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The HCV Diagnostics Pipeline

July 2018
by Bryn Gay

Simpler Rapid, Point-of-Care Hepatitis C Diagnostics: Are We There Yet?1

To increase treatment rates and make a dent in the epidemic by 2030, countries need to scale up access to affordable, simple, rapid diagnostics, ensuring services reach people most at risk of HCV infection, including people who inject or use drugs, people using harm reduction services, people coinfected with HIV on ARVs, prisoners, and migrants. Furthermore, market forecasting estimates the need of screening tests for high-burden LMICs to reach the global elimination targets by 2020 to be 826 million tests, compared with the 178 million tests that are projected to be demanded for LMICs by 2022, highlighting the capacity and supply chain gaps that need to be addressed to increase HCV diagnoses. To help guide countries on in vitro diagnostics (IVDs), in May 2018, the WHO released the first edition of the Essential Diagnostics List, which includes 53 lab tests for infectious diseases, including four types of HCV tests.

Due to the efficacy of pangenotypic DAAs, such as sofosbuvir/velpatasvir, glecaprevir/pibrentasvir (G/P), and sofosbuvir/daclatasvir, genotype tests are no longer necessary. In this way, the diagnostic pipeline is moving towards simpler, rapid, decentralized point-of-care (POC) HCV tests. The ideal diagnostic product, as identified in the Target Product Profile, would use either HCV ribonucleic acid (RNA) or HCV core antigen (cAg) and confirm diagnosis in 20 minutes, cost less than USD 5 per test (including the reagent cost), enable a person to initiate a pangenotypic treatment immediately, then return for test of cure at 12 or 24 weeks (i.e., sustained virological response [SVR12 or SVR24]) after completion of antiviral treatment. Patients with cirrhosis or at risk of re-exposure could return for monitoring and retreatment, as needed. Yet this rapid, affordable, true POC diagnostics technology is still several years away. In the meantime, we can use existing diagnostics and monitor the international trials and demonstration studies underway for several simplified diagnostics strategies in different settings, particularly for hard-to-reach populations and in LMICs, that could diagnose, then treat-to-cure a person in two or more visits.

Advocacy points

For companies:

  • Develop and validate dried blood spot (DBS) protocols and submit them for in vitro diagnostics (IVD) stringent regulatory authority approval so that DBS is a manufacturer-recommended sample type.
  • Any oral-fluid professional use antibody test (e.g., OraQuick) should be prioritized and registered for self-testing.
  • Prioritize development of POC ribonucleic acid (RNA) test and POC HCV core antigen (cAg) with acceptable diagnostic accuracy (sensitivity/specificity), and promote operational research of these technologies as soon as possible to understand their advantages/disadvantages when used in routine settings.
  • Develop point-of-care device to enable non-invasive assessment of fibrosis (equivalent to AST to Platelet Ratio Index (APRI) or fibrosis-4 (FIB-4) index) and evaluate the performance of POC ALT, which could be used in community-based healthcare settings.
  • Minimize and ensure transparency of service/maintenance costs for instrument-based platforms to better inform procurement and health program costs in LMICs

For governments:

  • Utilize diagnostics to support surveillance studies to understand countries’ local epidemics; when feasible prioritize surveys across infectious diseases for efficient use of resources.
  • Promote the integration of HCV assays on polyvalent platforms with the goal of reducing operational costs and facilitating price negotiations with the manufacturer by bundling items that aid in bulk purchasing and ease distribution.
  • Standardize the in-country registration process for IVDs and establish fast-track procedures for quality-assured (i.e., stringent-regulatory approved) diagnostics products.
  • Establish national External Quality Assurance schemes for laboratories performing HCV screening and/or confirmatory testing (through external collaboration or internal development).
  • Ensure competition and diverse distributor options to bring down diagnostics prices in countries.
    • Minimize the layers of distribution; ensure transparency and accountability along the global and domestic supply chains to reduce costs.

For donors:

  • Include hepatitis C tests and reagents in diagnostics procurement.
  • Facilitate the integration of disease-specific programs by promoting the sharing of multiplexed platforms across different programs and offer cross-disease instead of vertical funding.

1 We thank the two anonymous reviewers whose comments and suggestions helped improve and clarify this section.

 

TABLE 1. HCV Diagnostics Pipeline: Products with New Published Data or Regulatory Updates since the 2017 Pipeline Report

Assay

Sample/
Setting

Company

Status (regulatory)

Price

ANTIBODY ASSAYS – SCREENING

Monalisa and Murex combination Ag/Ab assay

Dried serum spots
 

Lab

Bio-Rad and

Abbott, respectively

CE-IVD

TK

·       A 2017 poster presented data HCV Ag/Ab combination assays, which are intended as a screening test despite using antigen assay, and were used during a window period using HCV seroconversion panels (for GT 1a and GT 2b):

o   Screened HCV co-infection in dried serum spots on filter paper of 1683 newly diagnosed HIV-cases.

o   Murex detected HCV infection earlier than Monolisa: 9 days for GT 1a and 2 days for GT 2b.

o   Yet Murex had fluctuating and lower optical density values below the cut-off for higher dilutions, compared with Monolisa which showed steady increases in the optical density values for all dilutions in each panel.

o   219/1683 (12.9%) and 193/1683 (11.4%) dried serum spot samples were found positive in the Murex and Monolisa, respectively.

o   Further testing with ELISA reactive samples revealed 74 (44.3%) positive results for both Murex and Monolisa. By comparison, 10/167 (5.9%) of samples positive in the HCV RNA assay were only positive in the Monolisa, while 5/167 (2.9%) were positive in the Murex ELISA sample.

o   Monalisa provided more reliable results for the detection of HCV infections compared to Murex. However, Murex is able to detect lower dilution, and could be used for early detection.

RNA ASSAYS – CONFIRMATORY AND TEST OF CURE

Xpert® HCV RNA assay

Plasma, serum

 

POC or lab

Cepheid

CE-IVD

Ceiling price USD 17.10 for HCV; USD 12.45 per cartridge for 4M bundled tests/year

      Field evaluation of Xpert® HCV assay for RNA quantification in GT6 in Cambodia demonstrates high sensitivity of 100% (95% CI 99.2, 100) and specificity of 98.5% (95% CI 98.4, 99.9) compared with Roche Cobas Ampliprep-Cobas TaqMan® (reference method). Quantitative analysis showed a small mean difference between Xpert® and Roche assays: 0.01 log10 IU/ml for all genotypes and -0.07 log10 IU/ml for GT6 indicating absence of systematic bias.

      Xpert® has excellent performance, however, there are major operational constraints such as the need for sufficient laboratory infrastructure, may be an expensive platform with costly cartridges for smaller, community clinics, and its waste management requires high temperature incinerators, which can be enormous constraints for resource-limited settings.

      Bundling HIV (VL and EID), HCV and HPV tests can drop the cartridge prices (USD 12.35 for HCV; USD 11.35 for HPV; USD 11.98-13.35 for HIV) if the test volumes purchased increases beyond 4 million per year.

      A 2017 study of RNA and antigen detection for diagnosis of acute HCV among MSM on PrEP used serum samples with Xpert®. The HCV RNA tests were positive within a median of two months before the detection of antibodies and ALT elevation when patients were asymptomatic and had no increased ALT in the majority of cases.

      Xpert® tests could be appropriate for use in high-risk MSM for early diagnosis and acute HCV infection, in order to prevent transmission.

Xpert® HCV VL FS assay

100 µL, capillary blood, Fingerstick

 

Tertiary POC: harm reduction settings

Cepheid

Prototype, CE-IVD

Possible availability in 2018

TK

      The VL Fingerstick assay was a modified version of the HCV RNA assay.

      Xpert® HCV VL Fingerstick test for HCV RNA quantification demonstrates high sensitivity 100.0% (95% CI, 93.9%–100.0%) and specificity 100.0% (95% CI, 96.6%–100.0%), in less than one hour, among people attending drug user health and homelessness services.

      Potential as a screening tool for HCV RNA detection in high-prevalence settings, particularly in services for PWID.

      Major advance over Ab-based RDT tests.

Genedrive® HCV RNA

Plasma

 

POC

Genedrive

CE-IVD in EU in 2017

USD 5000 per device,

USD 30-40 per test

      In 2017, Genedrive showed 98.6% sensitivity (95% CI 96.9% to 99.5%) and 100% specificity (95% CI 99.3% to 100%).

      Two-step process with 90 minute turnaround time, possible for task-shifting, requires reliable electricity and connectivity which may not be available in some LMICs.

      Accuracy for all genotypes.

OMUNIS PUMA HCV Kit

Plasma 

 

Lab

Omunis

ND

USD 20-30 per test

·       A 2017 poster examined performance characteristics of a new commercial, generic  assay for HCV RNA viral load quantification in plasma, which could be further evaluated for resource-limited settings:

      There were patients with 2% GT 1a, 31% GT 1b, 14% GT 2a, and 53% GT 6.

      The study showed excellent 100% sensitivity and 100% specificity of the OMUNIS PUMA HCV RNA VL assay compared to Roche Cobas Ampliprep-Cobas TaqMan® (reference method). The Bland-Altman analysis shows significant correlation and good agreement between the two assays.

      However, there was under-quantification with the OMUNIS assay for samples with HCV RNA > 6 Log10  IU/mL.

Real Time HCV Viral Load

DBS

 

Lab

Abbott

ND

Global Fund price contingent on test volume/term commitment

      HCV plasma viral load using DBS.

      Study presented at CROI 2018 used 36 paired plasma and DBS samples; yet the study did not provide patient profiles and there is limited information other than conducted in resource-limited setting in India:

      A good correlation of r=0.97 between the PVL values obtained by the standard Abbott plasma and the DBS assay (r2 = 0.94, p<0.001). Mean difference in HCV RNA between plasma and DBS was 0.25 log10 IU/mL and 95% limit of agreement between -0.7321 and 1.23374.

      The study supported the use of DBS as an alternative to plasma for RNA and genotyping testing, particularly in resource-limited settings.

QIAGEN Symphony®

(QS-RGQ Kit version 2)

Plasma, serum

 

Lab

QIAGEN

ND

TK

      Detects HCV RNA using QIAsymphony® SP/AS and Rotor-Gene® Q instruments. Study presented at 2015 European Society for Clinical Virology conference detected all  27 strains/1-6 genotypes of HCV tested. Showed values of 14.7 (95% CI 10.7–25.94) IU/ml; no cross-reactivity when challenged with panel of 36 organisms suggesting 100% specificity.

      This assay could be a good option for monitoring the response to antiviral therapy in chronic HCV infection.

·       QS-RGQ Kit version 2 still under development; Qiagen Symphony currently, used with a reagent-rental formula for HIV VL in Malaysia.

Aptima HCV Quant

Plasma, serum

 

Lab

Hologic

CE-approved IVD; FDA approved

TK

      Quantitates patient’s HCV RNA to assess achievement of SVR/test of cure.

        Aptima HCV Quant compared to the High Pure system/Cobas TaqMan version 2 (HPS/CTM) as reference assay in 2016 paper:

o   The study collected 267 samples (13 serum and 254 plasma samples; fresh and frozen); patient age of ≥18 years of age, known HCV-positive status. One sample excluded for not being able to conduct repeat testing.

o   High agreement, detection rates between the two assays: ranging from 100% for replicates at 1,000 and 100 IU/ml HCV RNA to 98.9% (Aptima) and 96.7% (HPS/CTM) for replicates at 25 IU/ml.

o   Both assays detected all genotypes similarly (at concentrations of 1,000, 100, and 25 IU/ml), but quantitation of genotype 1b was limited for both assays at the nominal concentration of 25 IU/ml. HPS/CTM quantified 0 out of 30 replicates and Aptima quantified 4 out of 30.

o   The Aptima assay has the lowest LOD and LLOQ (5.1 and 10 IU/ml, respectively) of all the assays approved; sensitive, accurate, reproducible assay with performance equal to or above HPS/CTM.

CORE ANTIGEN – CONFIRMATORY

ARCHITECT

HCV cAg assay

Serum and DBS

 

Lab

Abbott

CE-IVD, Under WHO PQ review

USD 10 per test; USD 25-50 per test result

      Can perform liver chemistry tests in addition to immunoassays for HCV cAg or anti-HCV detection and quantification.

      One recent study evaluated HCVcAg in plasma and dried blood spot samples, using Roche Cobas Ampliprep-Cobas TaqMan® (reference method) to measure HCV RNA levels and ARCHITECT-i2000R Immunoassay Analyser to measure HCVcAg levels. When the HCV RNA > 70 3,000 IU/mL, the data demonstrated good sensitivity and specificity of HCVcAg in plasma samples. The level of HCVcAg quantified in plasma was higher than that in DBS. The study demonstrated high sensitivity and specificity of the ARCHITECT HCV Ag assay on paired plasma and DBS, and good correlation of HCVcAg to RNA on both sample types.

o  The assay for plasma samples had the sensitivity and specificity of 91.6% (95%CI, 84%, 96%) and 100% (95CI, 84%, 100%), respectively.

o  The assay for DBS samples had the sensitivity and specificity of 82.9% (95%CI, 74%, 90%) and 96.1% (95CI, 78%, 100%), respectively.

o  Amino acid mutations in the HCV core region may have unclear effects, but detection of HCV active infection using DBS may be a good screening tool for chronic infection. Further studies required.

      Study published in 2017 aimed to assess accuracy of serum and DBS HCV cAg testing among PWID at opioid substitution centers in Tanzania:

      DBS HCVcAg had modest sensitivity of 76.7% and high specificity of 97.3%, with an area under the receiver operating curve of 0.87 (95% CI 0.83–0.92).

      To identify patients using serum, HCV cAg showed higher sensitivity (99.1%) and specificity (94.4%).

      HCVcAg performance did not differ by HIV coinfection or HCV genotype (1a-4a).

GENOTYPING ASSAY

Genotyping assay

DBS

 

Lab

Abbott

ND

Global Fund price contingent on test volume/term commitment

      HCV genotype test using DBS.

      Study presented at 2018 CROI used 36 paired plasma and DBS samples; yet the study did not provide patient profiles and there is limited information other than conducted in resource-limited setting in India:

      A good correlation of r=0.97 between the PVL values obtained by the standard Abbott plasma and the DBS assay (r2 = 0.94, p<0.001). Mean difference in HCV RNA between plasma and DBS was 0.25 log10 IU/mL and 95% limit of agreement between -0.7321 and 1.23374.

      The study supported the use of DBS as an alternative to plasma for RNA and genotyping testing, particularly in resource-limited settings.

 

ABBREVIATIONS

Ab: Antibody

Ag: Antigen

ALT: Alanine aminotransferase

APRI: AST to Platelet Ratio Index

ARV: Antiretrovirals

cAg: Core antigen

CE: Conformité Européene/European Conformity

DAA: Direct-acting antivirals

DBS: Dried blood spot           

FIB-4: Fibrosis-4 index

FS: Fingerstick

GT: Genotype

HCV: Hepatitis C virus

HPS/CTM: High Pure system/Cobas TaqMan version 2

IU/mL: International unit per milliliter

IVD: In vitro diagnostics

LMICs: Low- and middle-income countries

MSM: Men who have sex with men

ND: No data

POC: Point-of-care

PQ: Prequalification

PrEP: Pre-exposure prophylaxis

PVL: Plasma viral load

PWID: People who use drugs

RDT: Rapid diagnostic test

RNA: Ribonucleic acid, or HCV RNA test

SVR: Sustained virological response

TK: To come

uL: unit of liquid volume equal to one millionth of a liter, or 1 mm3

WHO: World Health Organization