CCO HCV Assays Kwo Slides

Understanding Optimal Use and Interpretation of Assays in HCV

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clinicaloptions.com/hepatitisUnderstanding the Optimal Use and Interpretation of Assays in HCV

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Faculty Affiliation and Disclosure

Paul Y. Kwo, MDProfessor of MedicineMedical Director of TransplantationDivision of Medicine/Gastroenterology/HepatologyIndiana University School of MedicineIndianapolis, Indiana

Paul Y. Kwo, MD, has disclosed that he has received consulting fees from Abbott, Anadys, Boehringer Ingelheim, Bristol-Myers Squibb, Gilead Sciences, Johnson & Johnson, Merck, Novartis, and Vertex; has received fees for non-CME services from Bristol-Myers Squibb, Gilead Sciences, Merck, and Roche; and has contracted research with Abbott, Anadys, Bayer, Bristol-Myers Squibb, Conatus, GlaxoSmithKline, Merck, Novartis, Roche, and Vertex.

New Standard of Care for Patients With HCV


Patterns of Virologic Response

7

6

5

4

3

2

1

00-2-4-8 4 8 12 16 20 24 32 40 48 52 60 72

Wks After Start of Therapy

HC

V R

NA

(lo

g10

IU/m

L)[1

]

Undetectable

RVR EVR EOT SVR

Relapse

Partial response

Null response

1. Ghany MG, et al. Hepatology. 2009;49:1335-1374. 2. McHutchison JG, et al. N Engl J Med. 2009;361:580-593.

40% chance of SVR with

pegIFN/RBV[2]


Addition of TVR or BOC to PegIFN/RBV Improves SVR in Genotype 1 Patients HCV NS3/4A protease inhibitors BOC and TVR approved by FDA, May 2011[1,2]

– Indicated in combination with pegIFN/RBV for treatment of genotype 1 HCV–infected patients who are previously untreated or who have failed previous therapy

1. Boceprevir [package insert]. May 2011. 2. Telaprevir [package insert]. May 2011. 3. Poordad F, et al. N Engl J Med. 2011;364:1195-1206. 4. Jacobson IM, et al. N Engl J Med. 2011;364:2405-2416. 5. Bacon BR, et al. N Engl J Med. 2011;364:1207-1217. 6. Zeuzem S, et al. N Engl J Med. 2011;364:2417-2428. 7. Vierling J, et al. AASLD 2011. Abstract 931.

0

20

40

60

80

100

SV

R (

%)

Relapsers[5,6] Partial Responders[5,6]

PegIFN + RBV

NullResponders[6,7]

BOC/TVR + pegIFN + RBV

24-29

7-15

29-38

5

69-83

40-59

63-75

38-44

Treatment Naive[3,4]


Proper Use of HCV Assays Essential For Successful Management With HCV PIs HCV RNA level important throughout treatment to

determine

– Eligibility for shortened therapy (response-guided therapy)

– Discontinuation of therapy due to futility

– Minimizes risk of resistance and unnecessary adverse events

– Assessment of EOT response

– Assessment of SVR

Additional genetic testing may help predict response to treatment

HCV RNA Assays in the Protease Inhibitor Era


Key Challenges Regarding Use of HCV RNA Assays in Protease Inhibitor Era Package inserts for BOC and TVR specify different

time points for monitoring HCV RNA

Available HCV RNA assays in practice have different quantifiable ranges

Different HCV RNA thresholds used for RGT determination vs SVR

Different HCV RNA thresholds used for defining treatment futility with BOC vs TVR


HCV RNA Assays: LLOD Is Distinct From LLOQ LLOQ

– Lowest HCV RNA concentration within linear range of assay

– ie, smallest amount of HCV RNA that can be not only detected but also accurately quantified

LLOD

– Lowest amount of HCV RNA concentration that can be detected with 95% probability to determine presence or absence


0

1

2

4

6

8

0 Time

Lo

g10

Vir

al T

iter

Detectable/quantifiable

SVR

Not quantifiable/not detectable

LLOQLLOD

Detectable/not quantifiable

Adapted from Naeger LK, et al. Intl Workshop on Clinical Pharmacology of Hepatitis Therapy 2011. Abstract R-8.

HCV RNA Levels and Relationship to LLOD and LLOQ

HCV Treatment


Assay (Manufacturer ) Method LLOD, IU/mL Setting

Amplicor HCV v2.0 (Roche Molecular Systems)

Manual RT-PCR 50Diagnosis and

monitoring

Cobas Amplicor HCV v2.0 (Roche Molecular Systems)

Semiautomated RT-PCR 50Diagnosis and

monitoring

Ampliscreen (Roche Molecular Systems)

Semiautomated RT-PCR < 50 Blood

screening

Versant HCV RNA Qualitative Assay (Siemens Healthcare Diagnostics)

Semiautomated TMA 10Diagnosis and

monitoring

Procleix HIV-1/HCV Assay (Chiron Corporation)

Manual TMA < 50 Blood

screening

Ghany MG, et al. Hepatology. 2009;49:1355-1374.

All assays report HCV RNA as detected/not detected

FDA-Approved Qualitative HCV RNA Assays


Quantitative HCV RNA Assays

Assay (Manufacturer)[1] Method Dynamic Range, IU/mL(LLOQ-ULOQ)

LLOD, IU/mL FDA Approved

Amplicor HCV Monitor (Roche Molecular Systems) Manual RT-PCR 600-500,000 N/A Yes

Cobas Amplicor HCV Monitor V2.0 (Roche Molecular Systems) Semiautomated RT-PCR 600-500,000 600 Yes

Versant HCV RNA 3.0 Assay (bDNA) (Siemens Health Care Diagnostics)

Semiautomated bDNA signal amplification 615-7,700,000 615 Yes

LCx HCV RNA-Quantitative Assay (Abbott Diagnostics) Semiautomated RT-PCR 25-2,630,000 23 No

SuperQuant (National Genetics Institute) Semiautomated RT-PCR 30-1,470,000 30 No

Cobas TaqMan HCV Test (Roche Molecular Systems)

Semiautomated RT-PCR 43-69,000,000 18 Yes

COBAS TaqMan HCV Test v2.0 for use with High Pure System (Roche Molecular Systems)

Semiautomated RT-PCR 25-300,000,000 15 Yes

Abbott RealTime HCV Assay (Abbott Diagnostics) Semiautomated RT-PCR 12-100,000,000 12 Yes

Phase III registration trials for both BOC and TVR used COBAS TaqMan HCV Test v2.0 for use with High Pure System

– 1.3% false-positive rate[2]

1. Ghany MG, et al. Hepatology. 2009;49:1355-1374. 2. Naeger LK, et al. Intl Workshop on Clinical Pharmacology of Hepatitis Therapy 2011. Abstract R-8.


SVR Rate by HCV RNA Status (LLOQ vs LLOD) for BOC and TVR

BOC/PR RGT T12/PR

SVR rate lower when HCV RNA not undetectable at key time points during therapy

100

0

40

SV

R (

%)

60

20

80

100

0

40S

VR

(%

)

60

20

4

80

8 10 12 16 20

Undetectable (Below LLOD)Detectable/Below LLOQAbove LLOQ (> 25 IU/mL)

Treatment Wk

4 6 10 12 16 208

Naeger LK, et al. Intl Workshop on Clinical Pharmacology of Hepatitis Therapy 2011. Abstract R-8.

Treatment Wk

Using HCV RNA Assays in Clinical Practice


61

192/314

Predictive Value of Baseline HCV RNA for Achieving SVR

100

0

50

7874

SV

R (

%)

75

25

207/281

64/82

> 800,000 IU/mL≤ 800,000 IU/mL

ADVANCE (TVR)[1]

1. Jacobson IM, et al. N Engl J Med. 2011;364:2405-2416. 2. Poordad F, et al. N Engl J Med. 2011;364:1195-1206.

100

0

50

SV

R (

%)

75

25

SPRINT-2 (BOC)[2]

8576

41/54

45/53

63

197/313

BOC/PR48 BOC/PR RGTT12PR arm

n/N = n/N =

≥ 800,000 IU/mL< 800,000 IU/mL


Patients Responding Early Can Achieve High SVR Rates With Shortened Therapy Response-guided therapy: patients who achieve optimal virologic

response at early time points can receive abbreviated therapy without reducing their chance of SVR

Patients eligible for RGT

– Boceprevir: noncirrhotic treatment-naive patients, previous relapsers, and previous partial responders[1,2]

– RGT criterion: Must achieve undetectable HCV RNA at Wk 8 (ie, Wk 4 of triple therapy) and maintain it at Wk 24

– Telaprevir: noncirrhotic treatment-naive patients and previous relapsers*[2,3]

– RGT criterion: Must achieve undetectable HCV RNA at Wk 4 of triple therapy and maintain it at Wk 12

1. Boceprevir [package insert]. May 2011. 2. Ghany MG, et al. Hepatology. 2011;54:1433-1444. 3. Telaprevir [package insert]. May 2011.

*AASLD guidelines state that RGT may be considered with TVR in previous partial responders.


Response-Guided Therapy Paradigm With BOC + PegIFN/RBV in Tx-Naive Patients

BOC + PegIFN +RBV

480 28124

PegIFN + RBV

8 3624

Early response stop at Wk 28; f/u 24 wks

HCV RNA Undetectable Undetectable

480 28124

PegIFN + RBVPegIFN + RBV

8 36

BOC + PegIFN +RBV

24

HCV RNA Detectable Undetectable Slow response extend triple therapy

to Wk 36; PR to Wk 48; f/u 24 wks

< 100 IU/mL

< 100 IU/mL

Indicated for all noncirrhotic treatment-naive patients

Boceprevir [package insert]. May 2011. Ghany MG, et al. Hepatology. 2011;54:1433-1444.


Response-Guided Therapy Paradigm With BOC + PegIFN/RBV in Tx-Exp Patients

BOC + PegIFN +RBV

480 28124

PegIFN + RBV

8 3624

HCV RNA Undetectable Undetectable

480 28124

PegIFN + RBVPegIFN + RBV

8 36

BOC + PegIFN +RBV

24

HCV RNA Detectable Undetectable

< 100 IU/mL

< 100 IU/mL

Indicated for noncirrhotic previous relapsers or partial responders

Boceprevir [package insert]. May 2011. Ghany MG, et al. Hepatology. 2011;54:1433-1444.

Early response stop at Wk 36; f/u 24 wks

Slow responsePR to Wk 48; f/u 24 wks


Response-Guided Therapy Paradigm With TVR + PegIFN/RBV in Tx-Naive Patients

TVR + PegIFN + RBV

480 24124

eRVR stop at Wk 24, f/u 24 wksPegIFN + RBV

TVR + PegIFN + RBV

480 24124

PegIFN + RBV

HCV RNAUndetectable

Undetectable

Detectable (≤ 1000 IU/mL)

Undetectable or detectable (≤ 1000 IU/mL) No eRVR extend pegIFN +

RBV to Wk 48; f/u 24 wks

HCV RNA

Indicated for all noncirrhotic treatment-naive patients

Telaprevir [package insert]. May 2011. Ghany MG, et al. Hepatology. 2011;54:1433-1444.

Undetectable

Undetectable


Response-Guided Therapy Paradigm With TVR + PegIFN/RBV in Tx-Exp Patients

TVR + PegIFN + RBV

480 24124

PegIFN + RBV

Detectable (≤ 1000 IU/mL)

Undetectable/detectable (≤ 1000 IU/mL)

No eRVR extend pegIFN + RBV to Week 48; f/u 24 wks

HCV RNA

1. Telaprevir [package insert]. May 2011. 2. Ghany MG, et al. Hepatology. 2011;54:1433-1444.

*AASLD guidelines say RGT “may be considered” for prior partial responders [2] but package insert recommends 48 weeks of therapy[1]

Same as naives; indicated for noncirrhotic previous relapsers[1]*

TVR + PegIFN + RBV

480 24124

eRVR stop at Wk 24, f/u 24 wksPegIFN + RBV

HCV RNAUndetectable

Undetectable

Undetectable

Undetectable


HCV RNA Assay Characteristics for RGT With BOC or TVR A quantitative assay with an LLOQ of ≤ 25 IU/mL and an

LLOD of approximately 10-15 IU/mL must be used

“Confirmed detectable but below limit of quantification HCV RNA result should not be considered equivalent to an undetectable HCV RNA result”

Boceprevir [package insert]. May 2011. Telaprevir [package insert]. May 2011.

0

1

2

4

6

8

0 Time

Lo

g10

Vir

al T

iter

Detectable/quantifiable

Undetectable/not quantifiable

LLOQ

LLODDetectable/not quantifiable


Predictive Value of Response to 4-Wk Lead-in Phase ≥ 1 log10 vs < 1 log10 decline in HCV RNA following

4-wk lead-in phase with pegIFN/RBV strongly predicts SVR in patients receiving BOC-based therapy

– Treatment-naive patients[1]

– OR: 9.0; P < .001

– Treatment-experienced patients[2]

– OR: 5.2; P < .001

1. Poordad F, et al. N Engl J Med. 2011;364:1195-1206. 2. Zeuzem S, et al. EASL 2011. Abstract 484.


Boceprevir [package insert]. May 2011.

BOC + PegIFN +RBV

480 28124

PegIFN + RBV

PegIFN + RBV

8 36

BOC + PegIFN +RBV

24

Early response*; stop at Wk 28 or 36; f/u 24 wks

F/u 24 wks

*Undetectable HCV RNA at Wks 8 and 24 of therapy (Wk 4 of triple therapy).

Wks

Stop all treatment if HCV RNA ≥ 100 IU/mL

Stop all treatment if HCV RNA detectable

(> LLOD)

Use quantitative assay to determine if HCV RNA

< or ≥ 100 IU/mL at Wk 12

Use assay with LLOD of 10-15 IU/mL to determine if

HCV RNA detectable at Wk 24

Boceprevir Futility Rules: Wks 12 and 24 Key Time Points Treatment-naive and treatment-experienced patients


Telaprevir Futility Rules: Wks 4, 12, and 24 Key Time Points Treatment-naive and treatment-experienced patients

TVR + PegIFN +RBV

Wks480 24124

eRVR*; stop at Wk 24; f/u 24 wksPegIFN + RBV

No eRVR; PegIFN + RBV

Telaprevir [package insert]. May 2011.

F/u 24 wks

*Undetectable HCV RNA at Wks 4 and 12 of triple therapy.

Use quantitative assay to determine if HCV RNA ≤ or > 1000 IU/mL

at Wks 4 and 12

Use assay with LLOD of 10-15 IU/mL to determine if

HCV RNA detectable at Wk 24

Stop all treatment if HCV RNA > 1000 IU/mL

Stop all treatment if HCV RNA > 1000 IU/mL

Stop all treatment if HCV RNA detectable

(> LLOD)


EOT response defined as[1,2] – HCV RNA < LLOD at EOT

– Using an assay with a sensitivity of 10-15 IU/mL[1,2]

Detectable but < LLOQ values while on treatment predict lower SVR rates[3]

1. Boceprevir [package insert]. May 2011. 2. Telaprevir [package insert]. May 2011. 3. Naeger LK, et al. Intl Workshop on Clinical Pharmacology of Hepatitis Therapy 2011. Abstract R-8.

HCV RNA Thresholds for EOT Response With BOC or TVR


Use of HCV RNA Assays to Assess SVR With BOC or TVR-Based Therapy SVR to pegIFN/RBV previously defined as

– Absence of detectable HCV RNA in serum using assay with sensitivity of at least 50 IU/mL 6 mos after EOT[1]

SVR defined by FDA in BOC and TVR package inserts as

– HCV RNA < 25 IU/mL (LLOQ) 6 mos after EOT[2-3]

1. Lindsay KL, et al. Hepatology. 2002;36:S114-S120. 2. Boceprevir [package insert]. May 2011. 3. Telaprevir [package insert]. May 2011.


Summary: Use of HCV RNA Assays in Managing Patients Receiving BOC or TVR A quantitative assay with an LLOQ of ≤ 25 IU/mL and an LLOD of

approximately 10-15 IU/mL must be used

HCV RNA < LLOQ not identical to HCV RNA < LLOD

– HCV RNA < LLOD required to qualify for RGT

– HCV RNA < LLOQ appropriate for assessing SVR

Qualification/Endpoint BOC TVR

RGT HCV RNA < LLOD at Wks 8 and 24

HCV RNA < LLOD at Wks 4 and 12

Futility HCV RNA ≥ 100 IU/mL at Wk 12

HCV RNA > LLOD at Wk 24

HCV RNA > 1000 IU/mL at Wk 4 or 12

HCV RNA > LLOD at Wk 24

EOT response HCV RNA < LLOD at EOT

SVR HCV RNA < LLOQ 24 wks after EOT

Other Assays


P < .0001

P < .0001

P < .0001

P < .0001

P < .0001

P = .004

Thompson AJ, et al. Gastroenterol. 2010;139:120-129.

IL28B Genotype the Strongest Baseline Predictor of SVR With PegIFN/RBV

Metavir F0-2

White vs Black

Fasting Serum Glucose < 5.6 mmol/L

Hispanic vs Black

HCV RNA ≤ 600,000 IU/mL

CC vs Non-CC

Odds Ratio (95% CI)

0 1 2 3 4 5 6 7 8


1. Poordad F, et al. EASL 2011. Abstract 12. 2. Jacobson IM, et al. EASL 2011. Abstract 1369.

SPRINT-2: BOC + PR48[1]

SV

R (

%)

44/55

82/115

26/44

CC CT TT

80

100

80

60

40

20

0

71

59

n/N =

ADVANCE*: T12PR[2]

SV

R (

%)

45/50

48/68

16/22

CC CT TT

90100

80

60

40

20

0

71 73

n/N =

*IL28B testing in ADVANCE was in white pts only.

IL28B Genotype Also Predicts Likelihood of Achieving SVR With BOC or TVR


IL28B Genotype Predicts Likelihood of Shortened Therapy With BOC or TVR

1. Poordad F, et al. EASL 2011. Abstract 12. 2. Jacobson IM, et al. EASL 2011. Abstract 1369.

Elig

ibili

ty f

or

Sh

ort

ened

T

her

apy

(%)

118/132

158/304

CC CT/TT

89

52

Elig

ibili

ty f

or

Sh

ort

ened

T

her

apy

(%)

39/50

39/68

10/22

78

57

45

SPRINT-2: BOC + PR[1] ADVANCE*: T12PR[2]

*IL28B testing in ADVANCE was in white pts only.

80

60

40

20

0

n/N =

CC CT TT

100

80

60

40

20

0

n/N =

100


When to Consider IL28B Genotype Testing

IL28B genotype testing may be considered prior to therapy if more information about probability of response or treatment duration desired[1]

– Commercially available tests

If patients have favorable CC genotype

– Likelihood of SVR is high with pegIFN/RBV alone, but triple therapy may allow shorter therapy and, in one TVR study, higher SVR rates[2]

If patients have unfavorable CT/TT genotype

– Likelihood of SVR is higher with triple therapy than with pegIFN/RBV [2,3]

Limited value of IL28B genotyping in treatment-experienced patients

– Most have unfavorable TT or CT genotype

1. Ghany MG, et al. Hepatology. 2011;54:1433-1444. 2. Jacobson IM, et al. EASL 2011. Abstract 1369. 3. Poordad F, et al. EASL 2011. Abstract 12.


HCV Genotype and Subtype

HCV classified into 6 major genotypes (1-6)[1]

Genotype 1 (subtypes a and b) most common in United States (~ 75%)[2]

– Subtype 1a more common than subtype 1b

Determining major genotype recommended for proper clinical management and predicting likelihood of response[3]

No current recommendations regarding HCV subtype testing

1. Simmonds P, et al. Hepatology. 2005;42:962-973. 2. Zein N. Clin Microbiol Rev. 2000;13:223-235. 3. Ghany MG, et al. Hepatology. 2009;49:1335-1374.


Higher SVR Rates With TVR in Patients With HCV Genotype 1b vs 1a

Tx Naive[1]

T12/PR48Tx Naive[1]

T12/PR48

7171

4747

SV

R (

%)

SV

R (

%)

0

20

40

60

80

100 Genotype 1aGenotype 1a

Genotype 1bGenotype 1b

Relapsers*[2]Relapsers*[2] Null Responders*[2]

Null Responders*[2]

Partial Responders*[2]

Partial Responders*[2]

1. Jacobson IM, et al. N Engl J Med. 2011;364:2405-2416. 2. Zeuzem S, et al. EASL 2011. Abstract 5.

79798484

8888

2727

6868

3737

*Pooled TVR arms.*Pooled TVR arms.


Higher SVR Rates With BOC in Patients With HCV Genotype 1b vs 1a

BOC RGTBOC RGT

59595050

Genotype 1aGenotype 1a

Genotype 1bGenotype 1b

Treatment Naive[1]Treatment Naive[1]

BOC/PR48BOC/PR48 BOC/PR48BOC/PR48BOC RGTBOC RGT

Treatment Experienced[2]Treatment Experienced[2]

1. Poordad F, et al. N Engl J Med. 2011;364:1195-1206. 2. Bacon BR, et al. N Engl J Med. 2011;364:1207-1217.

6666 6363

7070

61616565

7373

0

20

40

60

80

100

SV

R (

%)

SV

R (

%)


Genotype assay Manufacturer Method

Trugene 5'NC HCV Genotyping kit

Siemens Direct sequence analysis of the 5' noncoding region

INNO-LiPa HCV II Innogenetics Reverse hybridization analysis using genotype-specific oligonucleotide

probes located in the 5' noncoding region

Versant HCV Genotyping Assay 2.0

Siemens Reverse hybridization analysis using genotype-specific oligonucleotide

probes located in the 5' noncoding region

Abbott RealTime HCV Genotype II

Abbott Genotype-specific real-time PCR of the 5' noncoding region and NS5b

Incorrect typing among major genotypes rare (< 3%)

Ghany MG, et al. Hepatology. 2009;49:1335-1374.

Commercially Available HCV Genotype Assays


Resistance-associated variants occur naturally[1]

– Present in 5% to 7% of subject samples prior to treatment[2,3]

– No apparent impact on likelihood of SVR

– Selected for/enriched in patients failing PI-based therapy

Following treatment failure, resistance-associated variants decline over time after withdrawal of PI but may remain detectable for up to 2.5 yrs[4,5]

Lower genetic barrier to resistance (number of mutations required to overcome virologic activity of the regimen) with genotype 1a vs 1b with BOC/TVR–based regimens

Strict adherence to futility rules, ensuring patient adherence and tolerability of regimen essential to avoid resistance

1. Pawlotsky JM. Clin Liver Dis. 2003;7:45-66. 2. Telaprevir [package insert]. May 2011. 3. Boceprevir [package insert]. May 2011. 4. Vierling JM, et al. EASL 2010. Abstract 2016. 5. Sullivan JC, et al. EASL 2011. Abstract 8.

HCV Resistance With TVR/BOC


HCV Resistance Testing

Commercial resistance test for HCV NS3/4 mutations now available

– Provides genetic sequence for the nonstructural proteins NS3 and NS4A of HCV genotypes 1a and 1b

Role of resistance testing prior to treatment remains to be defined

– No current recommendation to perform resistance testing for patients failing therapy


Summary: Use of Genotype and Resistance Assays With BOC/TVR IL28B genotype testing

– May be considered prior to therapy if more information about probability of response or treatment duration desired[1]

HCV subtype testing

– No current recommendation to test prior to treatment

– Patients with genotype 1b may be counseled that their chance of SVR is slightly higher than 1a patients

HCV resistance testing

– No current recommendation regarding testing at baseline or upon treatment failure

1. Ghany MG, et al. Hepatology. 2011;54:1433-1444.

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