Re-discover the value of protein binding assessments in hepatic and renal impairment studies and its contributions in drug labels and dose decisions.

One of the key pharmacokinetic properties of most small molecule drugs is their ability to bind to serum proteins. Unbound or free drug is responsible for pharmacological activity while the balance between free and bound drug can impact drug distribution, elimination, and other safety parameters. In the hepatic impairment (HI) and renal impairment (RI) clinical studies, unbound drug concentration is often assessed; however, the relevance and impact of the protein binding (PB) results is largely limited. We analyzed published clinical safety and pharmacokinetic studies in subjects with HI or RI with PB assessment up to October 2022 and summarized the contribution of PB results on their label dose recommendations. Among drugs with HI publication, 32% (17/53) associated product labels include PB results in HI section. Of these, the majority (9/17, 53%) recommend dose adjustments consistent with observed PB change. Among drugs with RI publication, 27% (12/44) of associated product labels include PB results in RI section with the majority (7/12, 58%) recommending no dose adjustment, consistent with the reported absence of PB change. PB results were found to be consistent with a tailored dose recommendation in 53% and 58% of the approved labels for HI and RI section, respectively. We further discussed the interpretation challenges of PB results, explored treatment decision factors including total drug concentration, exposure-response relationships, and safety considerations in these case examples. Collectively, comprehending the alterations in free drug levels in HI and RI informs treatment decision through a risk-based approach.

[Current status and challenges of clinical research and development of new drugs for liver diseases].

Liver disease is a serious public health problem worldwide, affecting human health. However, there are still many unmet needs for the treatment of liver disease, especially with new therapeutic drugs. At present, there is no treatment method to eradicate the hepatitis B virus, nor are there therapeutic drugs for liver fibrosis, liver failure, and others. Chemotherapy and targeted immunotherapy are still unsatisfactory for liver cancer. This article provides an overview of the current status and challenges that arise in new drug research and development for liver diseases.

Antagonizing Activin A/p15INK4b Signaling as Therapeutic Strategy for Liver Disease.

BACKGROUND/AIM: Activin A is involved in the pathogenesis of human liver diseases, but its therapeutic targeting is not fully explored. Here, we tested the effect of novel, highly specific small-molecule-based activin A antagonists (NUCC-474/555) in improving liver regeneration following partial hepatectomy and halting fibrosis progression in models of chronic liver diseases (CLDs). METHODS: Cell toxicity of antagonists was determined in rat hepatocytes and Huh-7 cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay. Hepatocytes and hepatic stellate cells (HSCs) were treated with activin A and NUCC-555 and analyzed by reverse transcription-polymerase chain reaction and immunohistochemistry. Partial hepatectomized Fisher (F)344 rats were treated with NUCC-555, and bromodeoxyuridine (BrdU) incorporation was determined at 18/24/36/120/240 h. NUCC-555 was administered into thioacetamide- or carbon tetrachloride-treated F344 rats or C57BL/6 mice, and the fibrosis progression was studied. RESULTS: NUCC-474 showed higher cytotoxicity in cultured hepatic cells; therefore, NUCC-555 was used in subsequent studies. Activin A-stimulated overexpression of cell cycle-/senescence-related genes (e.g., p15INK4b, DEC1, Glb1) was near-completely reversed by NUCC-555 in hepatocytes. Activin A-mediated HSC activation was blocked by NUCC-555. In partial hepatectomized rats, antagonizing activin A signaling resulted in a 1.9-fold and 2.3-fold increase in BrdU+ cells at 18 and 24 h, respectively. Administration of NUCC-555 in rats and mice with progressing fibrosis significantly reduced collagen accumulation (7.9-fold), HSC activation indicated by reduced alpha smooth muscle actin+ and vimentin+ cells, and serum aminotransferase activity. CONCLUSIONS: Our studies demonstrate that activin A antagonist NUCC-555 promotes liver regeneration and halts fibrosis progression in CLD models, suggesting that blocking activin A signaling may represent a new approach to treating people with CLD.

A patent review on HMGB1 inhibitors for the treatment of liver diseases.

INTRODUCTION: HMGB1 is a non-histone chromatin protein released or secreted in response to tissue damage or infection. Extracellular HMGB1, as a crucial immunomodulatory factor, binds with several different receptors to innate inflammatory responses that aggravate acute and chronic liver diseases. The increased levels of HMGB1 have been reported in various liver diseases, highlighting that it represents a potential biomarker and druggable target for therapeutic development. AREAS COVERED: This review summarizes the current knowledge on the structure, function, and interacting receptors of HMGB1 and its significance in multiple liver diseases. The latest patented and preclinical studies of HMGB1 inhibitors (antibodies, peptides, and small molecules) for liver diseases are summarized by using the keywords 'HMGB1,' 'HMGB1 antagonist, HMGB1-inhibitor,' 'liver disease' in Web of Science, Google Scholar, Google Patents, and PubMed databases in the year from 2017 to 2023. EXPERT OPINIONS: In recent years, extensive research on HMGB1-dependent inflammatory signaling has discovered potent inhibitors of HMGB1 to reduce the severity of liver injury. Despite significant progress in the development of HMGB1 antagonists, few of them are approved for clinical treatment of liver-related diseases. Developing safe and effective specific inhibitors for different HMGB1 isoforms and their interaction with receptors is the focus of future research.

Exploring the potential of drug repurposing for liver diseases: A comprehensive study.

Drug repurposing involves the investigation of existing drugs for new indications. It offers a great opportunity to quickly identify a new drug candidate at a lower cost than novel discovery and development. Despite the importance and potential role of drug repurposing, there is no specific definition that healthcare providers and the World Health Organization credit. Unfortunately, many similar and interchangeable concepts are being used in the literature, making it difficult to collect and analyze uniform data on repurposed drugs. This research was conducted based on understanding general criteria for drug repurposing, concentrating on liver diseases. Many drugs have been investigated for their effect on liver diseases even though they were originally approved (or on their way to being approved) for other diseases. Some of the hypotheses for drug repurposing were first captured from the literature and then processed further to test the hypothesis. Recently, with the revolution in bioinformatics techniques, scientists have started to use drug libraries and computer systems that can analyze hundreds of drugs to give a short list of candidates to be analyzed pharmacologically. However, this study revealed that drug repurposing is a potential aid that may help deal with liver diseases. It provides available or under-investigated drugs that could help treat hepatitis, liver cirrhosis, Wilson disease, liver cancer, and fatty liver. However, many further studies are needed to ensure the efficacy of these drugs on a large scale.

A comprehensive review of bupleuri radix and its bioactive components： with a major focus on treating chronic liver diseases.

ETHNOPHARMACOLOGICAL RELEVANCE: Bupleuri Radix (BR) has been recognized as an essential herbal medicine for relieving liver depression for thousands of years. Contemporary research has provided compelling evidence of its pharmacological effects, including anti-inflammatory, immunomodulatory, metabolic regulation, and anticancer properties, positioning it as a promising treatment option for various liver diseases. Hepatitis, steatohepatitis, cirrhosis, and liver cancer are among the prevalent and impactful liver diseases worldwide. However, there remains a lack of comprehensive systematic reviews that explore the prescription, bio-active components, and underlying mechanisms of BR in treating liver diseases. AIM OF THE REVIEW: To summarize the BR classical Chinese medical prescription and ingredients in treating liver diseases and their mechanisms to inform reference for further development and research. MATERIALS AND METHODS: Literature in the last three decades of BR and its classical Chinese medical prescription and ingredients were collated and summarized by searching PubMed, Wiley, Springer, Google Scholar, Web of Science, CNKI, etc. RESULTS: BR and its classical prescriptions, such as Xiao Chai Hu decoction, Da Chai Hu decoction, Si Ni San, and Chai Hu Shu Gan San, have been utilized for centuries as effective therapies for liver diseases, including hepatitis, steatohepatitis, cirrhosis, and liver cancer. BR is a rich source of active ingredients, such as saikosaponins, polysaccharides, flavonoids, sterols, organic acids, and so on. These bioactive compounds exhibit a wide range of beneficial effects, including anti-inflammatory, antioxidant, immunomodulatory, and lipid metabolism regulation. However, it is important to acknowledge that BR and its constituents can also possess hepatotoxicity, which is associated with cytochrome P450 (CYP450) enzymes and oxidative stress. Therefore, caution should be exercised when using BR in therapeutic applications to ensure the safe and appropriate utilization of its potential benefits while minimizing any potential risks. CONCLUSIONS: To sum up, BR, its compounds, and its based traditional Chinese medicine are effective in liver diseases through multiple targets, multiple pathways, and multiple effects. Advances in pharmacological and toxicological investigations of BR and its bio-active components in the future will provide further contributions to the discovery of novel therapeutics for liver diseases.

Glycyrrhizin and the Related Preparations: An Inspiring Resource for the Treatment of Liver Diseases.

Liver diseases and their related complications endanger the health of millions of people worldwide. The prevention and treatment of liver diseases are still serious challenges both in China and globally. With the improvement of living standards, the prevalence of metabolic liver diseases, including non-alcoholic fatty liver disease and alcoholic liver disease, has increased at an alarming rate, resulting in more cases of end-stage liver disease. Therefore, the discovery of novel therapeutic drugs for the treatment of liver diseases is urgently needed. Glycyrrhizin (GL), a triterpene glycoside from the roots of licorice plants, possesses a wide range of pharmacological and biological activities. Currently, GL preparations (GLPs) have certain advantages in the treatment of liver diseases, with good clinical effects and fewer adverse reactions, and have shown broad application prospects through multitargeting therapeutic mechanisms, including antisteatotic, anti-oxidative stress, anti-inflammatory, immunoregulatory, antifibrotic, anticancer, and drug interaction activities. This review summarizes the currently known biological activities of GLPs and their medical applications in the treatment of liver diseases, and highlights the potential of these preparations as promising therapeutic options and their alluring prospects for the treatment of liver diseases.

First-in-class MKK4 inhibitors enhance liver regeneration and prevent liver failure.

Diminished hepatocyte regeneration is a key feature of acute and chronic liver diseases and after extended liver resections, resulting in the inability to maintain or restore a sufficient functional liver mass. Therapies to restore hepatocyte regeneration are lacking, making liver transplantation the only curative option for end-stage liver disease. Here, we report on the structure-based development and characterization (nuclear magnetic resonance [NMR] spectroscopy) of first-in-class small molecule inhibitors of the dual-specificity kinase MKK4 (MKK4i). MKK4i increased liver regeneration upon hepatectomy in murine and porcine models, allowed for survival of pigs in a lethal 85% hepatectomy model, and showed antisteatotic and antifibrotic effects in liver disease mouse models. A first-in-human phase I trial (European Union Drug Regulating Authorities Clinical Trials [EudraCT] 2021-000193-28) with the clinical candidate HRX215 was conducted and revealed excellent safety and pharmacokinetics. Clinical trials to probe HRX215 for prevention/treatment of liver failure after extensive oncological liver resections or after transplantation of small grafts are warranted.

Pharmacokinetics of long-acting lenacapavir in participants with hepatic or renal impairment.

Lenacapavir is a novel, first-in-class, multistage inhibitor of HIV-1 capsid function approved for the treatment of multidrug-resistant HIV-1 infection in combination with other antiretrovirals for heavily treatment-experienced people with HIV. Two Phase 1, open-label, parallel-group, single-dose studies assessed the pharmacokinetics (PK) of lenacapavir in participants with moderate hepatic impairment [Child-Pugh-Turcotte (CPT) Class B: score 7-9] or severe renal impairment [15 ≤ creatinine clearance (CLcr) ≤29 mL/min] to inform lenacapavir dosing in HIV-1-infected individuals with organ impairment. In both studies, a single oral dose of 300 mg lenacapavir was administered to participants with normal (n = 10) or impaired (n = 10) hepatic/renal function who were matched for age (±10 years), sex, and body mass index (±20%). Lenacapavir exposures [area under the plasma concentration-time curve from time 0 to infinity (AUCinf) and maximum concentration (Cmax)] were approximately 1.47- and 2.61-fold higher, respectively, in participants with moderate hepatic impairment compared to those with normal hepatic function, whereas lenacapavir AUCinf and Cmax were approximately 1.84- and 2.62-fold higher, respectively, in participants with severe renal impairment compared to those with normal renal function. Increased lenacapavir exposures with moderate hepatic or severe renal impairment were not considered clinically meaningful. Lenacapavir was considered generally safe and well tolerated in both studies. These results support the use of approved lenacapavir dosing regimen in patients with mild (CPT Class A: score 5-6) or moderate hepatic impairment as well as in patients with mild (60 ≤ CLcr ≤ 89 mL/min), moderate (30 ≤ CLcr ≤ 59 mL/min), and severe renal impairment.

Effectiveness and safety of direct oral anticoagulants among patients with non-valvular atrial fibrillation and liver disease: A multinational cohort study.

BACKGROUND AND AIMS: The effects of direct oral anticoagulants (DOACs) in patients with non-valvular atrial fibrillation (NVAF) and liver disease remain poorly understood. Our multinational cohort study assessed the effectiveness and safety of DOACs in this high-risk population. METHODS: We assembled two population-based cohorts in United Kingdom and in Québec of NVAF patients with liver disease initiating DOACs or vitamin K antagonists (VKAs) between 2011 and 2020. Using an as-treated exposure definition, we compared DOACs to VKAs and apixaban to rivaroxaban. After inverse probability of treatment weighting, Cox proportional hazards models estimated site-specific hazard ratios (HRs) and 95 % confidence intervals (CIs) of ischemic stroke and major bleeding. Site-specific estimates were pooled using random-effects models. Analyses were repeated among NVAF patients with cirrhosis. RESULTS: There were 11,881 NVAF patients with liver disease (2683 with cirrhosis). Among those, 8815 initiated DOACs (4414 apixaban, 2497 rivaroxaban) and 3696 VKAs. The HRs (95 % CIs) for DOACs compared to VKAs were 1.01 (0.76-1.34) for ischemic stroke and 0.87 (0.77-0.99) for major bleeding. Results were consistent among patients with cirrhosis. The HRs (95 % CIs) for apixaban compared to rivaroxaban were 0.85 (0.60-1.20) for ischemic stroke and 0.80 (0.68-0.95) for major bleeding. This decreased bleeding risk was not observed among patients with cirrhosis (HR, 1.01; 95 % CI 0.72-1.43). CONCLUSIONS: Among NVAF patients with liver disease, DOACs were as effective and slightly safer than VKAs, and apixaban was as effective but safer than rivaroxaban. The safety benefit with apixaban was not present among patients with cirrhosis.

Autoimmune liver disease triggered by SARS-CoV-2: a case report and review of the literature.

BACKGROUND: An increasing number of coronavirus disease 2019 (COVID-19) related autoimmune hepatitis (AIH) and autoimmune liver disease (AILD) has been already described so far in the last three years. This rise has set up some diagnostic and therapeutic concerns, although steroid therapy has mostly been efficient, avoiding main significant side effects. CASE REPORT: We report the case of a 52-year-old subject displaying liver function impairment at the laboratory tests while positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) swab. Needle liver biopsy showed severe portal inflammation, interface hepatitis, lobular inflammation, abundant plasma cells, bridging necrosis, endothelialitis, bile duct vanishing disease, and ductular reaction. The diagnosis of autoimmune liver disease (AILD) was performed. After a month of steroid and ursodeoxycholic acid medications, liver function fully recovered. Azathioprine was introduced, and steroids were gradually reduced. CONCLUSIONS: Probably triggered by the SARS-CoV-2-induced cytokine storm, the association between COVID-19 and autoimmune-related inflammatory injury may display a particular paradigm of AILD pathogenesis.

UDCA for Drug-Induced Liver Disease: Clinical and Pathophysiological Basis.

Drug-induced liver injury (DILI) is an adverse reaction to medications and other xenobiotics that leads to liver dysfunction. Based on differential clinical patterns of injury, DILI is classified into hepatocellular, cholestatic, and mixed types; although hepatocellular DILI is associated with inflammation, necrosis, and apoptosis, cholestatic DILI is associated with bile plugs and bile duct paucity. Ursodeoxycholic acid (UDCA) has been empirically used as a supportive drug mainly in cholestatic DILI, but both curative and prophylactic beneficial effects have been observed for hepatocellular DILI as well, according to preliminary clinical studies. This could reflect the fact that UDCA has a plethora of beneficial effects potentially useful to treat the wide range of injuries with different etiologies and pathomechanisms occurring in both types of DILI, including anticholestatic, antioxidant, anti-inflammatory, antiapoptotic, antinecrotic, mitoprotective, endoplasmic reticulum stress alleviating, and immunomodulatory properties. In this review, a revision of the literature has been performed to evaluate the efficacy of UDCA across the whole DILI spectrum, and these findings were associated with the multiple mechanisms of UDCA hepatoprotection. This should help better rationalize and systematize the use of this versatile and safe hepatoprotector in each type of DILI scenarios.

Pathophysiological-Based Nutritional Interventions in Cirrhotic Patients with Sarcopenic Obesity: A State-of-the-Art Narrative Review.

In recent decades, following the spread of obesity, metabolic dysfunction has come to represent the leading cause of liver disease. The classical clinical presentation of the cirrhotic patient has, therefore, greatly changed, with a dramatic increase in subjects who appear overweight or obese. Due to an obesogenic lifestyle (lack of physical activity and overall malnutrition, with an excess of caloric intake together with a deficit of proteins and micronutrients), these patients frequently develop a complex clinical condition defined as sarcopenic obesity (SO). The interplay between cirrhosis and SO lies in the sharing of multiple pathogenetic mechanisms, including malnutrition/malabsorption, chronic inflammation, hyperammonemia and insulin resistance. The presence of SO worsens the outcome of cirrhotic patients, affecting overall morbidity and mortality. International nutrition and liver diseases societies strongly agree on recommending the use of food as an integral part of the healing process in the comprehensive management of these patients, including a reduction in caloric intake, protein and micronutrient supplementation and sodium restriction. Based on the pathophysiological paths shared by cirrhosis and SO, this narrative review aims to highlight the nutritional interventions currently advocated by international guidelines, as well as to provide hints on the possible role of micronutrients and nutraceuticals in the treatment of this multifaceted clinical condition.

The Impact of Red Yeast Rice Extract Use on the Occurrence of Muscle Symptoms and Liver Dysfunction: An Update from the Adverse Event Reporting Systems and Available Meta-Analyses.

Red yeast rice (RYR) has a cholesterol-lowering effect due to the presence of bioactive components (monacolins, mainly monacolin K) that act by inhibiting the activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. The European Food Safety Authority (EFSA) assessed the use of RYR and, while pointing out several uncertainties regarding the available data, raised a warning related to the safety of RYR when used as a food supplement at a dose of monacolin as low as 3 mg/day. In their decision in June 2023, EFSA approved the use of monacolins from RYR at doses less than 3 mg/day. We therefore decided to interrogate the different adverse event reporting systems (FAERS and CAERS) and analyse the characteristics of the cases reported to be associated with RYR supplements, and we reviewed the most recent meta-analyses with a focus on the occurrence of muscle symptoms and liver dysfunction. In terms of all musculoskeletal disorders from September 2013 (when the first case related to RYR consumption was recorded) to 30 September 2023, 363,879 cases were reported in the FAERS, with the number of cases related to RYR consumption being very small and accounting for 0.008% of cases. In the same time frame, 27,032 cases of hepatobiliary disorders were reported, and the cases attributable to RYR ingestion accounted for 0.01% of all cases. A low rate of muscle symptoms and liver dysfunction attributed to RYR ingestion was also observed in the CAERS database, where only 34 cases of adverse muscle events and 10 cases of adverse liver events reported RYR as the suspect product, while 19 cases of both muscle events and 10 cases of adverse liver events reported it as a concomitant product. This profile mirrors that of meta-analyses of randomised clinical trials of RYR, in which RYR use was not associated with either liver dysfunction or muscular adverse symptoms.

Mortality and associated factors among people living with HIV admitted at a tertiary-care hospital in Uganda: a cross-sectional study.

BACKGROUND: Hospital admission outcomes for people living with HIV (PLHIV) in resource-limited settings are understudied. We describe in-hospital mortality and associated clinical-demographic factors among PLHIV admitted at a tertiary-level public hospital in Uganda. METHODS: We performed a cross-sectional analysis of routinely collected data for PLHIV admitted at Kiruddu National Referral Hospital between March 2020 and March 2023. We estimated the proportion of PLHIV who had died during hospitalization and performed logistic regression modelling to identify predictors of mortality. RESULTS: Of the 5,827 hospitalized PLHIV, the median age was 39 years (interquartile range [IQR] 31-49) and 3,293 (56.51%) were female. The median CD4 + cell count was 109 cells/µL (IQR 25-343). At admission, 3,710 (63.67%) were active on antiretroviral therapy (ART); 1,144 (19.63%) had interrupted ART > 3 months and 973 (16.70%) were ART naïve. In-hospital mortality was 26% (1,524) with a median time-to-death of 3 days (IQR 1-7). Factors associated with mortality (with adjusted odds ratios) included ART interruption, 1.33, 95% confidence intervals (CI) 1.13-1.57, p 0.001; CD4 + counts ≤ 200 cells/µL 1.59, 95%CI 1.33-1.91, p < 0.001; undocumented CD4 + cell count status 2.08, 95%CI 1.73-2.50, p < 0.001; impaired function status 7.35, 95%CI 6.42-8.41, p < 0.001; COVID-19 1.70, 95%CI 1.22-2.37, p 0.002; liver disease 1.77, 95%CI 1.36-2.30, p < 0.001; co-infections 1.53, 95%CI 1.32-1.78, p < 0.001; home address > 20 km from hospital 1.23, 95%CI 1.04-1.46, p 0.014; hospital readmission 0.7, 95%CI 0.56-0.88, p 0.002; chronic lung disease 0.62, 95%CI 0.41-0.92, p 0.019; and neurologic disease 0.46, 95%CI 0.32-0.68, p < 0.001. CONCLUSION: One in four admitted PLHIV die during hospitalization. Identification of risk factors (such as ART interruption, function impairment, low/undocumented CD4 + cell count), early diagnosis and treatment of co-infections and liver disease could improve outcomes.

Recent trends in the delivery of RNA drugs: Beyond the liver, more than vaccine.

RNAs are known for versatile functions and therapeutic utility. They have gained significant interest since the approval of several RNA drugs, including COVID-19 mRNA vaccines and therapeutic agents targeting liver diseases. There are increasing expectations for a new class of RNA drugs for broader applications. Successful development of RNA drugs for new applications hinges on understanding their diverse functions and structures. In this review, we explore the last five years of literature to understand current approaches to formulate a spectrum of RNA drugs, focusing on new efforts to expand their applications beyond vaccines and liver diseases.

Improvement of liver histology in hepatic sarcoidosis due to treatment with corticosteroids and ursodeoxycholic acid: a case report.

We report the case of a 48-year-old male with a history of pulmonary and ocular sarcoidosis. Non-caseating granulomas, identified histologically, are the most characteristic manifestation of sarcoidosis. Hepatic sarcoidosis is difficult to diagnose using radiological imaging. In the patient reported in this study, ultrasound and contrast-enhanced computed tomography scans identified multiple intra-abdominal lymphadenopathies, with evidence of liver and splenic infiltrations. The first liver biopsy revealed non-caseating granulomatous hepatitis consistent with hepatic sarcoidosis. The patient was treated with ursodeoxycholic acid (UDCA), but his laboratory parameters did not improve. Prednisone was initiated at a dose of 30 mg daily and slowly tapered. At a dose of 12.5 mg daily, marked improvements in the fibrotic and sarcoid-like lesions were noted at the second biopsy. A third biopsy was performed, with the patient on a prednisone taper of 5 mg/day showed mild fibrous expansion in the portal tracts and mild parenchymal necro-inflammatory lesions. However, overall, fibrosis marker levels remained stable over the course of treatment. A fourth biopsy was performed after a 5-year course of 5 mg/day prednisone. This revealed minimal lobular inflammation without fibrosis. Thus, treatment of this patient with corticosteroids and UDCA resulted in marked improvements in his biochemical and histological parameters.

Targeting Protein Phosphatases for the Treatment of Chronic Liver Disease.

There exists a huge number of patients suffering from chronic liver disease worldwide. As a disease with high incidence and mortality worldwide, strengthening the research on the pathogenesis of chronic liver disease and the development of novel drugs is an important issue related to the health of all human beings. Phosphorylation modification of proteins plays a crucial role in cellular signal transduction, and phosphatases are involved in the development of liver diseases. Therefore, this article summarized the important role of protein phosphatases in chronic liver disease with the aim of facilitating the development of drugs targeting protein phosphatases for the treatment of chronic liver disease.