Contents lists available at ScienceDirect Clinica Chimica Acta journal homepage: www.elsevier.com/locate/cca A homozygous ABCB4 mutation causing an LPAC syndrome evolves into cholangiocarcinoma Boudour Khabou a, ⁎ , Ayman Trigui b , Tahya Sellami Boudawara c , Leila Keskes d , Hassen Kamoun d , Véronique Barbu e , Faiza Fakhfakh a a Laboratory of Molecular and Functional Genetics, Faculty of Science, University of Sfax, Tunisia b Department of General Surgery, Habib Bourguiba Hospital, 3027 Sfax, Tunisia c Anatomo Pathology's Department, Habib Bourguiba Hospital, Sfax, Tunisia d Laboratory of Molecular and Human Genetics, Faculty of Medecine, University of Sfax, Tunisia e Sorbonne University Medical School, APHP, St Antoine Hospital, Medical Biology and Pathology Department, LCBGM, 75012 Paris, France ARTICLE INFO Keywords: LPAC syndrome ABCB4 mutation QPCR Cholangiocarcinoma UDCA therapy ABSTRACT Low phospholipid-associated cholelithiasis (LPAC) is characterized by the association of ABCB4 mutations and low biliary phospholipid concentration with symptomatic and recurring cholelithiasis. In the present study, we reported a case of a 63-year-old woman, who presented a biliary pain beginning at the age of 30, recurrent after cholecystectomy, along with “comet-tail shadows” revealed by ultrasonography thus, fulfilling the diagnosis of LPAC. This disease evolved into a cholangiocarcinoma. To understand the molecular basis of this phenotype, we performed the ABCB4 gene sequencing, followed by in silico analysis and Q-RT-PCR assay. The results displayed a homozygous missense sequence variation (c.140G > A, p.Arg47Gln), predicted as pathogenic according to MutPred. Accordingly, this gave rise to a decreased hepatic ABCB4 mRNA level and structural alterations of the mutated protein. Eventually, we reported, here, the first description of an ABCB4 missense mutation (p.Arg47Gln) at homozygous state in a Tunisian LPAC syndrome. An elucidation of its functional consequences was performed. Besides, this case suggests that the delayed diagnosis of LPAC syndrome and the lack of UDCA treatment may contribute in the development of complications, such as cholangiocarcinoma. 1. Introduction Low Phospholipid-Associated Cholelithiasis (LPAC syndrome – OMIM #600803) is a cholestatic disorder that affects the biliary func- tion of the liver among young adults. Clinically, the presence of in- trahepatic cholesterol stones, the recurrence of biliary symptoms after cholecystectomy and the onset of first symptoms before the age of 40 represent the three criteria characterizing LPAC. The presence of at least two of these criteria, establishes the diagnosis, which is con- solidated by a personal history of gravidic cholestasis and/or a family history of cholelithiasis[1]. This disease has recently been described in the European population and considered as a peculiar form of in- trahepatic cholelithiasis. It is caused by genetic defects in the ABCB4 gene, encoding the MDR3 protein which is the transporter of the phosphatidylcholine (PC) during bile secretion. The transported PC is crucial for the solubilization of the cholesterol and the neutralization of bile acids [2]. Indeed, the lack of PC in the bile leads to the production of lithogenic bile, with a detergent effect, responsible for the crystallization of cholesterol as stones and an excessive secretion of the gamma-glutamyltransferase (GGT) by the damaged cholangiocytes [3]. In 2013, 79 disease-causing mutations were shown in a large in- teresting study, comprising 156 LPAC patients. All truncating and 74% of missense variants were detected at heterozygous state, while 26% of the missense ones were homozygous or compound heterozygous [4]. The functional characterization pertaining to some of these mutations indicated that they altered the MDR3 protein within different degrees; however, a residual activity of MDR3 is always maintained in LPAC patients [5–7]. In addition to the LPAC syndrome, it has been shown that the MDR3 deficiency in adults is associated with a wide spectrum of hepatobiliary disorders with variable severity [8]. This variable phenotypic expression confirmed the implication of further factors, ong with the genetic component, in the development of these ABCB4-re- lated diseases, as previously suggested [9,10]. For the treatment of LPAC patients, UDCA is systematically used. This therapy allows prompt relief of symptoms, taking into account its several beneficial effects [11]. In fact, a favorable outcome has been https://doi.org/10.1016/j.cca.2019.06.007 Received 4 March 2019; Received in revised form 16 May 2019; Accepted 6 June 2019 ⁎ Corresponding author at: Molecular and Functional Genetics Laboratory, Faculty of Science, Route de la Soukra km 4, 3038 Sfax, Tunisia. E-mail address: Boudour.khabou.bio@gmail.com (B. Khabou). Clinica Chimica Acta 495 (2019) 598–605 Available online 07 June 2019 0009-8981/ © 2019 Published by Elsevier B.V. T