HCV: Diagnostic Testing and...

David E. Bernstein, MD, FACG

HCV: Diagnostic Testing and Staging

HCV: Diagnostic Tests

Necessary Not Necessary• HCV Antibody• HCV-RNA• Genotype

• ALT• IL28 B

ACG's Hepatitis School - Las Vegas, NV Copyright 2015 American College of Gastroenterology

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ALT as a Sole Trigger for Screening Misses Some Infected Patients

Patients* With Hepatitis C Infection

15%PersistentlyElevated ALT

42%PersistentlyNormal ALT

43%IntermittentlyElevated ALT

Elevated ALT

*4 serum ALT level measurements during 25 months of follow-up (n = 1042). These results are from a prospective community-based study that evaluated liver enzyme levels in patients with hepatitis C who had a history of prior drug use.

ALT = alanine aminotransferase.

Inglesby TV, et al. Hepatology. 1999;29:590-596.

Hepatitis C Antibody Testing

• 2 types of tests– Standard-goes to lab– Point of Care Testing


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Detection of Anti-HCV IgG:Rapid Tests

• Do not require complicated instrumentation/testing by skilled staff• Potentially generate results within an hour (potential point-of-care use)• CDC evaluation of 3 rapid tests (Orasure, Chembio,and Medmira)

– Assays based on recombinant antigens derived from core, NS3, NS4, and NS5 proteins in an immunochromatographic format

– Specificity: >99%; sensitivity: 86% to 99%• OraQuick HCV Rapid Antibody Test (CLIA-waived)

– FDA approved with fingerstick, whole-blood, and venous blood specimens from individuals aged >15 years and at risk for HCV infection or persons with signs and symptoms of hepatitis

– Not approved for general screening• Typically more expensive versus conventional immunoassays• Not designed for testing large batches of specimens• Results available in ~20 minutes

Kamili S, et al. Clin Infect Dis. 2012;55(suppl 1):S43-S48.

Steps for Fingerstick or Venipuncture Specimen Collection


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Detection of Anti-HCV IgG:Immunoassays

• Diagnostic specificity– >99% for 3rd generation assays

F l ti lt

Signal-to-Cutoff Ratios(FDA-Approved, Screening Assays)

(Detect Anti-HCV IgG)• False-negative results– Undergoing hemodialysis– Immunocompromised patients

• Signal-to-cutoff ratios – Predict a true antibody positive results

>95% of the time, regardless of the anti-HCV prevalence or characteristics of the population tested

(Detect Anti HCV IgG)

Ratio

Enzyme immunoassay (manual)Ortho HCV Version 3.0Abbott HCV EIA 3.0

>3.8>3.8

Chemiluminescence immunoassay (automated)

Vitros anti-HCVAdvia Centaur HCV

>8.0>11.0


Microparticle immunoassay (automated)

Architect anti-HCV >10.0

Chemiluminescence microparticle immunoassay (automated)

Architect anti-HCV >5.0

HCV Antibody

• Does not diagnose disease• Positive antibody may mean:

– Active disease– Previous infection now cleared– False positive

• All positive antibodies requires confirmatory• All positive antibodies requires confirmatory HCV-RNA testing


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Detection of Virus-Specific Molecules:HCV RNA and Genotype

• Nucleic acid testing for HCV RNA remains the gold standard

PCR b h d DNA i l

HCV RNA AssaysLLD/– PCR, branched DNA signal

amplification, and transcription-mediated amplification

– Specificity: up 99% across all genotypes 1-6

– Essential for monitoring response to therapy

Method (IU/mL)

QualitativeAmplicor HCV v2.0 (Roche)COBAS Amplicor HCV 2.0 (Roche)Ampliscreen (Roche)Versant HCV RNA (Gen-Probe)UltraQual HCV (National Genetics)Procleix HIV-1/HCV (Gen-Probe)

RT-PCR (manual)RT-PCR (semi-auto)RT-PCR (semi-auto)

TMA (manual)RT-PCR

TMA (manual)

5050

<501010

<50

QuantitativeAmplicor HCV Monitor (Roche)COBAS Amplicor HCV Monitor

(Roche)Versant HCV RNA 3.0 (Gen-Probe)

RT-PCR (manual)RT-PCR (semi-auto)

bDNA (semi-auto)

5050

615


( )COBAS Ampliprep/TaqMan

(Roche)Real Time HCV/m2000sp/m2000rt

(Abbott)HCV SuperQuant

(National Genetics)LCx HCV RNA-Quantitatie (Abbott)

( )qPCR (semi-auto)

qPCR (semi-auto)

RT-PCR (semi-auto)

RT-PCR (maual)

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12

20

25

General Clinical Statements on HCV Viral Load

• Amount of virus not clinically significant10 000 000 IU/ l b ild di i h i• 10,000,000 IU/ml may be mild disease or cirrhosis

• 50,000 IU/l may be mild disease or cirrhosis• Viral load not predictive of disease progression• Viral load predictive of maternal-fetal transmission• Viral load critical for assessing adherence to therapyg py


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Hepatitis C Genotype

• Important in determining which therapy to use

• Important in determining duration of therapy• Sub-typing important• Common US genotypes

1 1b 2 3 4– 1a, 1b, 2 ,3 ,4– Less common 5 and 6

Recommended Laboratory Testsfor Chronic HCV Infection

Test Purpose

Hepatitis C antibody by Screening for past or presentHepatitis C antibody byenzyme immunoassay (EIA)

Screening for past or present HCV infection

PCR for HCV RNA Confirmation of positive EIAGenotype Determines therapy and

duration

AASLD and IDSA.Available at: http://www.hcvguidelines.org/full-report-view. Version January 29, 2014.


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HCV Diagnostic Assays:What the Results Mean

Anti-HCV

HCV RNA Interpretationp

+ + Acute or chronic HCV depending on the clinical context

+ False positive HCV antibodyResolved infectionLow-level intermittent viremia

+ Early acute HCV infectionChronic HCV in setting of immunosuppressed stateFalse positive HCV RNA test

Absence of HCV infection

AASLD and IDSA.Available at: http://www.hcvguidelines.org/full-report-view. Version January 29, 2014.

Pre-Treatment Assessment

• HgB• Cr• Liver enzymes• HCV-RNA• Genotype• Assessment of disease severity

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Options for Liver Fibrosis Assessment:Degree of Fibrosis = Disease severity

Liver Biopsy FibroScanSerum Biomarkers

Serum Biomarkers assessing Fibrosis in Chronic HCV

– Fibrotest– Fibrosis Score 4

– HepascoreL k I d– Fibrosis Score 4

– AST to platelet ratio (APRI)

– FibroSpect II– Enhanced liver fibrosis

score (ELF)

– Lok Index– Virahep– Fibroindex– AST to ALT ratio– Platelet count

– Fibrosis probability index

Castera L. Gastroenterology. 2012;142:1293-1302.


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Serum FibroTest: A Continuous Variable (n=1,270)

Expected FibroTest

0.20

0.40

0.60

0.80

1.00

Fib

roTe

st F3-F40.73-0.74

F30.59-0.72

F20.49-0.58

F1-F20.32-0.48

F40.75-1.00

FibrosisFibroTest

Poynard, Clin Chem 2004; 50:1344-55.

0.000 1 2 3 4

Fibrosis StageF00.00-0.21

F0-F10.22-0.27

F10.28-0.31

Serum biomarkers

• Excellent at predicting minimal fibrosis• Excellent at predicting cirrhosis• May need 2-3 tests• Unclear if cost effective


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Liver Biopsy for assessing fibrosis

• “Gold standard”• Sampling error (< 1/50,000th of the liver)

– Cirrhosis missed in up to 20%– Grade of inflammation and stage of fibrosis

under-scored in shorter and narrower specimens

Adequate specimen:q p≥ 1.5 cm long (≥ 2 cm preferable; ≥ 2.5 cm best)

≥ 1.0 mm wide (≥ 1.4 mm preferable)≥ 6 portal triads (≥ 11 is better) RegevRegev A, et al. Am J A, et al. Am J GastroenterolGastroenterol 2002; 97:26142002; 97:2614--8.8.

Crawford AR, et al. Crawford AR, et al. HepatologyHepatology 1998; 28:3231998; 28:323--31.31.ColloredoColloredo G, et al. J G, et al. J HepatolHepatol 2003; 39:2392003; 39:239--44.44.

DemetrisDemetris AJ, AJ, RuppertRuppert K. J K. J HepatolHepatol 2003; 39:2752003; 39:275--7.7.BedossaBedossa P, et al. P, et al. HepatologyHepatology 2003; 38:14492003; 38:1449--57.57.

Malik AH, et al. Aliment Malik AH, et al. Aliment PharmacolPharmacol TherTher 2004; 19:5452004; 19:545--9.9.

Fibrosis Area: 65%

Sampling Error of Liver Biopsy

Fibrosis Area: 15%Courtesy of M. Pinzani, Florence


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Liver Biopsy: Potential Complications

• PainBl di• Bleeding

• Bile Peritonitis• Puncture of gallbladder, colon, kidney• Pneumothorax, hemothorax, pleural effusion• Arteriovenous fistulaArteriovenous fistula• Sepsis• Needle track seeding with tumor

Bravo AA, et al. N Bravo AA, et al. N EnglEngl J Med 2001; 344:495J Med 2001; 344:495--500.500.PiccininoPiccinino F, et al. J F, et al. J HepatolHepatol 1986; 2:1651986; 2:165--73.73.

McGill DB, et al. Gastroenterology 1990; 99:1396McGill DB, et al. Gastroenterology 1990; 99:1396--400.400.

Bleed following Liver Biopsy


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Summary : Liver Biopsy

• Gold standard• Poor patient acceptance• Safe with potential complications• Limited availability

Fibrosis = Stiffness• A key parameter of soft biological tissues

– Often related to a pathological state– Palpation widely used in routine clinical practice– Palpation widely used in routine clinical practice

10 9

10 10Bone

Stiff

ness

(Pa)

10 2

10 3

10 4

10 5

10 6

10 7

10 8 Cartilage

Contracted musclePalpable nodulesCirrhotic liverNormal breast tissues Relaxed muscleNormal liver

“When the liver is stiff, prognosis is bad”

Aphorisms, Hippocrate (460-370 BC)


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Transient Elastography measures stiffness

• Fibroscan®• MRI elastography

Fibroscan

•Examination time < 5 minutes•Median value of 10 successful acquisitions•Fasting 2-4 hours•Performed on expiration


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The FibroScan ProbeThe FibroScan Probe

Ultrasound(Shear wave speed measurement)

Actuator(Shear wave generator)

Skin

2.5 cm6.5 cm

Measurement PositionMeasurement Position

4 cm

1 cm 4 cm3

Explored Tissue


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Shear Wave PropagationShear Wave Propagation Map Map

Depth

TimeSpeed = Distance / Time

Summary Data DisplaySummary Data DisplayMedian shear wave speed, stiffness & IQR data


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FIBROSCAN

Transient elastography: What the Clinician needs to know

• What is the underlying disease• Other evidence of advanced liver disease• Factors that can affect the test

– Is the patient fasting?– BMI

B d f i fl ti (ALT l l)– Burden of inflammation (ALT level)– ETOH use– Cholestasis

Tapper et al. Clin Gastro Hep 2015;13:27-36


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kPa CutoffsDisease Cirrhosis Comments

Hepatitis C 12.5 kPa 7.3 kPa suggests significant fibrosis

Hepatitis B 11.7 kPa If normal ALT, consider treating at 9.0 kPa

NAFLD 10.3 kPa Consider performing CAP* (controlled attenuation parameter) assessment

ETOH 22.7 kPa if drinking12.5 kPa if abstinent

Biliary liver disease 17.9 kPa Must know alkaline phosphatase

Portal HTN 20 kPa suggests HVPG > 1050.7 kPa suggests high risk of varicealbleeding

* Assessed in decibels per meter, currently proprietary algorithm

Confounders of VCTE

Know the ALT

Confounders

Inflammation

Non-fasting

Ch l t i

Inexperience

Congestion

Fast 3-4 hours

Cholestasis

Alcohol Obesity

Know the alk phos

Use XL probe for BMI > 30Determine drinkingstatus

Examine for Right heartfailure


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Magnetic resonance elastography for detecting hepatic fibrosis Magnetic resonance elastography uses a vibrating device to induce shear waves in internal organs, which are

detected by a modified magnetic resonance imaging machine.

CAREY E , and CAREY W D Cleveland Clinic Journal of Medicine 2010;77:519-527

©2010 by Cleveland Clinic

Importance of Determining Degree of Fibrosis

• Determines prognosis• With new therapies, timing may be before,

during or after therapy– If possible, do not delay therapy

• May determine eligibility for treatment with anti-viral therapyanti-viral therapy


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Recommendations for when and in whom to initiate treatment

• Treatment is recommended for patients with chronic HCV pinfection.

• Rating: Class I, Level A• Treatment is assigned the highest priority for those

patients with advanced fibrosis (Metavir F3), those with compensated cirrhosis (Metavir F4), liver transplant recipients, and patients with severe extrahepatic hepatitis C

• Based on available resources, treatment should be prioritized as necessary so that patients at high risk for liver-related complications and severe extrahepatichepatitis C complications are given high priority

Accessed 1/20/2015:http://www.hcvguidelines.org/full-report/when-and-whom-initiate-hcv-therapy

Diagnosis and Staging Combined for Efficiency

• Initial Visit (PCP)HCV A tib d

• Second visit (Specialist)R i HCV RNA– HCV Antibody

• POCT

– Send off • HCV-RNA• Genotype

– Review HCV-RNA– Review genotype– Obtain Fibroscan®– Initiate anti-viral therapy


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