Controlled Therapy Interruption Improves Host Immune Control of Chronic Hepatitis B Virus Infection in HBeAg-Positive Patients.

Prolonged viral suppression with oral antiviral drugs allows partial immune reconstitution. Controlled therapy interruption (CTI), by leveraging secondary immune response, proposes further augmentation in chronic hepatitis B virus (HBV) infection. Transient treatment interruptions (TIs) at months 0, 1, and 3 during otherwise continuous oral antiviral therapy allow viremic bursts, simulating autovaccination. Four weekly injections of Hepatitis B Immunoglobulin are given before the second and third TI to simulate prime boosting, which specifically amplifies the immune response. Fourteen patients (10 males; four controls, four HBeAg positive, and six anti-HBe positive) aged 28-46 years were studied. The period between TI and reappearance of viremia, time to relapse (TTR) (weeks) estimated immune control. The other endpoints included reduction in serum HBsAg IU/mL and loss of HBeAg. TTR after the first TI was significantly shorter in HBeAg-positive patients, indicating low baseline immunity. TTR increased significantly after the second and subsequent TI in all four HBeAg-positive patients. One patient persistently lost HBeAg. Mean HBsAg levels fell significantly in three of four patients after the second TI. In contrast, in the anti-HBe-positive group, TTR was unchanged after all three TI. Furthermore, no significant changes in HBsAg levels were detected after the second or subsequent TIs. No significant differences in adverse events were noted between groups. HBeAg-positive patients have low baseline levels of host immune control against HBV. CTI consistently boosts this immunity. CTI did not influence immunity in anti-HBe-positive patients.

Hagen-Poiseuille's law: The link between cirrhosis, liver stiffness, portal hypertension and hepatic decompensation.

The onset of hepatic decompensation in cirrhosis heralds an accelerated downhill course with poor outcome. The sole predictor of this decompensation in cirrhosis is increased hepatic vein to portal vein gradient hepatic venous pressure gradient (HVPG). Surrogate markers of liver function or hepatic reserve appear to be less relevant. The hepatic sinusoids become less elastic and more rigid as liver fibrosis and cirrhosis progress. We propose that the Hagen-Poiseuille's law, which applies to rigid, but not elastic vessels, determines the pressure-flow characteristics in the sinusoids. In the rigid cirrhotic liver, HVPG rises dramatically with any change in net surface area or radius, r(4) of the vasculature that follows surgical resection. This review relates liver stiffness to the risk of decompensation in patients with cirrhosis. The liver has a unique dual blood supply comprising a low pressure portal vein and high pressure hepatic artery. We compare the complexity of autoregulation in the normal elastic liver with that in the rigid cirrhotic liver. Therapeutic modalities to reduce portal pressure may reduce the risk of hepatic decompensation and improve outcomes in cirrhosis.

Management of Hepatocellular Carcinoma in Cirrhotic Patients with Portal Hypertension: Relevance of Hagen-Poiseuille's Law.

BACKGROUND: Hepatic decompensation in cirrhosis heralds an accelerated course with poor survival. An increase in hepatic venous pressure gradient (HVPG), rather than surrogate tests of liver function, appears to be the sole predictor of decompensation after surgical resection. We propose that hepatic sinusoidal walls become less elastic as cirrhosis progresses. Decompensation signals the development of increased vessel wall rigidity. The pressure-flow characteristics then become subject to Hagen-Poiseuille's law, which applies only to rigid, cylindrical vessels. Thereafter, HVPG rises exponentially (by a factor inversely proportional to the fourth power of the net radius of functional sinusoidal vessels, 1/r(4), at any given hepatic blood flow rate. This review attempts to correlate liver stiffness, risk of decompensation and outcomes from hepatocellular carcinoma (HCC) in patients with cirrhosis. SUMMARY: We compare the complexity of autoregulation in the normal elastic liver, which has a unique dual blood supply, with that in the rigid cirrhotic liver. We also review, in the context of background liver cirrhosis, the management of HCC which is in essence, a solid mass of unorganized cells that exacerbates liver stiffness. We discuss the differential effects of various therapeutic modalities such as liver transplantation, loco-regional therapy and drugs on HCC outcomes, based on their effects on HVPG. KEY MESSAGES: Increased hepatic artery supply, or the hepatic artery buffer response, may be the only available method for autoregulation or maintenance of hepatic blood flow in the cirrhotic liver. In HCC, loco-regional therapies, including partial resection of the cirrhotic liver, can exacerbate portal hypertension by increasing blood flow within the remnant organ. We conclude that studies of HVPG reduction as part of HCC management may be beneficial and are warranted.

Minimising cold ischaemic time is essential in cardiac death donor-associated liver transplantation.

BACKGROUND: An important issue in the transplantation of livers procured from cardiac death donors (CDDs) concerns why some centres report equivalent outcomes and others report inferior outcomes in transplantations using CDD organs compared with standard criteria donor (SCD) organs. Resolving this discrepancy may increase the number of usable organs. OBJECTIVES: This study aimed to test whether differences in cold ischaemic time (CIT) are critical during CDD organ transplantation and whether such differences might explain the disparate outcomes. METHODS: Results of CDD liver transplants in our own centre were compared retrospectively with results in a matched cohort of SCD liver recipients. Endpoints of primary non-function (PNF) and ischaemic cholangiopathy (IC) were used because these outcomes are clearly associated with CDD organ use. RESULTS: In 22 CDD organ transplants, CIT was a strong predictor of PNF or IC (P = 0.021). Minimising CIT in CDD organ transplants produced outcomes similar to those in a matched SCD organ transplant cohort at our centre and in SCD organ transplant results nationally (1- and 3-year graft and patient survival rates: 90.9% and 73.3% vs. 77.6% and 69.2% in CDD and SCD grafts, respectively. A review of the published literature demonstrated that centres with higher CITs tend to have higher rates of PNF or IC (correlation coefficient: 0.41). CONCLUSIONS: These findings suggest that a targeted effort to minimise CIT might improve outcomes and allow the safer use of CDD organs.

Acute wrist and foot drop associated with hepatitis C virus related mixed cryoglobulinemia: rapid response to treatment with rituximab.

The nature of the B-cell subsets associated with chronic hepatitis C virus related type II mixed cryoglobulinemia (HCV-MC) is unclear. We report the case of a 64-year-male with acute onset wrist drop and foot drop, secondary to HCV-MC related mononeuritis multiplex, who was treated with rituximab, an anti-CD20(+) antibody directed against B cells. We monitored the frequency of B-cell subsets in peripheral blood before and after rituximab, and correlated B-cell subset changes with clinical response. Significant improvements in his wrist and foot drop, as well as his vasculitic rash, depression and erectile dysfunction were evident within six days of starting rituximab and have persisted several months after B-cell recovery. More than 95% of CD20(+) B cells had disappeared from peripheral blood within 1 week, returning to baseline by week 21. CD20(+)CXCR3(+) frequency at baseline was similar to that at week 21. CD20(+)CD5(+), the human equivalent of B1 B cells and CD20(+)IgM(+)IgD(+), naïve B cells were increased. By contrast, CD20(+)CD27(+) memory cell frequency was reduced. These data suggest that CD27(+) memory B cells, but not CD5(+) and IgM(+)IgD(+) B cells may play a role in the clinical manifestations of cryoglobulinemia.

The effect of controlled therapy interruption in chronic HCV infection: enhanced host immune response? A hypothesis.

BACKGROUND: We have hypothesized that prolonged viral suppression partially reverses immune tolerance in chronic hepatitis C virus infection. Brief periods of treatment interruption can then simulate 'auto-vaccination' and evoke powerful secondary host immune responses. OBJECTIVE: To determine the effect of controlled therapy interruption CTI on viral load in previous relapsers to interferon and ribavirin treatment. STUDY DESIGN: Virus is maintained at undetectable levels for 2-8 weeks with pegylated interferon and ribavirin and then briefly interrupted, restarting as soon as viremia returns (cycle 1). It is suppressed for at least a further 4 weeks, then briefly interrupted again (cycle 2). RESULTS: Viremia relapsed within 2-4 weeks (time to relapse TTR) after the first treatment interruption in all four patients in cycle 1. TTR increased sevenfold with the second treatment interruption in patient 1 and was followed by sustained virological response with cycle 3. In patient 2, TTR increased threefold after cycle 2 and subsequent cycles. Serum ALT and bilirubin rose significantly with treatment interruption during cycles 2 and 3, returning to baseline with treatment resumption. Serum bilirubin rose to 12.3mg/dl when two doses of pegylated interferon were missed during cycle 4. In patients 3 and 4, TTR was unchanged after three consecutive cycles. However, VL has remained more than 1 log below baseline for up to 18 months in both. CONCLUSIONS: These observations suggest that CTI exerts significant control of chronic hepatitis C viremia.

Naturally occurring dominant resistance mutations to hepatitis C virus protease and polymerase inhibitors in treatment-naïve patients.

UNLABELLED: Resistance mutations to hepatitis C virus (HCV) nonstructural protein 3 (NS3) protease inhibitors in <1% of the viral quasispecies may still allow >1000-fold viral load reductions upon treatment, consistent with their reported reduced replicative fitness in vitro. Recently, however, an R155K protease mutation was reported as the dominant quasispecies in a treatment-naïve individual, raising concerns about possible full drug resistance. To investigate the prevalence of dominant resistance mutations against specifically targeted antiviral therapy for HCV (STAT-C) in the population, we analyzed HCV genome sequences from 507 treatment-naïve patients infected with HCV genotype 1 from the United States, Germany, and Switzerland. Phylogenetic sequence analysis and viral load data were used to identify the possible spread of replication-competent, drug-resistant viral strains in the population and to infer the consequences of these mutations upon viral replication in vivo. Mutations described to confer resistance to the protease inhibitors Telaprevir, BILN2061, ITMN-191, SCH6 and Boceprevir; the NS5B polymerase inhibitor AG-021541; and to the NS4A antagonist ACH-806 were observed mostly as sporadic, unrelated cases, at frequencies between 0.3% and 2.8% in the population, including two patients with possible multidrug resistance. Collectively, however, 8.6% of the patients infected with genotype 1a and 1.4% of those infected with genotype 1b carried at least one dominant resistance mutation. Viral loads were high in the majority of these patients, suggesting that drug-resistant viral strains might achieve replication levels comparable to nonresistant viruses in vivo. CONCLUSION: Naturally occurring dominant STAT-C resistance mutations are common in treatment-naïve patients infected with HCV genotype 1. Their influence on treatment outcome should further be characterized to evaluate possible benefits of drug resistance testing for individual tailoring of drug combinations when treatment options are limited due to previous nonresponse to peginterferon and ribavirin.

Immune suppression leading to hepatitis C virus re-emergence after sustained virological response.

Sustained virological response SVR is defined as undetectable HCV RNA in plasma 6 months after therapy has been discontinued. Relapse or re-emergence of viremia after SVR is rare. We report two patients that relapsed when immune suppressive therapy was given within a few weeks of achieving SVR. Patient 1 received prednisone for bronchitis and patient 2 relapsed soon after immune suppression was started post renal transplantation. These data suggest that the early phase of SVR might be associated with incomplete protective immunity. They suggest that sterilizing immunity with complete elimination of virus is unlikely. The cases also caution against the use of immune suppressive therapy in the immediate aftermath of SVR.

Concomitant diffuse large B-cell lymphoma and hepatocellular carcinoma in chronic hepatitis C virus liver disease: a study of two cases.

Chronic hepatitis C virus HCV infection progresses through liver fibrosis and cirrhosis to hepatocellular carcinoma HCC. It appears to be causally related to B-cell non-Hodgkin's lymphoma since regression after antiviral therapy has been described. Two cases are described of non-Hodgkin's lymphoma and HCC arising simultaneously in two patients. The first patient did not have cirrhosis on liver biopsy. HCV had been undetectable in plasma following successful therapy with interferon and ribavirin treatment 7 years earlier. The second patient developed an aggressive form of hepatocellular carcinoma HCC within weeks of stopping treatment with interferon and ribavirin. Therapy had induced complete viral suppression for over 40 weeks. The two cases suggest that non-Hodgkin's lymphoma and HCC can develop in the absence of detectable hepatitis C viremia and argues for continued surveillance even after sustained virological response to treatment.

Clonal expansion of immunoglobulin M+CD27+ B cells in HCV-associated mixed cryoglobulinemia.

Hepatitis C virus (HCV) is associated with B-cell lymphoproliferative disorders such as mixed cryoglobulinemia (MC) and B-cell non-Hodgkin lymphoma (B-NHL). The pathogenesis of these disorders remains unclear, and it has been proposed that HCV drives the pro-liferation of B cells. Here we demonstrate that certain HCV(+)MC(+) subjects have clonal expansions of immunoglobulin M (IgM)(+)kappa(+)IgD(low/-)CD21(low)CD27(+) B cells. Using RT-PCR to amplify Ig from these singly sorted cells, we show that these predominantly rheumatoid factor-encoding V(H)1-69/J(H)4 and V(kappa)3-20 gene segment-restricted cells have low to moderate levels of somatic hypermutations. Ig sequence analysis suggests that antigen selection drives the generation of mutated clones. These findings lend further support to the notion that specific antigenic stimulation leads to B-cell proliferation in HCV MC and that chronic B-cell stimulation may set the stage for malignant transformation and the development of B-NHL. The finding that these hypermutated, marginal zone-like IgM(+)CD27(+) B cells are clonally expanded in certain subjects with MC offers insight into mechanisms of HCV-associated MC and B-cell malignancy. This study was registered at www.clinicaltrials.gov as NCT00219999.

Hepatitis C Virus-related Mixed Cryoglobulinemia: Pathogenesis, Clinica Manifestations, and New Therapies.

Within the past decade, it has been recognized that a majority of patients with essential mixed cryoglobulinemia (MC) are chronically infected with hepatitis C virus (HCV). Although the underlying mechanisms have not been fully elucidated, cryoglobulin formation is clearly linked to the attempt of the host to clear the significant quantities of virions generated daily by the chronic infection. This review summarizes the current understanding of the relationship between chronic HCV infection and the development of MC, and discusses the interaction between the immune system and HCV and how this interaction can lead to the development of lymphoproliferative disorders.

Effect of ribavirin and amantadine on early hepatitis C virus RNA rebound and clearance in serum during daily high-dose interferon.

The early rebound in serum HCV RNA during HCV dynamic studies with high-dose interferon may be due to de novo infection with interferon escape quasispecies. We simultaneously measured serum alanine aminotransferase (ALT) and HCV RNA at rapid intervals in chronic HCV liver disease patients during interferon therapy alone or in combination with ribavirin and amantadine. HCV RNA declined rapidly between 0 and 48 hr in all patients (phase 1). Ribavirin and amantadine significantly increased this phase 1 decline. In all four monotherapy patients with viral rebound, the increasing levels of HCV RNA were associated with a parallel increase in serum ALT, consistent with a hepatitis flare or de novo infection. By contrast, in the four monotherapy patients without viral rebound, and all eight patients receiving combination therapy, the slow progressive phase two decay was associated with declining serum ALT levels. Ribavirin or ribavirin and amantadine significantly and incrementally increased the phase two HCV RNA clearance. Dynamic sequencing in the HVR1 region in one rebound patient confirmed the potential for rapid evolutionary changes during interferon therapy. These preliminary data suggest that early viral rebound might be associated with de novo infection with interferon escape HCV variants, which in turn are attenuated by ribavirin and amantadine.

Digital image analysis of the distribution of proliferating cell nuclear antigen in hepatitis C virus-related chronic hepatitis, cirrhosis, and hepatocellular carcinoma.

Viral dynamic studies in chronic hepatitis C virus (HCV) infection indicate a significantly shortened survival of virus-infected cells. Since at the steady state of chronic viral infection, the rate of infected cell elimination equals new cell regeneration, this would imply a high rate of hepatocyte turnover in chronic HCV liver disease. We estimated the fraction of regenerating hepatocytes in liver biopsy sections in chronic HCV liver disease, cirrhosis, and hepatocellular carcinoma (HCC). We used antibodies to proliferating cell nuclear antigen (PCNA) to detect proliferating cell nuclei in liver biopsy specimen from controls and patients with chronic hepatitis, cirrhosis, and HCC. We also used bis-benzimide to label fluorescently all hepatocyte nuclei simultaneously. Using digital image analysis, we calculated the area occupied by PCNA-stained hepatocyte nuclei, as a fraction of the total area occupied by fluorescently labeled hepatocyte nuclei (labeling index; LI). Antibody staining was negligible in the control specimen. The mean +/- SE PCNA LI increased from 0.21 +/- 0.1 in chronic hepatitis to 0.63 +/- 0.15 in HCC. There was no significant difference between chronic hepatitis and cirrhosis. The fraction of cells undergoing regeneration is increased in chronic HCV liver disease, HCV-related cirrhosis, and HCC. Increased hepatocyte turnover could provide the link between chronic HCV liver disease and HCC.