Comparison of Qualitative (COBAS AMPLICOR HCV 2.0 versus VERSANT HCV RNA) and Quantitative (COBAS AMPLICOR HCV Monitor 2.0 versus VERSANT HCV RNA 3.0) Assays for Hepatitis C Virus (HCV) RNA Detection and Quantification: Impact on Diagnosis and Treatment of HCV Infections

Desombere, Isabelle; Vlierberghe, Hans Van; Couvent, Sibyl; Clinckspoor, Filip; Leroux‐Roels, Geert

doi:10.1128/jcm.43.6.2590-2597.2005

Cited by 37 publications

(31 citation statements)

References 45 publications

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“…In fact, in most of the currently used methods for HCV RNA detection, the total RNA is isolated from a maximum of 250 l of serum (8,10,11,13). By contrast, the use of ultracentrifugation permits the concentration of viral particles from large volumes of serum that would otherwise be difficult to manage for the direct isolation of total RNA, increasing the sensitivity of HCV RNA detection.…”

Section: Discussionmentioning

confidence: 99%

Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C

Bartolomé

López‐Alcorocho

Castillo

et al. 2007

J Virol

105

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Occult hepatitis C virus (HCV) infection of patients with abnormal liver function tests of unknown originwho are anti-HCV and serum HCV RNA negative but who have HCV RNA in the liver has been described. As HCV replicates in the liver cells of these patients, it could be that the amount of circulating viral particles is under the detection limit of the most sensitive techniques. To prove this hypothesis, serum samples from 106 patients with occult HCV infection were analyzed. Two milliliters of serum was ultracentrifuged over a 10% sucrose cushion for 17 h at 100,000 ؋ g av , where av means average, and HCV RNA detection was performed by strand-specific real-time PCR. Out of the 106 patients, 62 (58.5%) had detectable serum HCV RNA levels after ultracentrifugation, with a median load of 70.5 copies/ml (range, 18 to 192). Iodixanol density gradient studies revealed that HCV RNA was positive at densities of 1.03 to 1.04 and from 1.08 to 1.19 g/ml, which were very similar to those found in the sera of patients with classical chronic HCV infection. The viral genome of hepatitis C virus (HCV) is a 9.6-kb linear single-stranded RNA molecule of positive polarity that replicates via an HCV RNA strand of negative polarity (18).Chronic HCV infection is a progressive disease that may lead to liver cirrhosis and hepatocellular carcinoma (6, 21). The hallmark of chronic hepatitis C is the presence of anti-HCV and HCV RNA in serum for more than 6 months after acute infection. Recently, a new form of chronic HCV infection called "occult HCV infection," characterized by the presence of genomic HCV RNA in liver in the absence of anti-HCV and serum HCV RNA, in patients with abnormal liver function tests of unknown etiology was described (3). In 84% of patients with occult HCV infection, the negative-polarity (antigenomic) HCV RNA strand was also detected in liver, indicating that the virus was replicating. Furthermore, 70% of these patients had HCV RNA in peripheral blood mononuclear cells (PBMCs) (3), and viral replication in these cells was also reported (4).Since HCV replicates in the livers of patients with occult HCV infection, the reason why HCV RNA is not detected in serum is unknown. One possible explanation is that the number of circulating viral particles is too low to be detected even using the most sensitive reverse transcription (RT)-PCR techniques.In the present study, by using ultracentrifugation and a sensitive real-time PCR technique, we have demonstrated that sera from patients with occult HCV infection contain low amounts of viral particles and that the physical characteristics of these virions are similar to those found in patients with classical chronic HCV infection. MATERIALS AND METHODS Patients.One hundred six patients (74 males) with occult HCV infection were analyzed in this study. Patients had abnormal liver function tests, were anti-HCV negative (INNOTEST HCV Ab IV; Innogenetics, Ghent, Belgium), and did not have serum HCV RNA levels tested by real-time RT-PCR (see below). Other causes of liver disease...

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Section: Discussionmentioning

confidence: 99%

Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C

Bartolomé

López‐Alcorocho

Castillo

et al. 2007

J Virol

105

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“…Our testing with the TMA assay identified a subset of patients with persistent low-level viremia (PCR-negative but TMA-positive) on combination therapy who did not achieve SVR. Desombere et al 19 described a small number of treatment-naive patients who exhibited a similar pattern of treatment nonresponsiveness and who subsequently relapsed. Thus, within the context of the HALT-C study, consecutive TMA testing at weeks 20 and 24 could have been used in conjunction with EVR at W12 to avoid unnecessary therapy in an additional 45 patients, without missing any subjects who could have achieved SVR.…”

Section: Discussionmentioning

confidence: 99%

HCV RNA detection by TMA during the hepatitis C antiviral long-term treatment against cirrhosis (Halt-C) trial

et al. 2006

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For making treatment decisions related to chronic hepatitis C, the utility of HCV RNA tests with increased sensitivity has not been defined. Prior interferon nonresponders with advanced fibrosis (n ‫؍‬ 1,145) were retreated with peginterferon alpha-2a and ribavirin. Patients who were HCV RNA-negative by a polymerase chain reaction (PCR)-based assay (Roche COBAS Amplicor ™ HCV Test, v. 2.0; lower limit of detection [LOD] 100 IU/mL) at week 20 (W20) received treatment for 48 weeks. Stored specimens were tested using the Bayer VERSANT HCV RNA Qualitative (TMA) Assay (LOD 9.6 IU/mL) and compared to PCR results for the ability to predict sustained virological response (SVR; defined as undetectable HCV RNA by PCR at W72). Nearly all PCR-positive samples (1006/1007, 99.9%) were positive as assessed by TMA. Among 1,294 PCR-negative samples, 22% were TMApositive. Negative TMA results were more predictive of SVR than were negative PCR results at W12 (82% vs. 64%, P < .001) and at W20 (66% vs. 52%, P ‫؍‬ 0.001). SVR was more likely the earlier TMA had become negative during treatment (82% at W12, 44% at W20, 20% at W24). Among 45 patients who were TMA-positive but were PCR-negative at W20 and W24, none achieved SVR (95% CI: 0%-8%). Approximately 10% of patients with a Abbreviations: HCV, hepatitis C virus; SVR, sustained virological response. From the

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“…In this group 40 subjects were sustained responders to therapy, 40 were relapsers, and 40 were non-responders. Quantitative HCV-RNA data were available from these 120 patients (Table I) [Desombere et al, 2005]. ''Group 2'' consisted of 35 genotype 1-positive chronic HCV patients that were treated with peg-IFNa/ribavirin.…”

Section: Patient Selection and Sample Collectionmentioning

confidence: 99%

Serum levels of anti‐NS4a and anti‐NS5a predict treatment response of patients with chronic hepatitis C

Desombere

Vlierberghe

Weiland

et al. 2007

Journal of Medical Virology

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In order to understand better the clinical significance and prognostic value of antibody responses to HCV proteins and in search for parameters that may allow the early identification of non-sustained responders to therapy, antibody levels were measured against NS3, NS4a and NS5a at baseline in the serum of 120 patients chronically infected with HCV of genotype 1 that were classified as sustained responders, relapsers, or non-responders to therapy. The capacity of these antibody tests to predict therapy-outcome was evaluated. While no differences were observed in the anti-NS3 responses in these different response groups, anti-NS4a and anti-NS5a antibodies were observed more frequently and at higher titres in sustained responders versus non-responders or non-sustained responders (=non-responders + relapsers). Based on this observation, a combination of test results consisting of 'the absence of NS4a (AA 1687-1718) antibody at baseline and the presence of HCV-RNA exceeding 10(5) IU/ml after 1 week of treatment' was identified which predicts non-sustained response to treatment with 100% certainty. Replacing the HCV-RNA decision limit by a HCV-core antigen level of >15 pg/ml resulted in the same predictive value. The proposed algorithm also holds for patients treated with peg-interferon and ribavirin. In conclusion, in patients with chronic HCV infection, the decision to continue or stop treatment can be made after 1 week of treatment with (peg)-interferon alpha and ribavirin.

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Comparison of Qualitative (COBAS AMPLICOR HCV 2.0 versus VERSANT HCV RNA) and Quantitative (COBAS AMPLICOR HCV Monitor 2.0 versus VERSANT HCV RNA 3.0) Assays for Hepatitis C Virus (HCV) RNA Detection and Quantification: Impact on Diagnosis and Treatment of HCV Infections

Cited by 37 publications

References 45 publications

Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C

Ultracentrifugation of Serum Samples Allows Detection of Hepatitis C Virus RNA in Patients with Occult Hepatitis C

HCV RNA detection by TMA during the hepatitis C antiviral long-term treatment against cirrhosis (Halt-C) trial

Serum levels of anti‐NS4a and anti‐NS5a predict treatment response of patients with chronic hepatitis C

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