis - Today and Tomorrow

“…In Mdr2 (Abcb4) -/-mice interruption of bile acids enterohepatic circulation via potent ASBT inhibitors A4250 or SC-435 increased fecal bile acid lost and markedly reduced biliary concentrations of potentially toxic bile acids. In line with 'toxic bile' hypothesis, biliary bile acid depletion led to the amelioration of bile duct injury, inflammation and fibrosis in cholestasis [129,130] .…”

Bile Acid Uptake Transporters as Targets for Therapy

“…The ASBT inhibitor A4250, previously used in a clinical phase I study [50], showed improvement of cholestatic liver injury in multi-drug resistance P-glycoprotein 2-deficient mice [51]. In a similar study, 2 weeks of treatment with SC-435, a different ASBT inhibiting small molecule, demonstrated reduced bile acid pool size and attenuation of cholestasis in the same mouse model [52]. At present, human studies are being performed, but the outcome of these promising studies is not published yet (https://clinicaltrials.gov/ct2/show/NCT02061540).…”