September 17, 2012

September 17, 2012 | By John Carroll

Brian Starkey didn't have insurance coverage when he was diagnosed with hepatitis C. So when a doctor pointed him to a study of Bristol-Myers Squibb's ($BMY) experimental BMS-094, he leaped at a chance to get free care. And as The Kansas City Star reports in an in-depth profile, that decision may have cost him his life.

Starkey's sudden death after taking the treatment--along with severe heart ailments suffered by a big group of the 113 enrollees in the drug study--scuttled a program that inspired Bristol's $2.5 billion acquisition of Inhibitex. And the attorney representing Starkey's family tells the Midwestern newspaper that the risks of toxicity were already apparent when Bristol rushed into clinical studies, anxious to grab a lead in the frenzied race to develop a new hep C treatment that would not require interferon.

"This was a poorly designed study that caused serious injury," attorney Robert Hilliard told the Star. "I'm convinced that they rushed it into trials in order to get FDA approval." Hilliard's firm represents a half dozen of the injured patients, several with severe heart damage. One of the patients requires a heart transplant. All are suing Bristol.

BMS' response: "Based on our due diligence, we believed that we could develop BMS-986094 at a dose that would provide sufficient viral suppression and resistance coverage while also having an acceptable safety and tolerability profile." Bristol has also promised to help officials and other companies like Idenix ($IDIX) to get to the bottom of the drug reactions as quickly as possible.

The patient reactions in the 094 study have also forced Idenix to slam the brakes on two of its experimental programs as the FDA carefully considers whether patients are being exposed to unnecessary risks.

For Starkey's family, including his 19-month-old son, the exercise in caution came too late.

- here's the story from The Kansas City Star

Related Articles:
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Bristol mulls write-down of key hep C drug
Analyst reads last rites after heart failure scuttles Bristol hep C study

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Why 88% of US military veterans with HCV are not treated

Journal of Hepatology
Volume 57, Issue 4 , Page 924, October 2012

Bennet Cecil

Hepatitis C Treatment Centers, 1009A Dupont Square N, Louisville, KY 40207, USA

Received 21 March 2012; accepted 28 March 2012. published online 16 April 2012.

To the Editor:

The article in the February issue of the Journal of Hepatology reported that less than 12% of American military veterans identified with HCV were treated with antiviral therapy [1]. The Veterans Administration does not want to spend adequate funds to cure patients with hepatitis C. Dr. Kenneth Kizer, Under Secretary for Health in the US Department of Veterans Affairs (VA), gave HCV a high priority but unfortunately he left the VA in 1999. Subsequent leadership has not shown enthusiasm for treating HCV.

The Director of Pharmacy and the Chief of Staff at my local VA hospital told me that I spent too much money treating HCV. Boceprevir and telaprevir are both on the hospital formulary but telaprevir prescriptions are routinely denied because it is more expensive. Patients must jump multiple hurdles before qualifying for antiviral therapy. No one would refuse to give coronary artery stents or bypass grafts to a veteran who smokes but veterans who do not completely abstain from alcohol for three months are refused antiviral therapy. In spite of difficulties, 585 of 1372 (43%) HCV RNA positive patients received antiviral therapy between 1998 and 2010 at our local VA hospital; 226 of 583 treated (39%) achieved SVR [2]. 36% of deaths were from HCC or liver failure. Veterans with sustained viral response had substantially improved survival. Effective antiviral therapy improves prognosis [3], [4]. Less than 2% of Americans die from liver disease, but more than one third of veterans with HCV die prematurely from complications of cirrhosis [2], [5]. According to a 2010 national VA report, deaths in veterans with HCV have more than tripled, “Between 2000 and 2008, the annual number of all cause deaths recorded for Veterans with chronic HCV rose from 1259 (1129 per 100,000 in VHA care) to 5967 (4049 per 100,000 in VHA care), respectively” [6].

Legislation should be passed allowing veterans with HCV to prequalify for their choice of Medicaid or Medicare so that they can obtain antiviral therapy in the private sector. Since Dr. Kizer is no longer in charge of the VA, it is very clear that the VA is not going to treat very many of them.

Conflict of interest

Speakers Bureau Vertex Pharmaceuticals.

References

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Key Clinical Endpoints Met: JX-594 can be Safely and Efficiently Delivered Through Systemic Route, and Standard-of-Care Sorafenib Can Be Safely Administered Sequentially After JX-594, Opening Door to New Clinical Perspectives

BERLIN, Sept. 17, 2012 /PRNewswire/ -- Jennerex, Inc., a private, clinical-stage biotherapeutics company focused on the development and commercialization of first-in-class targeted oncolytic immunotherapies, presented Phase 2 clinical data of JX-594 delivered first intravenously and subsequently through intra-tumoral route demonstrating safety as well as disease control and tumor responses in patients with hepatocellular carcinoma (liver cancer, HCC). The data were presented in an oral presentation at the International Liver Cancer Association (ILCA) Annual Meeting in Berlin, Germany, by Mong Cho, M.D., from Pusan National University Yangsan Hospital, South Korea.

Twenty five Asian patients with advanced HCC, 20 of whom were refractory to sorafenib, were treated with an initial intravenous dose of JX-594, and the majority of patients then received sequential intra-tumoral doses of JX-594 at week one and three. The majority of patients subsequently received treatment with sorafenib.

The primary objective of this study was to determine the safety of JX-594 followed by sorafenib in patients with advanced HCC. The sequential treatment regimen was well tolerated with transient flu-like symptoms and transient leukopenia being the most common side effects related to JX-594. The sorafenib side effects observed were consistent with the expected toxicity profile of this product.

Secondary endpoints included the effect of the sequential treatment of JX-594 followed by sorafenib on disease control and tumor response. Evidence of antitumor activity was observed in both sorafenib-naive and sorafenib-refractory patients.

Importantly, this trial also demonstrated the feasibility of the systemic administration of the product (through intravenous injection).

"Our ability to deliver JX-594 intravenously to liver cancer tumors, further confirmed by these encouraging data, is a key attribute that sets it apart from other therapies in the class of oncolytic immunotherapies," stated David H. Kirn, M.D., founder, chief medical officer and president of R&D of Jennerex. "In the Phase 2 trial presented at ILCA, JX-594 demonstrated its ability to selectively target and destroy tumors following intravenous infusion. This finding confirms the ability of JX-594 to target both primary and metastatic, or distant, tumors which we believe is important in this HCC patient population and most cancers."

"We have treated more than 160 patients with JX-594 to date and are actively enrolling a multinational Phase 2b study in second line treatment of liver cancer patients, a Phase 2 all-IV trial in first line HCC patients, and a Phase 2 study in colorectal cancer. The data presented today build on the growing body of promising clinical data showing that JX-594 has a direct anti-tumor effect and can stimulate an immune response killing cancer cells," stated Laurent Fischer, M.D., president and chief executive officer of Jennerex. "We are excited with the progress we are making in our JX-594 program and believe it has the potential to advance patient care across multiple types of cancer."

The abstract (#2012-1304) entitled "Phase 2 Trial Of JX-594, A Targeted Multi-Mechanistic Oncolytic Vaccinia Virus, Followed By Sorafenib In Patients With Advanced Hepatocellular Carcinoma (HCC)" was presented at the International Liver Cancer Association Annual Meeting in Berlin.

About this Trial:

Twenty five Asian patients with advanced HCC, 20 of whom were refractory to sorafenib, were treated with an initial intravenous dose of JX-594, and the majority of patients then received sequential intratumoral doses of JX-594 at week one and three. The majority of patients subsequently received treatment with sorafenib.

Following treatment with JX-594 alone at four weeks, 62 percent of patients had disease control as measured by modified RECIST (tumor burden measurement). Tumor biopsies of four patients following intravenous infusion showed four of four patients had local infection of JX-594 in tumor tissue while normal liver tissue was not affected, providing further evidence of JX-594's tumor selectivity and the ability to administer JX-594 intravenously. Furthermore, after six or 12 weeks, 59 percent of patients had disease control as measured by modified RECIST and 75 percent of patients had objective responses by Choi criteria. 85 percent of patients had disease control by mRECIST and /or Choi response.

JX-594: A Multi-Mechanistic Approach To Targeting Cancer
JX-594 is a proprietary, engineered oncolytic immunotherapy designed to selectively target and destroy cancer cells through three diverse mechanisms of action: 1) the lysis of cancer cells 2) the stimulation of an immune response against cancer cells, (i.e., active immunotherapy), and 3) the shutdown of the blood supply to tumors. Phase 1 and Phase 2 clinical trials in multiple cancer types to date have shown that JX-594, delivered either directly into tumors or intravenously, induces tumor shrinkage and/or necrosis and is well-tolerated (over 160 patients treated to date). Objective tumor responses have been demonstrated in a variety of cancers including liver, colon, kidney, lung cancer and melanoma. JX-594 has had a favorable, predictable and generally mild safety profile to date which includes flu-like symptoms that resolve in 24 to 48 hours.

JX-594 takes advantage of the natural attributes of poxviruses and was engineered to target and destroy solid tumors both systemically and locally. The vaccinia virus backbone of JX-594 has been used safely in millions of people as part of a worldwide vaccination program. This strain naturally targets cancer cells due to common genetic abnormalities in cancer cells. JX-594 was engineered to enhance this cancer-selectivity by inactivating its thymidine kinase (TK) gene and encode the immunogenic GM-CSF gene, to enhance the immune response against cancer cells.

Hepatocellular Carcinoma: A Global Unmet Need
Hepatocellular carcinoma is the fifth most common cancer worldwide and the third leading cause of cancer death, with over 600,000 new cases diagnosed annually resulting in more than 90 percent mortality. The annual incidence rate in the U.S., Europe, Japan and China are estimated to be 20,000, 55,000, 40,000 and 350,000 patients, respectively. The only treatment approved for HCC is sorafenib. There is no treatment approved for patients who fail sorafenib.

About Jennerex's Partners for JX-594
Transgene (NYSE Euronext Paris: FR0005175080), a bio-pharmaceutical company specialized in the development of immunotherapeutic products, holds an exclusive license to develop and commercialize JX-594 in Europe and neighboring countries. Green Cross Corporation, a leading company in the development, manufacturing, and commercialization of viral vaccines and other biological products, holds an exclusive license to develop and commercialize JX-594 in South Korea, and Lee's Pharmaceutical Ltd. holds an exclusive license to develop and commercialize JX-594 in China.

Transgene, a member of the Institut Merieux Group, is a publicly traded French biopharmaceutical company dedicated to the development of therapeutic vaccines and immunotherapeutic products in oncology and infectious diseases, and has five compounds in clinical development: TG4010 and JX-594 (TG6006) having completed initial phase II trials, TG4001 in phase IIb trial, TG4040 in phase II trial and TG4023 in phase I trial. Transgene has concluded strategic agreements for the development of two of its immunotherapy products, an option agreement with Novartis for the development of TG4010 to treat various cancers, and an in-licensing agreement with U.S.-based Jennerex Biotherapeutics, Inc., to develop and market JX-594 (TG6006), an oncolytic product. Transgene has bio-manufacturing capacities for viral-based products. Additional information about Transgene is available on the internet at www.transgene.fr

Green Cross Corp. is a publicly traded and leading Korean biopharmaceutical company specialized in development and commercialization of vaccines, plasma-derivatives, recombinant proteins and therapeutic antibodies in oncology and infectious diseases. Green Cross Corp. has been collaborating with Jennerex in Korea since 2006 to jointly conduct the Phase 1 and 2 clinical trials in patients with liver cancer. Additional information about Green Cross Corp. is available on the internet at www.greencross.com.

Lee's Pharmaceutical Holdings Limited is a public biopharmaceutical company with over 16 years operation in China's pharmaceutical industry. It is fully integrated with solid infrastructures in drug development, clinical development, regulatory, manufacturing, sales and marketing in China with global perspectives and currently markets nine products. Lee's Pharma focuses on several different areas such as cardiovascular and infectious diseases, dermatology, oncology, gynecology and others. It has more than 30 products under different development stages stemming from both internal R&D as well as from the recent acquisition of licensing and distribution rights from various U.S. and European companies. The mission of Lee's is to become a successful biopharmaceutical group in Asia providing innovative products to fight diseases and improve health and quality of life. Additional information about Lee's Pharma is available on the internet at www.leespharm.com.

About Jennerex
Jennerex, Inc. is a clinical-stage biotherapeutics company focused on the development and commercialization of first-in-class, breakthrough targeted oncolytic immunotherapy products for cancer. The Company's lead product JX-594 is currently in an international, randomized Phase 2b clinical trial (TRAVERSE) in patients with advanced primary liver cancer who have failed sorafenib therapy. In addition, JX-594 is being tested in the same patient population in combination with sorafenib. JX-594 is also in a Phase 1/2 clinical trials in patients with treatment-refractory colorectal cancer. Published studies designed to establish optimal dose levels and the safety profile of JX-594 have shown its ability to selectively target and cause destruction of a variety of common solid tumor types and trigger a potent immune response. JX-594 and other product candidates under development are designed to attack cancer tumors through three diverse mechanisms of action: the lysis of cancer cells through targeted viral replication, the ablation of the blood supply to tumors and the stimulation of the body's immune response against the cancer. Jennerex is headquartered in San Francisco and has related research and development operations in Ottawa, Canada and Pusan, South Korea. For more information about Jennerex, please visit www.jennerex.com. For studies evaluating JX-594 please visit www.clinicaltrials.gov.

SOURCE Jennerex, Inc.

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SAN FRANCISCO, Sept. 17, 2012 /PRNewswire-USNewswire/-- Getting reliable, easy-to-understand information about a groundbreaking new HIV prevention strategy just got easier. The strategy, known as pre-exposure prophylaxis (PrEP), is the focus of a new website, PrEPfacts.org, developed by San Francisco AIDS Foundation in collaboration with the San Francisco Department of Public Health (SFDPH), National Institute of Allergy and Infectious Diseases, and other community partners.

"Ever since PrEP first emerged as a viable and effective strategy to prevent new HIV infections, we have seen a lot of misinformation about it," said Neil Giuliano, CEO of San Francisco AIDS Foundation. "So we set out to create a website that delivers all the facts, in a way that's engaging and interactive, so that gay and bisexual men and transwomen in our community have the best possible information to determine if PrEP might be a valuable tool in their lives."

In July, the U.S. Food and Drug Administration approved the drug Truvada for PrEP. It is for use by HIV-negative individuals at risk for infection as part of a comprehensive prevention package that includes regular condom use, HIV testing, and risk reduction and adherence counseling.

"Right now in San Francisco, we are launching the first demonstration project to determine how best to implement PrEP in different communities," said Dr. Albert Liu, Director of HIV Prevention Intervention Studies at SFDPH. "This new website, PrEPfacts.org, is an important resource to provide our community with accurate, timely, and relevant information about this new HIV prevention strategy."

PrEPfacts.org boldly states that "love may have a new defender" in the fight against HIV/AIDS. The site gives an overview of PrEP, answers commonly asked questions and dispels many of the myths about the strategy, provides information about the upcoming demonstration project in San Francisco, and looks ahead to other HIV prevention interventions that are on the horizon.

The website is a collaboration between San Francisco AIDS Foundation; Project Inform; San Francisco HIV Vaccine Trials Network and HIV Prevention Trials Network; Bridge HIV; Magnet, a program of San Francisco AIDS Foundation which is leading demonstration project recruitment efforts in the Castro neighborhood; and San Francisco City Clinic, which is enrolling participants for the demonstration study. The website is sponsored and funded by Be The Generation, an initiative of National Institute of Allergy and Infectious Diseases.

For more information, visit www.PrEPfacts.org.

About San Francisco AIDS Foundation
No city experienced epidemic levels of HIV faster than San Francisco. At San Francisco AIDS Foundation, we work to end the epidemic where it first took hold, and eventually everywhere. Established in 1982, our mission is the radical reduction of new infections in San Francisco. Through education, advocacy, and direct services for prevention and care, we are confronting HIV in communities most vulnerable to the disease. We refuse to accept that HIV transmission is inevitable. For more information, visit www.sfaf.org.

Contact: Ryan McKeel
(415) 487-3071
rmckeel@sfaf.org

SOURCE San Francisco AIDS Foundation

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Alabama's segregation for inmates with HIV faces court scrutiny

By Verna Gates

BIRMINGHAM, Alabama | Mon Sep 17, 2012 7:53am EDT

BIRMINGHAM, Alabama (Reuters) - Alabama, one of two U.S. states that segregate inmates with HIV from the rest of their prison population, will seek to defend the policy against a class action lawsuit headed to trial in federal court on Monday.

The American Civil Liberties Union sued Alabama in 2011 for what the group contends is a discriminatory practice that prevents most HIV-positive inmates from participating in rehabilitation and retraining programs important for their success after prison.

The state says the civil liberties group has failed to prove that there would be no significant risk of the infection being transmitted to other prisoners if inmates with HIV were fully integrated, according to court documents.

An appeals court upheld the segregated housing policy in 1999, but ACLU attorney Margaret Winter said advances in treatment for HIV infection warranted the court taking another look at the practice.

"It is based on an uneducated view on HIV and how it is transmitted, which really goes back to the dark ages of when it first started and there was hysteria," she said.

South Carolina is the only other U.S. state that houses inmates with HIV away from other prisoners. Mississippi ceased a similar practice in March 2010 and has since integrated inmates with the infection, Winter said.

Two of Alabama's 29 prisons have dormitories set aside specifically for prisoners with HIV. A handful of prisoners have been allowed to live and work in non-segregated settings in two work-release programs, Winter said.

Approximately 270 inmates out of the 26,400 in the state prison system have tested positive for the virus and none has developed AIDS, according to Alabama Department of Corrections spokesman Brian Corbett.

The inmates with HIV live, eat and have their recreation time apart from the general population, according to court documents filed by the ACLU. Male inmates in the HIV dormitories are given white arm bands that signal their medical status and are to be worn at all times, the group said.

One HIV-positive prisoner received 21 disciplinary days and lost six months of good-behavior time off his sentence for sitting in the general population cafeteria, according to court documents.

"First, we are isolated ... like we are contagious animals," Dana Harley, another prisoner who is a plaintiff in the case, said in a letter included in the court file. "It is like punishment three times over."

Restricting inmates with HIV from programs such as prison jobs and work release "could harm their reentry into society," Winter said.

At the trial, expected to last one month in Montgomery, Alabama, the ACLU will argue that Alabama's policy violates the Americans with Disabilities Act. The state of Alabama contends that HIV does not qualify as an impairment under federal law.

(Editing by Colleen Jenkins and David Brunnstrom)

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Journal of Hepatology
Volume 57, Issue 4 , Pages 897-909, October 2012

Matthew T. Kitson, Stuart K. Roberts

Department of Gastroenterology, The Alfred, Melbourne, Australia

Received 20 February 2012; received in revised form 29 March 2012; accepted 1 April 2012. published online 24 May 2012.

Summary

Vitamin D is synthesized predominantly in the liver and functions as an important secosteroid hormone with pleiotropic effects. While its key regulatory role in calcium and bone homeostasis is well established, recently there is increasing recognition that vitamin D also regulates cell proliferation and differentiation, and has immunomodulatory, anti-inflammatory and anti-fibrotic properties. These non-skeletal effects are relevant in the pathogenesis and treatment of many causes of chronic liver disease. Vitamin D deficiency is frequently present in chronic liver disease and may predict non-response to antiviral therapy in chronic hepatitis C. Small studies suggest that vitamin D supplementation improves sustained viral response rates, while 1α-hydroxylase polymorphisms and vitamin D-binding protein are also implicated in therapeutic outcomes. Vitamin D deficiency also closely relates to the severity of non-alcoholic fatty liver disease (NAFLD) and is implicated in the pathogenesis of insulin resistance, a key factor in the development of NAFLD. In preclinical studies, phototherapy and vitamin D supplementation ameliorate NAFLD histopathology, while vitamin D is a powerful anti-fibrotic against thioacetamide liver injury. In liver transplant recipients severe vitamin D deficiency predicts, and vitamin D supplementation prevents, acute cellular rejection. The role of vitamin D in the activation and regulation of both innate and adaptive immune systems may explain its importance in the above liver diseases. Further prospective studies are therefore warranted to investigate the therapeutic impact of vitamin D supplementation in chronic liver disease.

Keywords: Vitamin D, Cholecalciferol, Liver disease, Liver fibrosis

Introduction

Vitamin D is an important secosteroid hormone with pleiotropic effects (Table 1). While its role in the regulation of calcium and bone homeostasis is well established, recently there is increasing recognition that vitamin D has immunomodulatory, anti-inflammatory and anti-fibrotic properties and plays an important role in the regulation of cell proliferation and differentiation. These extraskeletal effects are relevant in the pathogenesis and treatment of many causes of chronic liver disease.

Table 1. Pleiotropic effects of vitamin D.

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Vitamin D synthesis and metabolism

Vitamin D undergoes a 3-step activation process before it interacts with the vitamin D receptor. The majority of circulating vitamin D is synthesized in the skin as a result of exposure to sunlight. The initial step involves ultraviolet-B radiation (wavelength 290–315nm) converting the cholesterol metabolite 7-dehydrocholesterol into previtamin D3 in the lower epidermis, which is rapidly converted to vitamin D3 in a heat-dependent process. However, excessive sunlight exposure does not cause vitamin D intoxication because excess vitamin D3 is destroyed by sunlight [1]. Only a small proportion of vitamin D is obtained from dietary sources such as fatty fish, eggs, UV-irradiated mushrooms, supplements, and artificially fortified foods (Table 2). Dietary-derived vitamins D2 (ergocalciferol) and D3 (cholecalciferol) are absorbed via a bile-acid dependent process whereby vitamin D is incorporated into micelles in the intestinal lumen, then absorbed by enterocytes and packaged into chylomicrons that are then transported to the venous circulation via lymphatic drainage. Vitamin D from both skin synthesis and dietary sources can either be stored in adipocytes or undergo 25-hydroxylation in the liver. This process is mediated by the 25-hydroxylases, which are cytochrome P450 isoforms that include the important microsomal CYP2R1 and the mitochondrial CYP27A1 enzymes. This produces the main circulating, though biologically inactive, form 25-hydroxyvitamin D [25(OH)D], or calcidiol, which has a long half-life of 2–3weeks and is therefore used to assess vitamin D status. The vast majority (88%) of serum 25(OH)D is bound to vitamin D-binding protein (DBP), which is also known as Gc or the group-specific component of globulin. DBP is a 58kDa α-macroglobulin almost exclusively synthesized by the liver and a member of the albumin gene family located on chromosome 4, with high sequence homology to albumin and α-fetoprotein [2]. It is highly polymorphic, having three common isoforms, Gc1F, Gc1S, and Gc2, that display marked racial variation [3], with the Gc1F isoform having the highest affinity for vitamin D metabolites. DBP has anti-inflammatory and immunomodulatory functions independent of its role as the carrier of vitamin D [4], [5].

Table 2. Sources of vitamin D.

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The final step in the synthesis of vitamin D is 1α-hydroxylation that predominantly occurs in the proximal tubule of the kidney but also to a lesser extent in lymphocytes and parathyroid tissue. It is mediated by 1α-hydroxylase (CYP27B1) that produces the active form 1α,25-dihydroxyvitamin D [1α,25(OH)2D] or calcitriol, which is also highly bound to DBP (85%) [2] and has a half-life of only 4h. 1α,25(OH)2D is the ligand that activates the vitamin D receptor (VDR). This then forms a heterodimer with the retinoid X receptor that acts as a transcription factor that binds to vitamin D response elements in the promoter region of target genes. 1α-hydroxylation is under the influence of factors such as serum phosphate and calcium concentration, parathyroid hormone (PTH), fibroblast growth factor 23 and genetic polymorphisms of CYP27B1. 1α,25(OH)2D acts in a negative feedback loop to decrease its own synthesis and increase the expression of 25-hydroxyvitamin D-24-hydoxylase (CYP24A1), which catabolizes 1α,25(OH)2D into calcitroic acid, a biologically inert agent excreted in the bile (Fig. 1).

Fig. 1. Vitamin D synthesis.

PIIS0168827812003601_gr1_lrg

VDR is expressed in most tissues and cells of the human body, including liver, pancreas, and several immune cells including monocytes, macrophages, T lymphocytes, B lymphocytes, natural killer (NK) cells, and dendritic cells (DC), with expression most abundant on the epithelial cells of the gastrointestinal tract. As a transcription factor activated by 1α,25(OH)2D, VDR directly or indirectly regulates the expression of more than 200 genes that influence cell proliferation, differentiation and apoptosis, as well as immunomodulation and angiogenesis [6]. Studies in VDR null mice highlight the broad physiologic function of vitamin D [7].

Vitamin D deficiency

Vitamin D deficiency is broadly defined as a serum 25(OH)D level <50nmol/L (<20ng/ml). Levels between 75 and 125nmol/L (30–50ng/ml) are considered optimal as PTH levels rise when 25(OH)D is <75nmol/L (30ng/ml); hence, levels between 50 and 75nmol/L (20–30ng/ml) are increasingly recognized to represent vitamin D insufficiency [8], [9], [10], [11]. Using these definitions, it is estimated that more than 1billion people worldwide are either vitamin D deficient or insufficient [12], with the elderly and those with chronic medical illness most at risk. However, even amongst healthy young people, vitamin D deficiency and insufficiency are still common [13], [14], [15]. A meta-analysis of 18 randomized-controlled trials involving 57,311 participants shows that subjects randomized to receive vitamin D supplementation, at a mean daily dose of 528 IU, have a statistically significant 7% reduction in all-cause mortality over a mean follow-up duration of 5.7years [16]. Vitamin D deficiency has a deleterious clinical impact on a number of important medical conditions (Table 3).

Table 3. Clinical impact of vitamin D deficiency.

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Vitamin D and chronic liver disease

The liver is a pivotal organ in the synthesis of vitamin D. It is the site where 25-hydroxylation occurs and where the vast majority of DBP is synthesized. In those with chronic liver disease (CLD) the prevalence of vitamin D insufficiency (<75nmol/L) is almost universal, with vitamin D deficiency (<50nmol/L) present in around two-thirds of subjects. Even in the absence of cirrhosis, vitamin D deficiency is present in the majority of subjects. In those with cirrhosis, the prevalence of severe vitamin D deficiency (<25nmol/L) increases with increasing severity of synthetic liver dysfunction [17], [18]. Notably, in those about to undergo liver transplantation, the frequency of 25(OH)D and 1α,25(OH)2D deficiency is 84% and 77%, respectively, with transplantation resulting in a marked increase in 25(OH)D, 1α,25(OH)2D, and DBP levels [19].

The high prevalence of vitamin D deficiency in this population occurs regardless of the etiology of liver disease [20], [21]. Synthetic liver dysfunction is not entirely responsible, as vitamin D deficiency is still highly prevalent in those with non-cirrhotic liver disease [17]. 25(OH)D levels normalize after oral or parenteral administration of vitamin D in patients with cirrhosis, indicating that 25-hydroxylation is preserved in this patient population [22], [23]. Serum DBP levels, which play a critical role in the transport and bioavailability of vitamin D, are moderately decreased in cirrhosis [24], [25]. However, as only 5% of DBP binding sites are occupied at any one time with vitamin D metabolites [2], profound liver dysfunction is required for low DBP levels to exert a significant contributing role to vitamin D deficiency in chronic liver disease.

Vitamin D deficiency in CLD is likely to result from a number of mechanisms. In addition to those described above, those patients with a chronic medical illness such as liver disease are more likely to have lower levels of sunlight exposure and/or inadequate dietary intake of vitamin D. Moreover, luminal absorption of dietary sources of vitamin D may be hindered by intestinal edema complicating portal hypertension and/or impaired bile salt dependent micellar incorporation due to cholestasis.

Vitamin D and chronic hepatitis C

Around 170million people worldwide have chronic hepatitis C (CHC) infection [26], causing a substantial burden of chronic liver disease globally [27]. Vitamin D deficiency is more prevalent in CHC subjects than healthy controls, even in those with minimal liver fibrosis. The majority of subjects with CHC are vitamin D deficient (<50nmol/L) with 25% having severe deficiency (<25nmol/L) [28], [29]. Current understanding of the mechanisms underlying the high prevalence of vitamin D deficiency in CHC is incomplete.

Nevertheless, recent evidence suggests that vitamin D may impact upon clinical outcomes and treatment response. Fundamental to this are several in vitro studies showing that vitamin D inhibits hepatitis C virus (HCV) replication in a dose-dependent manner [30], [31], [32]. Moreover, an association between baseline vitamin D status and treatment response to pegylated-interferon (PEG-IFN) and ribavirin (RBV) has recently been established (Fig. 2). Pre-treatment vitamin D deficiency is reportedly an independent predictor of failure to achieve a sustained virologic response (SVR) in HCV genotype 1 (HCV-1), [28], [33], and 2/3 infection [29]. However, 25(OH)D level is not associated with SVR in HCV–HIV co-infection [34]. In HCV-1 infection, the rs12979860 C/T polymorphism upstream of the interleukin-28B (IL28B) gene on chromosome 19 is the strongest pre-treatment predictor of SVR [35]. Baseline vitamin D status is independent of, but additive to, the IL28B genotype in predicting SVR in HCV-1. The highest SVR rate occurs in subjects who have the favorable CC genotype and 25(OH)D levels >50nmol/L [33].

PIIS0168827812003601_gr2_lrg

To date, there is limited data evaluating vitamin D supplementation in CHC treatment. Two small prospective randomized controlled studies from Israel showed that those subjects who received vitamin D3 supplementation of 2000IU/day, targeting a 25(OH)D level >80nmol/L in addition to PEG-IFN/RBV combination therapy, had higher rates of rapid virologic response (RVR; 44% vs. 17%, p<0.001), complete early virologic response (cEVR; 94% vs. 48%, p<0.001) and SVR (86% vs. 42%; OR 2.5, 95% CI 2.0-4.9, p<0.001) in HCV-1 [36] and SVR (95% vs. 77%, p<0.001) in HCV-2/3 infection [37] compared to subjects treated with standard therapy. Moreover, recipients of vitamin D3 supplementation were less likely to be relapsers or non-responders to antiviral therapy, and had improved insulin resistance indices [36]. Similarly, a small retrospective Italian study showed vitamin D3 supplementation improved SVR rate in the treatment of recurrent hepatitis C post liver transplantation (53.3% vs. 18.5%, p=0.02) [38]. It remains unclear whether these improvements in the clearance of HCV with vitamin D supplementation are the result of an alteration in innate and/or adaptive immune function, or are mediated via improvement in insulin resistance. Large, prospective, placebo-controlled studies are thus required to assess the impact of vitamin D supplementation on viral response in CHC treatment. However, these studies now seem unlikely to occur in the new and rapidly evolving era of direct acting viral therapy.

Vitamin D status also reportedly correlates with liver histology in CHC. Patients with vitamin D deficiency have a higher grade of hepatic necroinflammation [28], [33], more advanced fibrosis stage [28], [29], [34] and may possibly have more rapid fibrosis progression [39]. At a cellular level, vitamin D deficiency is associated with downregulation of the 25-hydroxylase enzyme CYP27A1 in liver tissue. This may have pathogenetic relevance, given the established inverse relationship between CYP27A1 expression and the severity of necroinflammatory activity [28].

The above findings highlight the potential role that proteins and enzymes involved in the synthesis and metabolism of vitamin D may have in liver inflammation and response to anti-viral therapy. Genetic variation in the rs10877012 A/C polymorphism in the promoter region of the 1α-hydroxylase enzyme CYP27B1, but not the rs10735810 FokI VDR polymorphism, is associated with SVR in HCV-1 infection. Subjects with the AA genotype have higher SVR rate and 1α,25(OH)2D level than those with the AC or CC genotype [29], suggesting a key role of vitamin D in CHC infection. Moreover, a recently published proteomic study has shown vitamin DBP to be one of three metaproteins associated with SVR [40]. DBP levels are significantly lower in subjects with significant or advanced fibrosis (METAVIR F2-4) compared with those with absent or minimal fibrosis (F0/1) and healthy controls [41], [42].

Thus, vitamin D deficiency appears to be common in CHC and may be associated with adverse outcomes such as lower treatment response, more advanced fibrosis stage and increased severity of necroinflammation. It remains, however, uncertain as to whether vitamin D supplementation improves the SVR rate in patients receiving combination anti-viral therapy with PEG-IFN and RBV. Still, the findings of a significant association between the CYP27B1 rs10877012 A/C polymorphism, higher 1α,25(OH)2D levels, and SVR rate, as well as the association between vitamin D-binding protein and SVR suggest that higher 25(OH)D and 1α,25(OH)2D levels directly improve the virologic response to PEG-IFN and RBV therapy, presumably by impacting on the downstream regulation of vitamin D target gene transcription. DBP determines how much free 25(OH)D substrate is available for 1α-hydroxylase as well as the amount of free 1α,25(OH)2D ligand available to activate the VDR and influences downstream gene transcription. Hepatic 1α-hydroxylase activity levels therefore represent a major additional factor regulating 1α,25(OH)2D concentration in the liver.

Vitamin D and non-alcoholic fatty liver disease

Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome. It is the most common liver disease in the developed world, with a prevalence of 20–30% [43]. Thirty percent of subjects with NAFLD have histologic evidence of non-alcoholic steatohepatitis (NASH) [44] and are at risk of disease progression and development of cirrhosis. The pathogenesis of NAFLD is yet to be fully elucidated, but insulin resistance (IR) is implicated as the key mechanism leading to hepatic steatosis. Apart from lifestyle modification that results in significant weight loss [45], there is currently no safe, effective therapy for NASH.

Vitamin D levels decrease by 1.3nmol/L with each 1kg/m2 increase in body mass index (BMI) [46]. Normal vitamin D status is associated with a two-thirds lower prevalence of metabolic syndrome compared to those with reduced levels [47]. In non-diabetic Caucasians low vitamin D levels are independently associated with insulin resistance [48] and are a predictor of increased 10-year risk of developing hyperglycemia and insulin resistance [49]. A vitamin D response element is present in the insulin gene promoter region, and 1α,25(OH)2D activates transcription of the insulin gene [50]. Both 1α-hydroxylase and the vitamin D receptor are expressed on pancreatic β cells, with an association between low vitamin D levels and impaired β cell function having been suggested [50], [51]. Two randomized placebo-controlled trials have shown that high dose vitamin D supplementation improved insulin sensitivity in non-diabetic South Asians [52], [53]. A large prospective cohort study of women demonstrated that those who received vitamin D supplementation had a significantly lower risk of developing type 2 diabetes [54].

Subjects with NAFLD have lower vitamin D levels when compared with controls. Low vitamin D levels are closely associated with histologic severity of steatosis, necroinflammation, and fibrosis in NAFLD, independent of age, gender, BMI, Homeostatic Model Assessment (HOMA)-IR score and presence of metabolic syndrome [55], [56]. These findings have been confirmed in children with NAFLD [57].

In a recent study of Lewis rats with diet-induced (choline-deficient and iron-supplemented l-amino acid or CDAA) NASH, phototherapy elevated 25(OH)D, and 1α,25(OH)2D levels while reducing hepatocyte inflammation, fibrosis, and apoptosis when compared to controls. Phototherapy also improved insulin resistance and increased serum adiponectin in association with reduced hepatic expression of the profibrotic transforming growth factor (TGF)-β and α-smooth muscle actin (α-SMA), a marker of hepatic stellate cell activation. In addition, oral vitamin D3 supplementation reportedly improved liver histology in a dose-dependent manner [58]. Furthermore, in a rodent high fat diet model of NAFLD, vitamin D deficiency exacerbated histologic features of NAFLD, increased insulin resistance, and upregulated liver tissue expression of genes involved in hepatic inflammation and oxidative stress [59]. Given the above findings, prospective studies that assess the impact of vitamin D supplementation on the histologic features of NASH are warranted as a priority, given the lack of an effective therapy for this condition.

Vitamin D and liver fibrosis

1α,25(OH)2D has anti-fibrotic effects in lung fibroblasts and mesenchymal multipotent cells in vitro [60], [61], as well as anti-proliferative and anti-fibrotic effects in both in vitro and in vivo rat models of liver fibrosis. VDR is expressed by hepatic stellate cells (HSC) and this expression is upregulated by 1α,25(OH)2D. In addition, 1α,25(OH)2D suppresses HSC proliferation, and expression of cyclin D1, tissue inhibitor of metalloproteinase 1 and collagen Iα1 in vitro. In vivo, 1α,25(OH)2D decreases α-SMA expression and collagen levels, and prevents the development of cirrhosis by thioacetamide (TAA) [62], [63]. A vitamin D level >50nmol/L may be associated with a decreased frequency of rapid fibrosis progression in CHC [39]. However, the clinical importance of vitamin D as an anti-fibrotic agent remains to be determined.

Vitamin D receptor polymorphisms and liver disease

The vitamin D receptor (VDR) gene is located on chromosome 12. It encodes a 48kDa soluble protein that is a member of the nuclear receptor family of ligand-activated transcription factors. Common single nucleotide polymorphisms (SNP) of the VDR gene include FokI (rs10735810), BsmI (rs1544410), ApaI (rs7975232), and TaqI (rs731236). There is a marked racial variation in the allele frequency of these VDR polymorphisms [64], but their influence on VDR function and signaling is unknown [65]. The BsmI, ApaI, and TaqI SNPs are all in the 3′ region of the VDR gene and are in linkage disequilibrium with each other [66].

In CHC infection, the bAt [CCA]-haplotype of the BsmI, ApaI, and TaqI alleles, and the CC genotype of the ApaI allele are associated with rapid fibrosis progression, cirrhosis and increased intrahepatic expression of the fibrosis marker gene MMP-9 [39]. In chronic hepatitis B (HBV) infection, the variation in allele frequency of BsmI, ApaI, and TaqI is associated with HBeAg positivity and HBV flare [67]. Variation in ApaI, and to a lesser extent TaqI, is also associated with a higher HBV viral load and more severe fibrosis and necroinflammation [68]. Variation in the TaqI VDR polymorphism is also associated with both chronic HBV infection [69] and occult HBV infection [70], in which there is a low degree of HBV replication present in HBsAg negative subjects.

In hepatocellular carcinoma (HCC), complicating cirrhosis variation in the allele frequency of the BsmI, ApaI, and TaqI, but not FokI VDR polymorphisms is associated with HCC development when compared to cirrhotic patients without HCC. This association is most marked in subjects with alcohol-related cirrhosis, where carriage of the BsmIApaITaqI A–T–C and G–T–T haplotypes is independently associated with an increased risk of HCC. Furthermore, there is a significant difference in allele frequency of these VDR polymorphisms in alcohol-related cirrhosis compared to cirrhosis complicating chronic viral hepatitis [66].

Multiple studies have confirmed an association between VDR polymorphisms and autoimmune liver disease in both European and Asian populations. Variation in the allele frequency of the BsmI polymorphism is associated with primary biliary cirrhosis [71], [72], while variation of the FokI polymorphism is associated with autoimmune hepatitis [64], [73]. Furthermore, carriage of the VDR BsmITaqI G–T/G–T diplotype is an independent predictor of acute cellular rejection post-liver transplantation [74]. Similarly, VDR polymorphisms are associated with a variety of other autoimmune and immune-mediated diseases, including type 1 diabetes [75], leprosy [76], Crohn’s disease [77], tuberculosis [69], [78], psoriasis [79], multiple sclerosis [80], and Graves’ disease [81] (Table 4).

Table 4. Genetic variation in vitamin D and disease.

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Vitamin D, the immune system, and the liver

There is an increased incidence and prevalence of autoimmune diseases such as type I diabetes, multiple sclerosis (MS) and Crohn’s disease in geographic regions at higher latitude [82], [83]. This phenomenon is suggested to be related to lower 25(OH)D levels resulting from decreased ultraviolet sunlight exposure. In support of this hypothesis, the incidence of MS decreases with increasing 25(OH)D levels [84] and vitamin D supplementation decreases the risk of developing both MS in women [85] and type 1 diabetes in children by 80% [86]. In this context, vitamin D has an important role in both the innate and adaptive immune system [87]. Macrophages, T cells, and DCs express both 1α-hydroxylase and vitamin D receptor, and are thus direct targets of 25(OH)D and 1,25(OH)2D [88], [89], [90].

Innate immunity

The innate immune response is mediated by pattern-recognition receptors (PRR). Toll-like receptors (TLR) are a family of transmembrane PRRs with broad specificity, expressed on immune cells such as polymorphonuclear cells, monocytes, and macrophages. They interact with pathogen-associated molecular patterns such as viral nucleic acids, and bacterial and fungal products, to trigger an inflammatory (TNF, IL-1β, and IL-6) or antimicrobial response in the host [91]. Several data from studies focusing on the immunology of mycobacterial infection suggest vitamin D and DBP play a significant part in the activation of the innate immune response. The risk of Mycobacterium tuberculosis (TB) infection is increased in subjects with vitamin D deficiency, with the greatest risk observed in subjects with the lowest 25(OH)D levels [78], [92], [93], [94]. At a cellular level, macrophages infected with M. tuberculosis initiate a TLR2/1 response that enhances 1α-hydroxylase and VDR expression and induction of the anti-microbial peptide cathelicidin. The anti-microbial activity of macrophages occurs via a vitamin D-dependent process. Addition of 1α,25(OH)2D to M. tuberculosis-infected macrophages reduces the number of viable bacilli, while both vitamin D and TLR2/1 are required for cathelicidin production [95]. DBP plays a key role in modulating monocyte responses to 25(OH)D and 1α,25(OH)2D, that varies according to the DBP genotype. Notably, monocytes cultured in serum from DBP null mice, and in human serum with the lower affinity Gc1S and Gc2 DBP isoforms, have more potent induction of cathelicidin than monocytes cultured in serum from DBP+/− mice or human serum with the high affinity Gc1F isoform [96]. The Gc2 isoform is associated with lower 25(OH)D levels [97], [98], and carriers of this allele have an increased susceptibility to active TB in the presence of severe vitamin D deficiency (<20nmol/L) [99]. The Gc2 isoform is less able to be converted into macrophage activating factor, resulting in reduced macrophage function [4]. These data indicate that vitamin D-dependent antimicrobial responses may be strongly influenced by genetic polymorphisms in DBP, especially in the presence of vitamin D deficiency. Moreover, adjunctive high-dose vitamin D significantly hastens the time to sputum culture conversion during intensive-phase antimicrobial treatment of pulmonary TB in the subset of patients with the tt genotype of the TaqI VDR polymorphism [100]. In addition, variation in allele frequency of the FokI and TaqI VDR polymorphisms in the presence of vitamin D deficiency is associated with increased risk of TB [69], [78].

The role of vitamin D in innate immunity has implications on liver disease. Chronic liver disease is characterized by ongoing increased exposure of the liver via the portal circulation to bacterial products such as lipopolysaccharide (LPS). Contributing factors include increased intestinal mucosal permeability, alcohol ingestion, and small bowel bacterial overgrowth [101], [102], [103]. Dietary factors, such as a high-fat diet that predisposes to NAFLD, may also contribute to increased intestinal permeability and result in increased hepatic exposure to LPS [104]. Kupffer cells, the resident macrophages of the liver, represent 80–90% of the macrophages in the body [105], and their innate immune vitamin D-dependent antimicrobial response is also likely to be influenced by the vitamin D status and genetic polymorphisms in DBP. They also express TLR2, TLR4, and TLR9, and are responsive to LPS, the main ligand of TLR4. Hepatocytes, hepatic stellate cells, sinusoidal epithelial cells, biliary epithelial cells, and hepatic DCs also express TLR4 and are responsive to LPS. The interaction of LPS with TLR4 in the liver is crucial during hepatic fibrogenesis [101], [106]. Serum vitamin D levels are inversely proportional to TLR2 and TLR4 expression in monocytes, with administration of 1α,25(OH)2D downregulating expression of TLR2, TLR4, and TLR9 [107], [108], [109], [110]. Intestinal microbiota play an essential role in hepatic fat accumulation. TLR2, TLR4, and TLR9 are implicated in the pathogenesis of NAFLD, with TLR4 and TLR9 signaling associated with worsening steatosis, inflammation and fibrosis [111], [112]. In obese rats, vitamin D deficiency increases hepatic mRNA levels of TLR2, TLR4, and TLR9, and the endotoxin receptor CD14, which is implicated in worsening histologic features of NAFLD [59]. In CHC infection, increasing hepatic necroinflammatory activity correlates with increasing hepatic mRNA expression of TLR2 and TLR4, and hepatic TNFα mRNA is also closely correlated with TLR2 and TLR4 mRNA expression [113]. Furthermore, the antiviral effect of vitamin D on hepatitis C inoculated HuH7.5 hepatoma cells is mediated by innate immune system activation of the interferon-mediated signaling pathways [30].

NK cells and DCs are both important innate immune effector cells. Studies in VDR knockout mice have shown that expression of VDR is necessary for NK cell development and function [114]. 1α,25(OH)2D enhances NK cell cytotoxicity [115] and suppresses DC maturation, inducing a more tolerant DC phenotype which, at the interface of the innate and adaptive immune systems, promotes T regulatory (Treg, CD4+CD25+) cell activity [116].

Adaptive immunity

Vitamin D is an important modulator of T cell response to pathogens, which is a key component of adaptive immunity. In particular, activation of naïve T cells is a vitamin D-dependent process. In the inactivated state, naïve T cells do not express VDR and express almost no phospholipase C-γ1 (PLCγ1), which is a key molecule required for subsequent classical T cell receptor signaling and T cell activation. Following stimulus exposure, VDR is expressed on T cells through T cell receptor signaling via the alternative mitogen-activated kinase p38 pathway. The VDR complex, activated by binding of 1α,25(OH)2D, upregulates transcription of the gene encoding PLCγ1 and results in a 75-fold increase in PLCγ1 expression, enabling activation of naïve T cells. T cells in patients with lower 25(OH)D and 1α,25(OH)2D levels have a lower proliferation index after stimulation than T cells from patients with normal 25(OH)D and 1α,25(OH)2D levels; this pattern is overcome by exogenous administration of 1α,25(OH)2D [117].

1α,25(OH)2D also has an anti-proliferative effect on adaptive immunity. It inhibits proliferation of T helper type 1 (Th1) lymphocytes, which produce interferon (IFN)-γ, interleukin (IL)-2, and activate macrophages [118], and shifts the balance to a T helper type 2 (Th2) phenotype with increased production of IL-4, IL-5, and IL-10 [119]. 1α,25(OH)2D increases Treg cells [120], [121], enhances DC secretion of IL-10, decreases DC secretion of IL-12, a critical cytokine in Th1 development [122], and inhibits Th17 development via inhibition of IL-6 and IL-23 production [121]. In patients with MS, 25(OH)D, but not 1α,25(OH)2D, levels correlate with the ability of Treg cells to suppress the proliferation of activated T responder cells and inversely correlated with Th1/Th2 ratio [123]. IL-2, IL-10, and IL-12 genes in T cells have regions which bind to VDR and 1α,25(OH)2D may directly play a role in the transcription of these cytokines in T cells [124]. The ability of 1α,25(OH)2D to modulate the adaptive immune system may explain the association of vitamin D supplementation and higher 25(OH)D levels with a lower risk of multiple autoimmune diseases.

In orthotopic liver transplant recipients, severe 25(OH)D deficiency (<12.5nmol/L) and VDR BsmITaqI G–T/G–T diplotype are independent predictors of moderate-severe acute cellular rejection, whilst vitamin D3 supplementation decreases the risk of acute rejection by 60% [74], [125]. These findings highlight the importance of optimizing the vitamin D status in liver transplant recipients, not only to prevent bone loss, but also to reduce the risk of T cell-mediated acute rejection. A lower Th1/Th2 ratio is an independent predictor of SVR in treatment of HCV-1 [126], which possibly explains why vitamin D supplementation may improve therapeutic outcomes with PEG-IFN plus RBV. The immune tolerant phenotype promoted by vitamin D may also be of therapeutic benefit in NASH, where activation of both innate and adaptive immunity is implicated in its pathogenesis.

Genome wide association studies of vitamin D

Only about a quarter of vitamin D variability between individuals is explained by factors such as reported dietary intake, latitude and season of measurement [127], [128]. Twin and family studies suggest that genetic factors play a significant role in the wide variation of vitamin D levels observed within and between populations [129]. Polymorphisms of the hydroxylases, DBP, and VDR may have a profound influence on serum vitamin D levels and the efficacy of vitamin D as a hormone. Two large genome wide association studies (GWAS), involving patients of European ancestry [130], [131], of SNPs and their association with 25(OH)D levels have revealed important information about genetic variation in the enzymes and carrier proteins which are integral to the synthesis and metabolism of vitamin D.

The NADSYN1/DHCR7 locus is closely related to the de novo synthesis of vitamin D3 in the skin from the precursor 7-dehydrocholesterol. There is an association between 25(OH)D levels and several SNPs including rs12785878, rs12800438, rs3794060, rs4945008, and rs4944957 of this locus [124]. SNPs in the 25-hydroxylase CYP2R1 locus rs10741657, rs2060793, rs12794714, rs10500804, and rs7116978 are also significantly associated with 25(OH)D levels [128], [129].

The highly polymorphic vitamin D-binding protein binds the majority of 25(OH)D and 1α,25(OH)2D. DBP is predominantly produced in the liver, but also in kidney, gonads, fat, and neutrophils. SNPs in the DBP locus associated with 25(OH)D levels are rs2282679, rs7041, rs3755967, rs17467825, rs2298850, and rs1155563 [124], [125], [126]. Response to vitamin D supplementation may vary with differing genotypes of DBP [132], [133]. In addition to the three common isoforms Gc1F, Gc1S, and Gc2, there are >120 rare variants of DBP. Haplotypes of the SNPs rs4588 and rs7041 in exon 11 of the gene result in the Gc1F, Gc1S, and Gc2 isoforms, which have differing affinities for vitamin D [134]. The DBP SNP rs2282679, which has the strongest association with vitamin D levels, lies in intron 12 near the actin subdomain III and may affect DBP binding of 25(OH)D [131].

24-hydroxylase (CYP24A1) is primarily responsible for the inactivation of 25(OH)D and 1α,25(OH)2D. The SNP rs6013897 from this locus is also associated with vitamin D levels [130].

These studies highlight the importance of genetic variation in vitamin D status and may explain in part the varying response seen to vitamin D supplementation. Polymorphisms in four specific loci involved in vitamin D synthesis and metabolism have a significant impact on circulating 25(OH)D levels in patients of European ancestry. Further GWAS that include patients of more diverse racial backgrounds may reveal more genetic associations with the vitamin D status.

Vitamin D and cancer

Vitamin D is also associated with the development of neoplasia. Higher 25(OH)D levels are associated with a lower risk of incident left-sided colorectal adenomas [135]. Multiple meta-analyses, and large prospective and retrospective observational studies have established that vitamin D deficiency is associated with an increased risk of colon [136], [137], [138], [139], breast [140], [141], [142], and prostate cancer [143], [144]. Furthermore, increased sunlight exposure is associated with a reduced risk of non-Hodgkin’s lymphoma [145] and VDR polymorphisms are associated with adenocarcinoma in the colon [146], ovary [147], breast, prostate, renal cell carcinoma, and melanoma [148]. With respect to the liver, a variety of VDR polymorphisms are associated with the development of HCC in at-risk patients as detailed above. However, it remains unclear as to whether vitamin D deficiency is associated with an increased risk of HCC.

Conclusions

Vitamin D deficiency is a common problem in chronic liver disease and is closely associated with disease severity. The anti-inflammatory and immune-modulatory properties of vitamin D provide plausible mechanisms by which vitamin D may impact on disease progression and severity, especially in CHC and NASH. However, there are few prospective studies evaluating the effect of vitamin D supplementation in chronic liver disease and these are clearly warranted in the areas of NASH and CHC based on preclinical, and limited retrospective and prospective clinical data. Genetic polymorphisms of the vitamin D receptor and of proteins and enzymes involved in vitamin D synthesis and activation have an association with vitamin D status and severity of liver disease. Further studies are also warranted in this area, to confirm known associations and evaluate other genetic polymorphisms, especially in the vitamin D binding protein, which plays a key role in vitamin D synthesis, activity and bioavailability (Table 5). In the interim period, we recommend vitamin D status to be assessed in all patients with CLD and, if deficiency is present (<50nmol/L or 20ng/ml), supplementation with 1000–4000IU/day of vitamin D3 should be initiated, with the initial dose dependent upon baseline 25(OH)D levels. However, increasing evidence suggests that supplementation should be considered for a 25(OH)D level <75nmol/L, especially in those considering interferon-based antiviral therapy for CHC. Further prospective studies are required to identify an optimal 25(OH)D level in subjects with CLD.

PIIS0168827812003601_fx6_lrg

Table 5. Key future research requirements.

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Journal of Hepatology
Volume 57, Issue 4 , Pages 715-717, October 2012

Harvey J. Alter

Department of Transfusion Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health (NIH), Bethesda, MD, USA

Received 22 June 2012; accepted 28 June 2012. published online 04 July 2012.

See Article, pages 730–735

To have B or not to have B, that is the question?

Whether tis nobler in the mind to suffer

The slings and arrows of contagious fortune

Or to vaccinate against a sea of troubles

And by opposing end them…

Liberally adapted from William Shakespeare

Development of a sterilizing vaccine is the ultimate strategy in the prevention of infectious diseases; treatment of established disease pales by comparison in its relevance to global health. Classic examples of vaccination-related disease eradication are smallpox and polio. Hepatitis B could follow a similar global trajectory given sufficient resources and the will of national health policies. The study of Yin-Hsian Ni, under the leadership of Ding Shen Chen, in this issue of the Journal of Hepatology chronicles how far we have come in the battle against hepatitis B. In 1984, when a universal vaccination program was initiated in Taiwan, 10% of the population was hepatitis B surface antigen (HBsAg) positive whereas the current rate among 3332 subjects in the Ni et al. study is 0.9%. Similarly, the prevalence of anti-core antibody (anti-HBc) has declined from 28% to 7% while the prevalence of protective surface antibody (anti-HBs) has risen from 24% to 56%. Serial 5-year interval analyses have shown that these trends are progressive and it is anticipated that as fully vaccinated children age and continue to dilute the HBsAg carrier population, Taiwan will near-eradicate hepatitis B infection and serve as a world model of what can be achieved when there is a national will to face, manage and then prevent seemingly insurmountable health issues.

The achievements in Taiwan are all the more remarkable given that only 50years ago, hepatitis B was a disease by name alone. There were no diagnostic assays, no observed particles, no known sequences, no culture methods, no treatments and no effective prevention strategies. This void began to fill in the early ′60s with the serendipitous finding of a precipitin line in agar that resulted from the interaction between the serum of a multiply transfused hemophiliac and that of an Australian aborigine. The aboriginal serum was part of a random panel that was being tested for lipoprotein polymorphisms in the NIH laboratory of Baruch Blumberg [1]. Early epidemiologic studies at NIH revealed a strong and unexplained association of the “Australia antigen” with leukemia. Subsequently, Blumberg moved to the Fox Chase Cancer Center in Philadelphia where on the hypothesis that the Australia antigen was genetically determined and had an association with leukemia, he and Tom London studied patients with Down’s syndrome who were known to have an inherited predisposition to leukemia. An overall prevalence of 10% was found among Down’s patients, initially supporting the genetic hypothesis [2]. However, subsequent studies of institutionalized versus non-institutionalized Down’s patients showed that the association was not with the disease, but rather with conditions of crowding and poor sanitation, providing the first clue to the infectious origin of this aboriginal antigen. The subsequent observation of antigen seroconversion in two Down’s syndrome patients and a technologist in the Blumberg lab, coincident with the onset of hepatitis, provided the definitive link between an unexpected precipitin reaction and human disease and established the first hepatitis specific serologic assay [3]. Fred Prince further defined the association showing that it was specific for serum and not infectious hepatitis [4]. Once a viral marker was in place, hepatitis research became goal directed and accelerated rapidly. Electron microscopy performed by Bayer and co-workers [5] demonstrated an abundance of spherical and tubular particles in Australia antigen positive specimens. Subsequently, Dane in England [6] showed by immune-electron microscopy that the spherical and tubular particles resided in complexes with larger enveloped particles that proved to be the complete hepatitis B virion and became known as the Dane particle. The antigen then was renamed Hepatitis B Surface Antigen (HBsAg). Meanwhile, Gerin and Purcell [7] used newly developed rate zonal ultracentrifugation to purify hepatitis B associated particles and to sort the infectious Dane particles from non-infectious spheres and tubules. These physical separations and chimpanzee infectivity studies [8] set the stage for subsequent commercial vaccine development by demonstrating that subunits of the virus were immunogenic, but not infectious. By 1976, only 9years after the first report that the Australia antigen was associated with hepatitis, and only 13years after the “aboriginal antigen” had set the chain in motion, Maurice Hilleman and co-workers at Merck manufactured a subunit, plasma-derived vaccine ready for clinical trial. The clinical trial, conducted by the late Wolf Szmuness and Cladd Stevens, was a classic in design and implementation. It was a placebo-controlled efficacy trial in high-risk male homosexuals and provided unequivocal documentation of vaccine efficacy [9]. Indeed, among the 95% who responded to a full course of vaccine, efficacy was 100%. Hence, by 1982, there was a licensed hepatitis B vaccine with the potential to eradicate this disease throughout the world. However, the vaccine was too expensive for developing nations where the risks of hepatitis B were, and are, exceedingly high.

Taiwan became a focal point for large scale epidemiologic studies of HBV prevalence, disease associations and prevention. Palmer Beasley and Lu-Yu Hwang conducted a massive prospective study of the incidence of hepatocellular carcinoma (HCC) among HBsAg positive and HBsAg negative subjects [10]. By following over 19,000 individuals, they showed that almost every case of cirrhosis and HCC occurred in the HBsAg positive group and that the relative risk of HCC was more than 200-fold greater in HBV-infected individuals than in uninfected controls. Definitive epidemiologic links between HBV and HCC were also demonstrated in South Africa by Michael Kew [11]. This unequivocal association also suggested that the hepatitis B vaccine would prevent HBV-related HCC and hence represent the first cancer vaccine. In a second series of studies, the Beasley team showed that the offspring of HBsAg and HBeAg positive mothers had a 90% chance of becoming chronic HBV carriers and that this risk could be reduced to 5% when hepatitis B immune globulin (HBIG) and hepatitis B vaccine were administered sequentially immediately after birth [12]. This set the stage for the administration of an HBIG-vaccine combination to children of known HBsAg positive mothers and ultimately to universal HBV vaccination of all newborns in Taiwan. The sequential 5-year analyses of universal HBV vaccination that was initiated in Taiwan in 1984 and is reported herein by Ni et al., documents the predicted efficacy of universal vaccination in the prevention of hepatitis B infection and its sequelae. The HBV/HBsAg carrier rate in Taiwan has decreased ten-fold to less than 1%, not only protecting the vaccine recipients, but also preventing a huge number of secondary infections. Importantly, there is no trend to increased prevalence with age indicating the prolonged protective effect of vaccine even in the absence of booster inoculations. The rate of occult HBV infection, characterized by the presence of HBV DNA in the absence of HBsAg has declined from 0.8% to 0.1% (p=0.003) over this same study interval. Although not measured in this study, other studies have already documented declining rates of HCC in Taiwan since the inception of universal vaccination [13].

It has been known since the original HBV vaccine trails in Taiwan that there is a small, but finite proportion of HBV vaccine recipients who nonetheless show serologic or molecular evidence of HBV infection. Since the primary mode of HBV transmission in HBV endemic regions is from an HBsAg and HBeAg positive mother to her offspring, the most likely reason for vaccine failure is that the fetus was infected in utero and that the infection was well established before vaccine was administered; this has suggested a strategy wherein HBsAg, and especially HBeAg, positive mothers would be given antiviral agents during pregnancy. A second reason for vaccine failure could be a very high viral load that would overwhelm vaccine efficacy. Third, a given individual might harbor a vaccine escape mutant. While this is a serious theoretical concern, there is little scientific evidence that vaccine escape mutants have entered the general population. The emergence of such mutants would presage that the vaccine would become less effective over time, but there is no evidence for loss of efficacy over the 25year duration of the Taiwanese study.

What then are the impediments to the global eradication of hepatitis B infection? These impediments do not reside in the scientific arena. Indeed, the science of recombinant HBV vaccine development is very advanced and it is probable that future formulations will provide only marginal increases in efficacy, though they might provide benefit through ease of administration and less frequent dosing. Further, the universal vaccination program in Taiwan has not only established vaccine efficacy, but has provided proof of principle that HBV vaccine can be effectively administered to virtually all neonates in a given population, can markedly diminish the HBV carrier rate and can break the treacherous cycle of maternal–fetal transmission that perpetuates the carrier state and recycles virus into the next generation. The concomitant of such vaccine efficacy is the prevention of chronic hepatitis, cirrhosis and HCC, all of which have been documented. With the Taiwan model in place, there is new impetus to attack the final hurdles to a global vaccination strategy, namely financial resources and the will of impoverished nations to make this a national health priority. More progress has been made in this socio-economic arena than would have seemed imaginable only a decade ago. In a unique and truly remarkable collaboration between the World Health Organization (WHO), UNICEF, the World Bank, the Bill and Melinda Gates Foundation, donor governments, developing countries, international development and finance organizations and the pharmaceutical industry, the Global Alliance for Vaccines and Immunization (GAVI) was founded in 1999. Statistics from WHO/GAVI indicate that each year 1.7 million children in developing countries die from a vaccine-preventable disease; in dramatic and sobering mathematics, this represents one life every 20seconds. These deaths are from a variety of vaccine preventable diseases including tuberculosis, diphtheria, tetanus, pertussis, measles, polio, pneumococcal pneumonia, and rotavirus-induced diarrhea. Deaths from hepatitis B are less acute, but perhaps equally devastating over a lifetime. GAVI has thus far fostered immunization of 326 million children and has set a goal to immunize 243 million additional children from 2011 to 2015 predicting this will prevent 4 million future deaths. 65 countries have now introduced a pentavalent vaccine that protects against diphtheria, tetanus, pertussis, hepatitis B, and hemophilus influenza. Hence it is apparent that hepatitis B vaccination can be applied on a global scale and can achieve efficacy in developing countries that are willing to partner with GAVI to protect their populations from infant and long-term mortality. The complete eradication of hepatitis B will take several more generations but all the pieces are in place and the unimaginable is now the conceivable.

Conflict of interest

The author declared that he does not have anything to disclose regarding funding or conflict of interest with respect to this manuscript.

References

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