Brief HTA report: Genotipo di polimorfismi del gene dell ...

HTA report: Genotipo di polimorfismi del gene dell’IL28B (IFNL3): utilizzo clinico in pazienti con HCV V4-2

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Rete Ligure HTA

Brief HTA report:

Genotipo di polimorfismi del gene dell’IL28B (IFNL3): utilizzo clinico in pazienti con HCV

Redazione a cura di:

Francesco Cardinale, Emilio Di Maria, Gaddo Flego per il Gruppo di Coordinamento della Rete Ligure HTA

Febbraio 2014

Indice:

Summary

Sintesi

Premesse e ambito di applicazione

Analisi farmacogenetica di IL28B in pazienti con HCV

o Terapia basata su PEG-Interferon-α.

o Test farmacogenetico: analisi di polimorfismi di IL28B

o Meccanismo biologico

o Applicazione in popolazioni di diversa origine

o Genotipo virale

o Co-infezione HCV-HIV

Validità analitica

Validità clinica

Utilità clinica

o Implicazioni etiche, legali e sociali

Valutazioni economiche

Conflitti di interesse

Censimento attività in Liguria

Discussione

Bibliografia

Appendice:

o Reference list: systematic reviews and meta-analyses

o Reference list: selection of primary studies


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Summary

Background:

PEG-Interferon-α and Ribavirin association is currently the standard regimen for treatment

of patients suffering from HCV infection. The genotype of polymorphisms located in the

IL28B (IFNL3) gene is considered a reliable predictor of response to therapy, as measured

by the mean of the sustained virologic response (SVR).

Aims:

The purpose of the present document is to provide information about the application in the

clinical practice of the pharmacogenetic test based on IL28B genotyping to guide the

administration of PEG-Interferon-α-based therapy in patients with HCV infection. The use

of this test within the Regional Health System was also evaluated. Economic evaluations

were not considered.

Methods:

A focused literature review was accomplished; a systematic search was performed to

identify available systematic reviews and meta-analyses. A telephone survey was

performed to identify laboratories offering the IL28B pharmacogenetic test within the

Regional Health System and collect current activity.

Analytic validity:

The genetic test is routinely based on real-time PCR allelic discrimination of the

rs12979860 single nucleotide polymorphism. Despite the lack of large studies on the

analytic performance, the test procedure is robust and was reported to be readily

implemented in the pipeline of molecular pathology laboratories.

Clinical validity:

According to all the meta-analyses reviewed in the present assessment, the genotype of

the IL28B rs12979860 polymorphism is a valid predictor of sustained virologic response in

previously untreated patients with HCV. Carriers of the rs12979860 CC genotype show a

more favourable response rate, as compared to non-CC carriers. Genotyping of other

IL28B polymorphisms is not suggested.

The association between response and IL28B genotypes was demonstrated for HCV

genotype 1 and 4, while in genotype 2 and 3 was not definitely confirmed. The association

with other endpoints, such as the rapid virologic response, was not extensively studied.

Clinical utility:

Though the sustained virologic response can be considered a valid measure of the clinical

outcome and was widely used as endpoint in clinical studies, no evidence was found

supporting the hypothesis that a regimen guided by IL28B genotyping is associated with a

more favourable clinical outcome with respect to standard protocol. No clinical measure of

efficacy, other than virologic response, was consistently evaluated in primary studies. Also

occurrence of adverse events was not explored in association studies on IL28B genotype.

Noteworthy, a multivariate model including both clinical variables and IL28B genotyping as

predictors was preliminarily demonstrated to more accurately predict virologic response

with respect to IL28B alone. Clinical pathways incorporating the pharmacogenetic test of

IL28B were not formally evaluated in clinical trials and currently are not recommended in

clinical guidelines. Ongoing studies were found in the clinical trial database.


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There is no literature exploring the ethical, legal and social implications of IL28B testing.

However, this type of test does not raise specific concerns, provided that good practices

are applied.

Provision of IL28B genotyping by laboratories of the Regional Health System:

The survey detected three laboratories offering pharmacogenetic IL28B testing.

Approximately 550 test/year were performed in the 2012-2013 period.

The ability of each laboratory to perform a larger number of test should be further

estimated. It could also be considered to what extent efficiency can be improved by

gathering samples in one laboratory. In addition, it should be also investigated to what

extent laboratories performing few tests can guarantee a timely response, as this is a

critical requisite to guarantee the proper use of the pharmacogenetic tests.


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Sintesi

Premesse:

L’associazione di PEG-Interferon-α and Ribavirina è attualmente il trattamento standard in

pazienti con infezione da HCV. Il genotipo di polimorfismi localizzati nel gene IL28B

(IFNL3) è considerato un predittore attendibile ella risposta alla terapia, misurata in termini

di risposta virale protratta (sustained virologic response, SVR).

Obiettivo:

Lo scopo del presente documento è fornire informazioni sull’applicazione nella pratica

clinica del test farmacogenetico basato sulla genotipizzazione di IL28B finalizzata

orientare la terapia basata su PEG-Interferon-α in pazienti con HCV. È stato inoltre

valutato l’utilizzo del test nell’ambito del Sistema Sanitario Regionale. Non sono state

considerate le valutazioni economiche.

Metodi:

È stata condotta una ricognizione della letteratura; una ricerca sistematica è stata

effettuata per identificare le rassegne sistematiche e le meta-analisi disponibili.

Un’indagine è stata condotta per rilevare le strutture che offrono il test farmacogenetico di

IL28B e l’attività attuale nell’ambito del Sistema Sanitario Regionale.

Validità analitica:

Il test genetico è comunemente basato su un saggio di discriminazione allelica mediante

real-time PCR del polimorfismo a singolo nucleotide rs12979860. Nonostante non siano

disponibili ampi studi sulle caratteristiche analitiche, la procedura del test è affidabile ed è

stata descritta come facilmente implementabile nel flusso di lavoro dei laboratori che

offrono indagini molecolari.

Validità clinica:

Secondo le meta-analisi prese in considerazione per questa valutazione, il genotipo del

polimorfismo rs12979860 di IL28B è un valido predittore della risposta virale protratta in

pazienti che non siano stati precedentemente trattati. I pazienti portatori del genotipo CC

di rs12979860 presentano un tasso di risposta più favorevole, in confronto ai portatori di

genotipo CT o TT. La genotipizzazione di altri polimorfismi non è raccomandata.

L’associazione tra risposta e genotipi di IL28B è stata dimostrata per i genotipi virali 1 e 4,

mentre non è stata confermata nei genotipi 2 e 3. L’associazione con altre misure di esito,

come la risposta rapida, non è stata studiata in modo esteso.

Utilità clinica:

Sebbene la risposta virale protratta possa essere considerata una misura valida dell’esito

clinico, e sia stata largamente adottata come endpoint negli studi clinici, non sono state

reperite prove a sostegno dell’ipotesi che un regime terapeutico orientato dal genotipo di

IL28B sia associato con un esito clinico più favorevole rispetto al protocollo standard di

trattamento. Negli studi primari non sono state sistematicamente valutate altre misure di

efficacia. Anche l’occorrenza di eventi avversi non è stata indagata negli studi di

associazione con il genotipo di IL28B.

È degno di nota che un modello multivariato, che include sia variabili cliniche sia il

genotipo di IL28B, sia stato dimostrato in grado di predire la risposta virale in maniera più

accurata rispetto al solo genotipo di IL28B. Percorsi clinici che comprendono il test

farmacogenetico di IL28B non sono stati formalmente valutati in studi clinici e attualmente


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non vengono raccomandati da linee guida. Nel catalogo degli studi clinici sono stati trovati

trial in corso.

Non si trova letteratura che abbia indagato gli aspetti etici, legali e sociali del test di IL28B.

Tuttavia questo tipo di test non solleva criticità peculiari, assumendo che siano applicate le

buone pratiche.

Erogazione del test nei laboratori del Sistema Sanitario Regionale:

L’indagine ha rilevato tre laboratori che offrono il test farmacogenetico di IL28B. Circa 550

test per anno sono stati erogati nel periodo 2012-2013. La capacità di ciascun laboratorio

di effettuare un maggiore numero di test dovrebbe essere ulteriormente stimata. Potrebbe

anche essere considerato in che misura si possa migliorare l’efficienza concentrando i

campioni in un singolo laboratorio. Dovrebbe inoltre essere indagato se i laboratori che

eseguono un numero limitato di test possano garantire la tempestività della risposta,

giacché questo è un requisito rilevante per garantire un utilizzo appropriato dei test

farmacogenetici.


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Premesse e ambito di applicazione

Il presente documento è un sintetico HTA report che analizza l’applicazione in ambito clinico del

test farmacogenetico di polimorfismi del gene IL28B per orientare la terapia basata su PEG-

Interferon-α in pazienti affetti da HCV.

È stata effettuata una ricerca bibliografica attraverso banche dati pubbliche (PubMed) focalizzata

su IL28B e HCV; una ricerca sistematica ha rilevato tutte le rassegne sistematiche e le meta-

analisi pubblicate fino a febbraio 2014 [si veda l’appendice: “Reference list: Systematic reviews

and meta-analyses”].

Sono state considerate le prove a supporto della validità clinica1 e dell’utilità clinica2 del test; è

stato inoltre effettuato un rilevamento delle strutture che offrono l’analisi di IL28B nell’ambito del

SSR e del volume di analisi attualmente erogate.

Non rientrano quindi nella valutazione: l’efficacia della terapia, i percorsi clinici, l’utilizzo di test

farmacogenetici diversi dalla genotipizzazione di IL28B, test farmacogenetici per trattamenti diversi

da quelli basati su PEG-Interferon-α.

Analisi farmacogenetica di IL28B in pazienti con HCV

Terapia basata su PEG-Interferon-α

L’associazione di PEG-Interferon-α (PEG-IFN) e Ribavirina (duplice terapia) costituisce la terapia

standard in pazienti con epatite sostenuta da HCV. In presenza del genotipo virale 1 o 4 si

raccomanda la somministrazione per 48 settimane, che può essere limitata a 24 nei casi con

genotipo virale 2 o 3. I trial più recenti (SPRINT-2 e ADVANCE) hanno dimostrato che la

combinazione con inibitori delle proteasi (boceprevir e telaprevir) migliora la risposta dal 38-44% al

66-75%. La triplice terapia è però gravata da maggiori effetti collaterali e implica un costo

notevolmente più elevato [per una rassegna si veda Kronenberger B 2009; per le linee guida sul

trattamento Ghany MG 2009; per un aggiornamento delle raccomandazioni Ghany MG 2011].

Gli studi clinici utilizzano come endpoint la risposta virale persistente (sustained virologic response,

SVR), ovvero la negativizzazione della carica virale plasmatica a sei mesi dalla fine della terapia.

Altri indicatori di risposta, come la risposta virale a breve termine (rapid virologic response, RVR)

sono stati impiegati prevalentemente come endpoint secondari.

Test farmacogenetico: analisi di polimorfismi di IL28B

Dopo il primo studio condotto da Ge e collaboratori [2009], numerosi studi primari hanno riportato

un’associazione statisticamente significativa tra polimorfismi del gene IL28B e la risposta al

trattamento in coorti di pazienti di diversa origine geografica. In particolare, l’associazione con il

polimorfismo a singolo nucleotide denominato rs12979860, presente con variabile frequenza

allelica nelle diverse popolazioni, è stata replicata in studi indipendenti. Il genotipo omozigote CC è

associato a una maggiore percentuale di risposta rispetto sia al genotipo eterozigote CT sia

all’omozigote TT [per una rassegna aggiornata si veda ad esempio Imran M 2013].

1 La validità clinica di un test genetico definisce la sua capacità di rilevare o predire un determinato fenotipo

(in genere un quadro patologico); nel caso di un test farmagenetico la validità clinica misura la capacità di predire la risposta a un trattamento, misurata attraverso un indicatore appropriato. 2 L'utilità clinica di un test genetico è la misura del suo valore in termini di salute per il paziente, misurato

attraverso un indicatore di esito.


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Sottolineiamo che gli stessi indicatori utilizzati nei clinical trials sono stati adottati negli studi di

farmacogenetica3. Poiché il SVR è l’endpoint comunemente utilizzato negli studi primari sui test

farmacogenetici nella terapia dell’HCV, è stato considerato nelle meta-analisi successive.

Sono state rilevate nove tra rassegne sistematiche e meta-analisi apparse negli anni recenti, otto

delle quali hanno analizzato pazienti con genotipo virale 1. Tutte hanno confermato la validità

dell’associazione tra genotipo CC di rs12979860 e il SVR in pazienti naïve con infezione da

genotipo 1 o 4, trattati sia con duplice sia con triplice terapia. Il tasso di SVR si dimostra

significativamente più elevato in pazienti con il genotipo CC rispetto agli altri genotipi (CT and TT).

L’odds ratio stimato nelle meta-analisi varia tra 3.8 e 5.2 [vedi appendice: Reference List:

Systematic Reviews & Meta-analyses]. Riportiamo come esempio i risultati pubblicati da Jia Z

[2012].

Tratta da: Jia, Z., et al. Test of IL28B polymorphisms in chronic hepatitis C patients treated with PegIFN and ribavirin depends on HCV genotypes: results from a meta-analysis. PLoS ONE 7[9], e45698. 2012.

Il Clinical Pharmacogenetics Implementation Consortium [Muir AJ 2013] ha sviluppato linee guida

basata su una rassegna sistematica e una valutazione strutturata della letteratura. Gli autori

concludono che il genotipo di IL28B è il principale predittore di risposta a PEG-IFN e Ribavirina in

pazienti non precedentemente trattati e che può essere utilizzato da pazienti e curanti come parte

di un percorso decisionale condiviso per la scelta del trattamento.

Un recente documento della European Medicine Agency (EMA) dal titolo “Guideline on clinical evaluation of medicinal products for the treatment of chronic hepatitis C”

3 La European Medicine Agency, attraverso la pubblicazione del documento “Note for guidance on

definitions for Genoimic biomarkers, pharmacogenomics, pharmacogenetics, genomica data and sample coding categories” (EMEA/CHMP/ICH/437986/2006), definisce la farmacogenetica come lo studio di varianti nelle sequenze di DNA in relazione alla risposta ai farmaci. In quanto tale, la farmacogenetica è un sottoinsieme della farmacogenomica, che è definita come lo studio delle caratteristiche del DNA e dell’RNA in relazione alla risposta ai farmaci. La principale differenza è che la farmacogenetica studia varianti ereditate proprie del DNA, mentre la farmacogenomica include anche l’analisi di varianti somatiche nel RNA (vedi anche la definizione dell’EMA di biomarker genomico: una caratteristica misurabile del DNA e/o del RNA, che sia indicatore di processi biologici normali, di processi patogenetici, e/o di risposta alla terapia o a altri interventi”).


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[EMEA/CHMP/51240/2011] riporta un paragrafo (4.1.2. Host IL28B genotype4) che menziona l’importanza della genotipizzazione del polimorfismo rs12979860 dell’ospite. Da sottolineare che il documento dell’EMA stabilisce le linee guida per la sperimentazione, e non per la pratica clinica.

Meccanismo biologico

Il gene IL28B, alias di IFNL3 (Interferon-Lambda-3) codifica per l’interferone λ, che contribuisce

alla risposta antivirale da parte dell’ospite. Il corrispondente biologico dell’associazione tra IL28B

rs12979860 e risposta alla terapia antivirale non è stato dimostrato. Il genotipo favorevole CC è

associato alla clearance spontanea nell’infezione acuta [Thomas DL 2009]; è associato inoltre alla

risposta a breve termine, RVR.

È quindi plausibile che un aplotipo funzionale identificato dall’allele C di rs12979860 favorisca la

risposta endogena al virus con genotipo 1 o 4. Secondo questa ipotesi, altre variabili cliniche

potrebbero essere validi indicatori di risposta [ad esempio: RVR; il tempo alla negativizzazione

della carica virale - estimated time point for 0 log viral copies/mL, Lindh M 2013].

Un recente studio di fase 2b sull’efficacia di trattamenti interferon-free non ha rilevato alcun effetto

sulla risposta associato al genotipo di IL28B [Kowdley KV 2014].

Applicazione in popolazioni di diversa origine

La variabilità tra popolazioni denominate “caucasian”, “hispanic”, “white non-hispanic”, “afro-

american”, etc., è ampiamente interpretabile sulla base della frequenza allelica del polimorfismo

rs12979860 nelle popolazioni di diverse origine5.

La popolazione di origine europea presenta una frequenza del genotipo favorevole CC pari al 38%

[Muir AJ 2013].

Non vi è alcuna evidenza che introdurre una variabile che descriva la genealogia o il gruppo etnico

migliori la corrispondenza del modello statistico basato sul genotipo predittore.

Genotipo virale

Ad oggi diversi studi hanno dimostrato un’associazione con il genotipo virale 1 ed un numero

minore di studi ha dimostrato un’associazione con il genotipo virale. Circa i genotipi 2 e 3 i risultati

sono contrastanti. La meta-analisi condotta da Lopez-Cortes LF [2012] non ha rilevato

un’associazione statisticamente significativa tra il genotipo di rs12979860 e la risposta in pazienti

co-infetti da HCV genotipo 3 e HIV.

In sintesi, le evidenze suggeriscono che i polimorfismi possano essere utili predittori pre-

trattamento solo nei genotipi 1 e 4 mentre per 2 e 3 la loro utilità sia limitata.

Co-infezione HCV-HIV

Il valore predittivo di tali polimorfismi è stato studiato per lo più in pazienti con sola infezione da

HCV, mentre pochi studi hanno esplorata l’associazione in pazienti HCV-HIV co-infetti. La meta-

analisi sopra citata [Lopez-Cortes LF 2012] non ha rilevato un’associazione in pazienti co-infetti

HCV genotipo 3 e HIV. Ulteriori studi sono necessari in tale contesto.

Validità analitica

4 “…Il-28 genotyping is rapidly becoming available in routine clinical practice. This should be considered,

when performing single arm confirmatory studies in populations where that is appropriate; the sponsor need to ensure that a particular IL28B genotype is not inappropriately selected for when recruiting for the study, e.g., by recruitment capping”. [http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2011/02/WC500102109.pdf] 5 Nella letteratura statunitense permane la stratificazione delle coorti di pazienti per “razza”. Ricordiamo per

inciso che l’attribuzione della variabile “race” non ha alcun razionale dal punto di vista genetico.


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Sono in commercio diversi kit per l’analisi del polimorfismo rs12979860 di IL28B; alcuni

comprendono anche la tipizzazione del polimorfismo rs8099917, che è stato analizzato in

numerosi studi. Tuttavia i polimorfismi studiati sono in linkage disequilibrium tra di loro; le

frequenze alleliche di rs12979860 riscontrate nella popolazione di origine europea (frequenza del

genotipo favorevole CC pari al 38% [Muir AJ 2013]) indicano una distribuzione informativa dei

genotipi; la tipizzazione di rs8099917 non sembra quindi apportare ulteriori informazioni

clinicamente utili [vedi di seguito il paragrafo “Validità clinica”].

Le metodiche (saggio mediante real-time PCR) sono comunemente adottate nei laboratori di

biologia molecolare e non presentano problematiche peculiari, purché la buone pratiche di

laboratorio siano rispettate [Cervinski MA et al 2014]. Non sono stati rilevati i valori di sensibilità e

specificità del test relativamente ai diversi prodotti commerciali. Il test può essere effettuato anche

utilizzando protocolli non commerciali sviluppati localmente.

Per la metodica basata su RT-PCR è stimabile un tempo totale di circa 3 ore dal check-in del

campione alla refertazione [vedi di seguito il paragrafo “Censimento delle attività in Liguria”].

Validità clinica

Le recenti meta-analisi [si veda l’appendice “Reference list: Systematic reviews and meta-

analyses”] indicano che il genotipo di IL28B rs12979860 è un valido predittore della SVR nei

pazienti affetti da HCV con genotipo 1 e genotipo 4, trattati sia con duplice terapia sia con triplice

terapia.

Non è definitivamente provata l’associazione con la risposta nelle infezioni da HCV genotipo 2 e 3,

anche se l’effetto è stato suggerito da alcuni studi recenti [Gupta AC et al 2014].

Una mole minore di risultati è disponibile per altri indicatori di risposta (e.g.: rapid virologic

response, RVR) [Gupta AC et al 2014].

Altri polimorfismi, oltre a rs12979860, risultano associati al fenotipo risposta; rs8099917 è stato

analizzato in numerosi studi. Tuttavia i polimorfismi studiati sono in linkage disequilibrium tra di

loro; non sono disponibili evidenze circa l’effetto di aplotipi rari. Non è pertanto indicata la

genotipizzazione di più varianti per la predizione della risposta.

Utilità clinica

Le raccomandazioni riportate nelle più recenti rassegne sistematiche riportano risultati sostenuti da

prove relativamente alla validità clinica. Come già riportato, l’endpoint adottato per l’analisi

dell’associazione tra risposta e genotipo di IL28B è il SVR.

Non sono stati reperiti studi che abbiano valutato l’influenza del genotipo di IL28B su misure

cliniche di esito, come la progressione dell’epatopatia, l’occorrenza di eventi avversi o la gravità di

effetti indesiderati.

Non sono state rilevate linee guida strutturate circa l’utilizzo clinico del test di farmacogenetica,

ovvero raccomandazioni di diversi protocolli di trattamento basati sul risultato del test.

http://www.ncbi.nlm.nih.gov/pubmed?term=Cervinski%20MA%5BAuthor%5D&cauthor=true&cauthor_uid=24462832


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Trial randomizzati sono pertanto necessari per stabilire che protocolli terapeutici basati sul

genotipo di IL28B, da solo o in modelli multivariati con altri predittori, garantisca un migliore esito

clinico6.

Ad oggi, sulla base delle prove di validità clinica disponibili, l’applicazione del test farmacogenetico

può contribuire a determinare, nei pazienti con genotipo virale 1 o 4, le seguenti opzioni.

La durata della terapia:

- PEG-IFN+Ribavirina per 24 settimane nei pazienti con genotipo favorevole CC di IL28B

rs12979860,

- vs 48 settimane di trattamento in pazienti con genotipo CT o TT.

La scelta del protocollo terapeutico:

- triplice terapia [PEG-IFN+Ribavirina associati a boceprevir o telaprevir] nei pazienti con

genotipo sfavorevole di IL28B rs12979860 CT o TT,

- vs duplice terapia [PEG-IFN+Ribavirina] in pazienti con genotipo CC.

Non ci sono indicazioni che il genotipo IL28B influenzi il dosaggio dei farmaci.

Una valutazione contestuale dell’utilità clinica del test farmacogenetico in pazienti con HCV potrà

essere effettuata con il contributo dei diversi attori coinvolti nel percorso clinico: in primo luogo i

prescrittori (epatologi e infettivologi), i laboratori eroganti il test, farmacisti ospedalieri, sentito il

parere dei pazienti e delle famiglie. Un percorso clinico condiviso dovrebbe stabilire:

- criteri di eleggibilità al test farmacogenetico di IL28B [Nota: questo permetterà anche di stimare

prospetticamente il numero di test/anno];

- il tempo di refertazione raccomandato;

- il protocollo terapeutico raccomandato nei pazienti con genotipo favorevole e genotipo

sfavorevole, rispettivamente;

- l’algoritmo terapeutico basato sul test farmacogenetico e su altri eventuali parametri clinici e

biomarker.

Recentemente è stato proposto un modello multivariato che comprende la genotipizzazione IL28B

(rs12979860) e quattro variabili cliniche (carica virale pre-trattamento, rapporto alanina / aspartato

transaminasi, punteggio Ishak per la fibrosi e precedente trattamento con ribavirina) [O’Brien TR

2011]. Gli autori hanno dimostrato che il modello multivariato è più efficace (AUC 78.5%) sia del

modello basato sui soli predittori clinici (AUC 73.0%) sia sul test farmacogenetico da solo (AUC

60.0%).

Un altro modello è stato descritto per pazienti con co-infezione HIV [Medrano J 2010].

Implicazioni etiche, legali e sociali.

La letteratura non riporta studi specifici che abbiano valutato l’accettabilità del test da parte dei

pazienti. Tuttavia si ritiene comunemente che i test genetici finalizzati alla predizione della risposta

al trattamento abbiano minori implicazioni rispetto ai test genetici associati a malattia. Va

sottolineato che questo assunto è valido se il test comprende l’analisi di varianti in un singolo gene,

e se queste non sono associate a altri fenotipi.

Variazioni di IL28B sono state riportate come fattori di rischio per alcuni fenotipi patologici (e.g.:

suscettibilità a cirrosi in infezione da HBV [Xia P et al 2014]). Va ricordato che l’analisi del genotipo

di IL28B è comunque un test genetico su DNA germinale. L’esame va effettuato previo consenso

informato specifico; il parere di un genetista medico deve essere disponibile qualora il paziente o la

6 Nel caso di un altro farmaco antivirale, l’Abacavir, un trial randomizzato sull’utilizzo di un test

farmacogenetico per ottimizzare la terapia ha dimostrato che la genotipizzazione di HLA riduce signifiticamente la frequenza di eventi avversi [Mallal S 2008].


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famiglia richieda informazioni ulteriori [secondo le “Linee guida per le strutture e le attività di

genetica medica”7].

Valutazioni economiche

Non sono disponibili valutazioni economiche o stime di costo-efficacia per il test farmacogenetico.

Conflitti di interesse

Sia gli studi primari, sia le rassegne sistematiche che raccomandano l’effettuazione del test

farmacogenetico (inclusa quella prodotta dal Clinical Pharmacogenetics Implementation

Consortium, [Muir AJ 2013]) riportano possibili conflitti di interessi8. Non sono state reperite

rassegne indipendenti [e.g. Cochrane reviews]. La meta-analisi condotta da Jiménez-Sousa [2013]

non ha rilevato bias di pubblicazione per quanto riguarda i risultati del polimorfismo rs12979860.

Censimento attività in Liguria

Allo scopo di valutare l’offerta sul territorio regionale del test farmacogenetico di IL28B, è stato

eseguito di un censimento delle strutture eroganti. Dall’indagine emerge che i centri che eseguono

il test sono tre, allocati presso:

1. E. O. Ospedali Galliera – Genova (S.C. Laboratorio di istocompatibilità);

2. P.O. Pietra ligure – Albenga (S.S.D Biologia Molecolare);

3. Az.Osp. IRCCS IST-San Martino (Laboratori di Igiene).

Per quanto concerne la tipologia di test si è rilevato che Pietra Ligure-Albenga ha eseguito nel

periodo 2012-2013 il test su due polimorfismi (rs12979860 e rs8099917); San Martino ha eseguito

nel 2013 il solo polimorfismo rs12979860.

Rilevati i centri (coerenti con la DGR 46 /2013), sono stati rilevati i volumi di attività e sono state

effettuate alcune valutazioni per quantificare l’impegno sostenuto sia dal punto di vista tempo-

operatore che da quello organizzativo-strutturale. Sono state pertanto rilevate la tempistica e la

tecnologia necessaria per l’esecuzione del test.

Sono stati riferiti i seguenti volumi di attività:

Galliera: 80 estrazioni nel 2013;

Pietra Ligure-Albenga: 368 nel periodo 2012-2013;

San Martino 350 nel 2013.

Per la metodica basata su RT-PCR è stimabile un tempo totale di circa 3 ore dal check-in del

campione alla refertazione. I tempi di refertazione appaiono quindi dipendenti dal flusso dei

campioni e dall’organizzazione del laboratorio, più che dai tempi analitici.

7 GU n.224 del 23.09.2004

8 La dimostrazione che uno strato di pazienti trattati risponde più favorevolmente può contribuire a

promuovere l’efficacia del farmaco.


12

Nota: se il test farmacogenetico mediante analisi di IL28B viene incluso nell’algoritmo decisionale

per la definizione del protocollo terapeutico, allora va garantita la tempestività della refertazione

entro un intervallo clinicamente ragionevole.

Un’ulteriore indagine potrebbe rilevare quali siano le strutture cliniche che hanno richiesto il test

farmacogenetico di IL28B, e se rientrino tra le strutture abilitate ai sensi della DGR 46 /2013. Il

numero di test erogati potrebbe inoltre essere comparato al numero di pazienti eleggibili

all’indagine, se i criteri di inclusione al test verranno stabiliti da linee guida applicabili localmente.

Discussione

Validità clinica e utilità clinica

C'è una crescente evidenza che i polimorfismi dell’IL28B possano rappresentare un valore

aggiunto nell’algoritmo terapeutico per l’infezione da HCV, quali predittori di un migliore SVR.

Tuttavia, il meccanismo di questa associazione rimane poco chiaro, ed è ancora in fase di studio.

Alcuni autori hanno proposto di incorporare la genotipizzazione di IL28B come criterio per la

definizione dell’iter terapeutico nella pratica clinica; permangono tuttavia alcune criticità,

sottolineate dagli stessi autori – si veda ad esempio l’algoritmo proposto da Holmes JA et al [2011]

sotto riportato, che comprende alcune domande tuttora aperte.

Tratta da: Holmes JA, Desmond PV, Thompson AJ: Redefining baseline demographics: the role of genetic testing in hepatitis C virus infection. Clin. Liver. Dis 2011, 15:497-513.

A conferma della presenza di alcuni margini di incertezza si fa presente che al momento sono in

corso 18 trial clinici9.

9 http://clinicaltrials.gov/

http://clinicaltrials.gov/ct2/results?term=12979860&Search=Search


13

Da quanto emerso nella valutazione dell’utilità clinica del test farmacogenetico di IL28B, le

applicazioni del test all’interno di un percorso clinico meritano ulteriori indagini. In particolare:

- per quali pazienti è indicato il test (tenuto conto del genotipo virale)

- come tale indagine influenza la decisione clinica (in assenza di un algoritmo definito non è

chiaro quale sia il valore clinico dell'informazione)

- se è utile per determinare il protocollo terapeutico, la durata del trattamento, o entrambe le

variabili, o se influenza il dosaggio dei farmaci.

- se il risultato del determina l'inclusione dei pazienti previsti responder, o l'esclusione dei non-

responder (questo aspetto è rilevante anche nella prospettiva dei farmaci IFN-free che non

dipendono da IL28B, ma essendo di costo elevato potrebbero essere indicati come prima linea

per i pazienti non-responder).

Appare quindi auspicabile che venga identificato e possibilmente standardizzato un algoritmo

diagnostico che incorpori la genotipizzazione di IL28B, eventualmente attraverso un modello

multivariato insieme con altri parametri clinici.

Dalle letteratura recente emerge che il modello multivariato che comprende la genotipizzazione

IL28B (rs12979860) combinato con variabili cliniche ha caratteristiche di sensibilità e specificità

significativamente migliori rispetto alla predizione basata sul solo genotipo di IL28B [O’Brien TR

2011].

Erogazione del test nelle strutture di laboratorio della Liguria

Per quanto riguarda l’allocazione dei centri abilitati all’esecuzione della metodica si ritiene che,

vista l’esiguità dei test eseguiti, vi siano possibili margini di razionalizzazione dell’offerta in ottica di

centri deputati e di utilizzo di risorse umane e tecnologiche, previa approfondita analisi delle

diverse attività laboratoristiche effettuate nei centri rilevati.

La metodica analitica viene descritta come robusta e si avvale di una tecnologia consolidata.

Non essendo stato rilevato in letteratura materiale utile a definire la qualità del prodotto finale in

rapporto al numero di test eseguiti, non è possibile esprimere un valore soglia di attività sotto al

quale possa essere messa in dubbio la qualità del test; pertanto un laboratorio potrebbe effettuare

pochi test all’anno senza pregiudicare la qualità del prodotto.

Visto il numero limitato di analisi di IL28B erogate, il costo del test farmacogenetico per singolo

paziente si può approssimare al costo dell’esperimento – in altri termini, il test su un singolo

campione comporta un costo complessivo comparabile a quello del test su pochi campioni, poiché

il costo è determinato prevalentemente da occupazione macchina e tempo uomo.

Per definire quindi la necessità di uno o più centri, si ritiene che la valutazione dovrebbe essere

focalizzata sulla capacità dei singoli laboratori di sostenere volumi di attività superiori a quelli

indicati. In ottica HTA andrebbe valutato il grado di utilizzo reale rispetto alle potenzialità effettive

delle piattaforme utilizzate (performance attuale vs performance teorica). È possibile ritenere che

almeno esista un sottoutilizzo dei dispositivi tecnologici dedicati, che potrebbero esprimere volumi

maggiori di attività attraverso una concentrazione dell’attività su uno o due poli.

Non è stato rilevato in che misura, nelle strutture liguri, il test farmacogenetico di IL28B venga

erogato nell’ambito di sperimentazioni. Si può considerare che la concentrazione dell’attività

permetterebbe una minore dispersione dei casi ed un migliore trattamento dei dati per la

conduzione di studi clinici, nonché di valutazioni che abbiano come obiettivo la stima dei costi

correlati all’esecuzione del test o la definizione dell’algoritmo diagnostico-terapeutico.

Ad integrazione di quanto riportato, si ricorda inoltre che analoghe considerazioni potrebbero

valere per altri aspetti diagnostici correlati all’HCV, ovvero genotipizzazione HCV e rilevazione

HCV-RNA.


14

Bibliografia Cervinski MA, Sam SS, Steinmetz HB, Wood B, Tsongalis GJ. Validation of interleukin 28B

genotyping assay for clinical use. Clinical Biochemistry (0), 2014. Ge D, Fellay J, Thompson AJ, Simon JS, Shianna KV, Urban TJ, Heinzen EL, Qiu P, Bertelsen

AH, Muir AJ, Sulkowski M, McHutchison JG, Goldstein DB. Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance. NATURE 461(7262):399-401, 2009.

Ghany MG, Nelson DR, Strader DB, Thomas DL, Seeff LB. An update on treatment of genotype 1 chronic hepatitis C virus infection: 2011 practice guideline by the American Association for the Study of Liver Diseases. Hepatology 54(4):1433-1444, 2011.

Ghany MG, Strader DB, Thomas DL, Seeff LB. Diagnosis, management, and treatment of hepatitis C: An update. Hepatology 49(4):1335-1374, 2009.

Gupta AC, Trehanpati N, Sukriti S, Hissar S, Midha V, Sood A, Sarin SK. Interleukin-28b CC genotype predicts early treatment response and CT/TT genotypes predicts non-response in patients infected with HCV genotype 3. Journal of Medical Virology 86(4):707-712, 2014.

Imran M, Manzoor S, Ashraf J, Khalid M, Tariq M, Khaliq HM, Azam S. Role of viral and host factors in interferon based therapy of hepatitis C virus infection. Virol J 10:299, 2013.

Jia Z, Ding Y, Tian S, Niu J, Jiang J. Test of IL28B polymorphisms in chronic hepatitis C patients treated with PegIFN and ribavirin depends on HCV genotypes: results from a meta-analysis. PLoS ONE 7(9):e45698, 2012.

Jimenez-Sousa MA, Fernandez-Rodriguez A, Guzman-Fulgencio M, Garcia-Alvarez M, Resino S. Meta-analysis: implications of interleukin-28B polymorphisms in spontaneous and treatment-related clearance for patients with hepatitis C. BMC Med 11:6, 2013.

Kowdley KV, Lawitz E, Poordad F, Cohen DE, Nelson DR, Zeuzem S, Everson GT, Kwo P, Foster GR, Sulkowski MS, Xie W, Pilot-Matias T, Liossis G, Larsen L, Khatri A, Podsadecki T, Bernstein B. Phase 2b Trial of Interferon-free Therapy for Hepatitis C Virus Genotype 1. New England Journal of Medicine 370(3):222-232, 2014.

Kronenberger B and Zeuzem S. Current and future treatment options for HCV. Ann Hepatol 8(2):103-112, 2009.

Lindh M, Arnholm B, Bjorkman P, Hellstrand K, Lagging M, Nilsson S, Wahlberg T, Wallmark E, Weiland O, Wejstal R, Westin J, Widell A, Norkrans G. Dynamic tailoring of treatment durations improves efficiency of hepatitis C treatment with pegylated interferon and ribavirin. Journal of Viral Hepatitis 20(4):e82-e89, 2013.

Lopez-Cortes LF, Ruiz-Valderas R, Jimenez-Jimenez L, Gonzalez-Escribano MF, Torres-Cornejo A, Mata R, Rivero A, Pineda JA, Marquez-Solero M, Viciana P. Influence of IL28B polymorphisms on response to a lower-than-standard dose peg-IFN-alpha 2a for genotype 3 chronic hepatitis C in HIV-coinfected patients. PLoS ONE 7(1):e28115, 2012.

Mallal S, Phillips E, Carosi G, Molina JM, Workman C, Tomazic J, Jagel-Guedes E, Rugina S, Kozyrev O, Cid JF, Hay P, Nolan D, Hughes S, Hughes A, Ryan S, Fitch N, Thorborn D, Benbow A. HLA-B*5701 screening for hypersensitivity to abacavir. The New England Journal of Medicine 358(6):568-579, 2008.

Medrano J, Neukam K, Rall+¦n N, Rivero A, Resino S, Naggie S, Caruz A, Calvino A, Mac+¡as J, Benito JM, S+ínchez-Piedra C, Vispo E, Barreiro P, McHutchison J, Pineda JA, Soriano V. Modeling the Probability of Sustained Virological Response to Therapy with Pegylated Interferon plus Ribavirin in Patients Coinfected with Hepatitis C Virus and HIV. Clinical Infectious Diseases 51(10):1209-1216, 2010.

Muir AJ, Gong L, Johnson SG, Lee MTM, Williams MS, Klein TE, Caudle KE, Nelson DR. Clinical Pharmacogenetics Implementation Consortium (CPIC) Guidelines for IFNL3 (IL28B) Genotype and PEG Interferon-[alpha]-Based Regimens. Clin Pharmacol Ther , 2013 .

O'Brien TR, Everhart JE, Morgan TR, Lok AS, Chung RT, Shao Y, Shiffman ML, Dotrang M, Sninsky JJ, Bonkovsky HL, Pfeiffer RM, and the HALT-. An IL28B Genotype-Based Clinical Prediction Model for Treatment of Chronic Hepatitis C. PLoS ONE 6(7):e20904, 2011.

Thomas DL, Thio CL, Martin MP, Qi Y, Ge D, 'hUigin C, Kidd J, Kidd K, Khakoo SI, Alexander G, Goedert JJ, Kirk GD, Donfield SM, Rosen HR, Tobler LH, Busch MP, McHutchison JG, Goldstein DB, Carrington M. Genetic variation in IL28B and spontaneous clearance of hepatitis C virus. NATURE 461(7265):798-801, 2009.


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Xia P, Zhou M, Dong D, Xing YN, Bai Y. Association of polymorphisms in interleukin-18 and interleukin-28B genes with outcomes of hepatitis B virus infections: a meta-analysis. Tumor Biol 35(2):1129-1137, 2014.


16

Appendice

1. Reference List: Systematic Reviews & Meta-analyses

1. Bota, Simona, Sporea, Ioan, Sirli, Roxana, Neghina, AdrianaMaria, Popescu, Alina, and Strain, Mihnea. Role of Interleukin-28B Polymorphism as a Predictor of Sustained Virological Response in Patients with Chronic Hepatitis C Treated with Triple Therapy: A Systematic Review and Meta-Analysis. Clin Drug Investig 33[5], 325-331. 2013. Abstract: Background and Objective: Chronic hepatitis C represents an important health problem. The aim of our meta-analysis was to establish the role of reference single nucleotide (rs) 12979860 allele of interleukin-28B (IL28B) CC versus CT+TT genotype (the most researched allele of IL28B) as a predictor of sustained virological response (SVR) in patients with chronic hepatitis C treated with triple therapy. Methods: The PubMed, MEDLINE, Lilacs, Scopus, Ovid, EMBASE, Cochrane and Medscape databases as well as abstract books from important gastroenterology and hepatology meetings were searched for all studies published until 15 July 2012 that analysed the relationship between the polymorphism of IL28B and SVR in patients with chronic hepatitis C, genotype 1, treated with pegylated interferon + ribavirin + direct antiviral agents (telaprevir or boceprevir). The following keywords were used: IL28B polymorphism, chronic hepatitis C, sustained virological response, SVR, triple therapy, telaprevir, boceprevir. Results: Odds ratios (ORs) with 95 % confidence intervals were pooled from five study populations (1,641 cases) using a random–effects model. The SVR rate was significantly higher in patients with the CC genotype of IL28B than in those with non-CC genotypes (CT and TT): OR = 3.91 (95 % CI 2.11–7.28), p < 0.0001. Higher SVR rates were obtained in chronic hepatitis C patients with the CC genotype of IL28B, regardless of their therapeutic status (naïve patients: OR = 3.99 [95 % CI 1.67–9.51], p < 0.0001; and previously treated ones: OR = 2.15 [95 % CI 1.35–3.43], p = 0.001). Conclusion: IL28B polymorphism seems to influence the SVR rate in patients with chronic hepatitis C treated with triple therapy, but further studies are needed to clarify the mechanism and the influence of other factors on the SVR rates.

2. Chen, Y., Xu, H. X., Wang, L. J., Liu, X. X., Mahato, R. I., and Zhao, Y. R. Meta-analysis: IL28B polymorphisms predict sustained viral response in HCV patients treated with pegylated interferon-alpha and ribavirin. Aliment.Pharmacol Ther 36[2], 91-103. 2012. Abstract: BACKGROUND: Interleukin (IL) 28B single nucleotide polymorphisms can predict sustained virological response (SVR) in hepatitis C virus (HCV) patients following pegylated interferon-alpha (PEG IFN-alpha) and ribavirin treatment. AIM: To design a meta-analysis to determine IL28B genotypes', rs12979860 CC and rs8099917 TT, correlation with SVR in PEG IFN-alpha/ribavirin-treated HCV patients. METHODS: Meta-analysis was performed in 17 studies of rs12979860 CC vs. CT/TT and 17 of rs8099917 TT vs. TG/GG. Odds ratios (OR) and confidence intervals (95% CI) were calculated by fixed- or random-effects models. Heterogeneity, sensitivity analysis and publication bias were also assessed. RESULTS: Of 4252 Asian, Caucasian and African HCV patients analysed for rs12979860, SVR was more frequent in CC (vs. CT/TT; OR = 4.76, 95% CI: 3.15-7.20). Moreover, CC was associated with SVR for HCV genotype-1 or -4 infections (OR(genotype 1) = 5.52, 95% CI: 3.74-8.15; OR(genotype 4) = 8.11, 95% CI: 4.13-15.93), regardless of ethnicity. Of 4549 Caucasian and Asian HCV patients analysed for rs8099917, SVR was more frequent in TT (vs. TG/GG; OR = 3.31, 95% CI: 2.39-4.59). Moreover, TT was associated with SVR for HCV-1 (OR(genotype 1) = 4.28, 95% CI: 2.87-6.38). Rs8099917 TT predictive value was stronger in Asians (OR(Asians) = 8.09, 95% CI: 5.63-11.61; OR(Caucasians) = 3.00, 95% CI: 2.03-4.45). Ethnicity stratification revealed that rs8099917 TT had slight predictive value in Asian HCV-2/3 patients (OR = 1.99, 95% CI: 1.09-3.62). CONCLUSIONS: IL28B rs12979860 CC and rs8099917 TT are strong SVR predictors for PEG IFN-alpha/ribavirin-treated HCV-1 patients, regardless of ethnicity. In HCV-2/3, rs12979860 CC has no SVR predictive value, but rs8099917 TT was slightly associated with SVR in Asians

3. Jia, Z., Ding, Y., Tian, S., Niu, J., and Jiang, J. Test of IL28B polymorphisms in chronic hepatitis C patients treated with PegIFN and ribavirin depends on HCV genotypes: results from a meta-analysis. PLoS ONE 7[9], e45698. 2012. Abstract: BACKGROUND: Many studies have been published on the association between single nucleotide polymorphisms (SNP) near the IL28B gene and response to the combined treatments of pegylated-interferon (PegIFN) and ribavirin (RBV) in chronic HCV-infected patients, but without identical conclusions. The aim of this study was to assess impact of the IL28B polymorphisms on the effect of HCV standard treatment using meta-analysis based method. METHODS: Association studies between polymorphisms of rs12979860 or rs8099917 and response to PegIFN/RBV treatment in chronic HCV patients were retrieved from PubMed. Data of qualified studies on sustained virological response (SVR) in different genotypes were extracted and analyzed using meta-analysis method in Stata 10 software. RESULTS: Thirty-four papers, containing 46 independent studies, were included in the analysis. In the HCV G1/4 patients without treatment history, individuals carrying rs12979860 CC genotype were more likely to achieve SVR (OR 3.97, 95%CI 3.29-4.80) compared to those carrying CT/TT genotypes. Similar results were observed in the HCV G1/4 patients with unsuccessful or unknown treatment history (OR 3.76, 95%CI 2.67-5.28) or in the patients co-infected with human immunodeficiency virus (OR 5.20, 95%CI 3.04-8.90). However, associations could not be observed in HCV G2/3 patients. For rs8099917, similar results were obtained for genotype TT compared


17

to genotypes TG/GG, indicating that TT genotype was significantly associated with better treatment response in patients infected with genotype 1 or 4 HCV, but not genotype 2 or 3 HCV. CONCLUSION: Polymorphisms of rs12979860 and rs8099917 near IL28B only associate with the treatment response to PegIFN/RBV in patients infected with HCV genotype 1 or 4 but not with genotype 2 or 3, irrespective of the previous treatment history or HIV co-infected status. Therefore, identification of IL28B genotypes is necessary only in patients infected with relatively difficult-to-treat genotype 1 or 4 HCV

4. Jimenez-Sousa, M. A., Fernandez-Rodriguez, A., Guzman-Fulgencio, M., Garcia-Alvarez, M., and Resino, S. Meta-analysis: implications of interleukin-28B polymorphisms in spontaneous and treatment-related clearance for patients with hepatitis C. BMC Med 11, 6. 2013. Abstract: BACKGROUND: Since 2009, several studies have identified single-nucleotide polymorphisms (SNPs) near the gene encoding for interleukin (IL)-28 (IL28B) that are strongly associated with spontaneous and treatment-induced hepatitis C virus (HCV) clearance. Because this large amount of data includes some inconsistencies, we consider assessment of the global estimate for each SNP to be essential. METHODS: Relevant studies assessing IL28B polymorphisms associated with sustained virologic response (SVR) and spontaneous clearance (SC) were identified from a literature search of PubMed up to 9 July, 2012. Studies were eligible studies if they included patients infected with HCV or HCV/HIV, or assessed any SNP located within or near the IL28B gene, SVR data available under standard treatment, and/or SC data in patients with acute HCV infection. Pooled odds ratios were estimated by fixed or random effects models when appropriate. Variables such as HCV genotype, ethnicity, and type of co-infection were studied. RESULTS: Of 282 screened studies, 67 were selected for SVR and 10 for SC. In total, 20,163 patients were studied for SVR and 3,554 for SC. For SVR, we found that all SNPs showed strong associations in patients with HCV genotypes 1 and 4, whereas the pooled ORs were almost three times lower for genotypes 2 and 3 (rs12979860 and rs8099917). Regarding ethnicity, the SNP most associated with SVR was rs12979860 in white patients, whereas in East Asians it seemed to be rs8099917. The most studied SNP (rs12979860) showed similar results for patients co-infected with HCV/HIV, as for those infected with HCV only. Finally, rs12979860 and rs8099917 both appeared to be associated with SC. CONCLUSIONS: IL28B polymorphisms influence both the outcome of interferon treatment and the natural clearance of HCV. However we did not identify a universal predictor SNP, as the best genetic markers differed depending on patient ethnicity, genotype, and type of infection. Nevertheless, our results may be useful for more precise treatment decision-making

5. Li, S., Hu, P., Zhang, Q. Q., Liu, Y. H., Hu, H. D., Zhang, D. Z., and Ren, H. Single nucleotide polymorphisms of the IL28B and sustained virologic response of patients with chronic hepatitis C to PEG-interferon/ribavirin therapy: A meta-analysis: Meta-analysis of IL28B. Hepat Mon. 11[3], 163-172. 2011. Abstract: BACKGROUND: Hepatitis C is a global health problem and represents a major cause of liver disease and socioeconomic burden. Effective antiviral therapy may prevent these complications, but the current treatment for patients with chronic hepatitis C virus (HCV) infection does not produce sustained virologic response. Therefore, identification of the determinants of response to treatment is a high priority. A number of host and viral factors have been associated with treatment outcomes. OBJECTIVES: To assess the associations of single nucleotide polymorphisms (SNP) of the IL28B and sustained virologic response (SVR) of patients with chronic hepatitis C to PEG-interferon/ribavirin therapy. MATERIALS AND METHODS: We searched PubMed, Medline and Cochrane Library, and found 7 eligible papers involved in this study. Then we performed a meta-analysis comparing the SVR rate at SNP of the IL28B in individuals with PEG-interferon/ribavirin therapy. Meanwhile, the SVR rate between different races and HCV genotypes was studied. RESULTS: The sustained virologic response rate was higher in patients with the rs12979860 CC and rs8099917 TT alleles in the IL28B SNP, comparing with the rs12979860 CT, or TT and rs8099917 TG or GG. Furthermore, a higher SVR was observed in the Caucasians than in Afro-Americans (OR = 3.85, 95% CI: 3.06-4.83); the percentage of rs12979860 TT genotype was lower in Caucasians (OR = 0.25, 95% CI: 0.20-0.31) and the percentage of rs12979860 CC genotype was higher in Caucasians than that of Afro-Americans (OR = 3.45, 95% CI = 2.68-4.44). Between different HCV genotypes, the SVR was much lower in those with HCV genotype 1 than those with genotype 2/3 (OR = 0.16, 95% CI: 0.11-0.24). CONCLUSIONS: IL28B is significantly associated with response to PEG-interferon/ribavirin therapy of patients with chronic HCV infection. Both the rs12979860 and rs8099917 alleles could be used as independent predictors of the treatment response. The rs12979860 allele in particular, is more important from our study. The polymorphism even explains part the difference in response rate between different ethnic groups and HCV genotypes

6. Luo, Y., Jin, C., Ling, Z., Mou, X., Zhang, Q., and Xiang, C. Association study of IL28B: rs12979860 and rs8099917 polymorphisms with SVR in patients infected with chronic HCV genotype 1 to PEG-INF/RBV therapy using systematic meta-analysis. GENE 513[2], 292-296. 25-1-2013. Abstract: Recently, genome-wide associated studies (GWAS) have identified that host genetics IL28B SNPs rs12979860 and rs8099917 were significantly associated with SVR in patients infected with chronic HCV genotype 1 to PEG-INF/RBV therapy. Results from these studies remain conflicting. We conducted this meta-analysis to estimate the overall association of SVR with rs12979860 and rs8099917. We searched the PubMed, Embase, Scholar Google, ISI Web of Knowledge, and Chinese National Knowledge Infrastructure (CNKI) databases for all articles before July 30, 2012. The odds ratio (OR) corresponding to the 95% confidence interval (CI) was used to assess the association. The statistical heterogeneity among studies was assessed with the I(2) statistics. Begg's test and Egger's test were performed to evaluate the publication bias. Eventually, twenty studies were selected for the meta-


18

analysis. The IL-28B SNPs rs12979860 genotype CC and rs8099917 genotype TT significantly positive associated with SVR in patients infected chronic HCV genotype 1 to PEG-INF/RBV therapy (OR=4.473, 95% CI=3.814-5.246, OR=5.171, 95% CI=4.372-6.117 respectively). The results suggested that rs12979860 genotype CC and rs8099917 genotype TT could be used as independent predictors of the HCV-1 infected patients

7. Muir, A. J., Gong, L., Johnson, S. G., Lee, M. T. M., Williams, M. S., Klein, T. E., Caudle, K. E., and Nelson, D. R. Clinical Pharmacogenetics Implementation Consortium (CPIC) Guidelines for IFNL3 (IL28B) Genotype and PEG Interferon-[alpha]-Based Regimens. Clin Pharmacol Ther . 13-11-2013. Abstract: Pegylated interferon-[alpha] (PEG-IFN-[alpha] or PEG-IFN 2a and 2b)- and ribavirin (RBV)-based regimens are the mainstay for treatment of hepatitis C virus (HCV) genotype 1. IFNL3 (IL28B) genotype is the strongest baseline predictor of response to PEG-IFN-[alpha] and RBV therapy in previously untreated patients and can be used by patients and clinicians as part of the shared decision-making process for initiating treatment for HCV infection. We provide information regarding the clinical use of PEG-IFN-[alpha]- and RBV-containing regimens based on IFNL3 genotype

8. Rangnekar, A. S. and Fontana, R. J. IL-28B polymorphisms and the response to antiviral therapy in HCV genotype 2 and 3 varies by ethnicity: a meta-analysis. Journal of Viral Hepatitis 20[6], 377-384. 1-6-2013. Abstract: Studies of IL-28B genotype in patients with hepatitis C virus (HCV) genotype 2/3 infection have yielded conflicting results. The aim of this meta-analysis was to obtain a pooled odds ratio (OR) of the impact of IL-28B genotype on achieving sustained virologic response (SVR) in patients with HCV genotype 2/3 infection treated with pegIFN and ribavirin. A meta-analysis with a random effects model was performed, and study heterogeneity and publication bias were assessed. Forty-three percent of the Caucasians (11 studies) and 86% of Asians (five studies) had the favourable IL-28B genotype. In Caucasians, the pooled OR of SVR with the favourable IL-28B genotype was

1.36 (95%CI: 0.98Çô1.88, P-á=-á0.07) in all patients and 1.55 (95%CI: 1.10Çô2.18, P-á=-á0.01) in patients

treated with pegIFN and ribavirin for ëÑ24-áweeks. In Asians, the pooled OR of SVR in patients with the favourable

IL-28B genotype was 1.99 (95%CI: 0.94Çô4.25, P-á=-á0.07). The favourable IL-28B genotype was also significantly

associated with rapid virologic response (RVR) in both groups (Caucasians: OR: 1.82, 95%CI: 1.12Çô2.96, P-á=-

á0.02; Asians: 2.39, 95%CI: 1.39Çô4.11, P-á=-á0.002), as well as the likelihood of an SVR in a subgroup of 350

Caucasian patients without an RVR (OR: 3.29, 95%CI: 1.67Çô6.51, P-á=-á0.001). The favourable IL-28B genotype is a statistically significant predictor of SVR and RVR in Caucasian patients treated with pegIFN and ribavirin for 24-áweeks. In contrast, the favourable IL-28B genotype is associated with RVR, but not SVR in Asian HCV genotype 2 patients

9. Rangnekar, A. S. and Fontana, R. J. Meta-analysis: IL-28B genotype and sustained viral clearance in HCV genotype 1 patients. Aliment.Pharmacol Ther 36[2], 104-114. 2012. Abstract: BACKGROUND: Polymorphisms in the IL-28B region are a strong predictor of sustained virologic response (SVR) in individual studies of HCV genotype 1 patients receiving peginterferon (pegIFN) and ribavirin. AIM: To obtain a pooled odds ratio (OR) of SVR in patients of varying race with the favourable IL-28B genotype compared to those with the unfavourable genotype. METHODS: A literature search was conducted using online databases and a review of conference abstracts. A random effects meta-analysis was performed and study heterogeneity and publication bias were assessed. RESULTS: There were 21 individual studies of HCV genotype 1 patients of varying ethnicity treated with pegIFN and ribavirin. The pooled prevalence of the favourable IL-28B genotype varied by race (73% vs. 41% vs. 13% in 2612 Asians, 3110 Caucasians and 452 African-Americans, respectively, P < 0.001). However, the strength of association of the IL-28B genotype with SVR was similar in all three racial groups (Caucasians: odds ratio (OR) 3.88, 2.75-5.49, African-Americans: OR 4.63, 2.52-8.50 and Asians OR 5.66, 3.99-8.02, all P < 0.001). The IL-28B genotype was also associated with SVR in 263 HIV/HCV co-infected Caucasians (OR 5.49, 3.02-9.96, P < 0.001). Study quality score and anti-viral treatment regimen did not impact the strength of the association in patient subgroups nor in the pooled population. CONCLUSIONS: IL-28B genotype is significantly associated with SVR in HCV genotype 1 patients of varying race, as well as in HIV co-infected patients, receiving pegIFN and ribavirin. IL-28B testing in conjunction with other pre-treatment parameters may prove useful in counselling HCV patients


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Appendice

2. Reference List: Primary studies 1. Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained

virologic response in genotype 1 hepatitis C virus. Thompson AJ et al. Gastroenterology. 2010;139(1):120. AIMS: We recently identified a polymorphism upstream of interleukin (IL)-28B to be associated with a 2-fold difference in sustained virologic response (SVR) rates to pegylated interferon-alfa and ribavirin therapy in a large cohort of treatment-naive, adherent patients with chronic hepatitis C virus genotype 1 (HCV-1) infection. We sought to confirm the polymorphism's clinical relevance by intention-to-treat analysis evaluating on-treatment virologic response and SVR. METHODS: HCV-1 patients were genotyped as CC, CT, or TT at the polymorphic site, rs12979860. Viral kinetics and rates of rapid virologic response (RVR, week 4), complete early virologic response (week 12), and SVR were compared by IL-28B type in 3 self-reported ethnic groups: Caucasians (n = 1171), African Americans (n = 300), and Hispanics (n = 116). RESULTS: In Caucasians, the CC IL-28B type was associated with improved early viral kinetics and greater likelihood of RVR (28% vs 5% and 5%; P<.0001), complete early virologic response (87% vs 38% and 28%; P<.0001), and SVR (69% vs 33% and 27%; P<.0001) compared with CT and TT. A similar association occurred within African Americans and Hispanics. In a multivariable regression model, CC IL-28B type was the strongest pretreatment predictor of SVR (odds ratio, 5.2; 95% confidence interval, 4.1-6.7). RVR was a strong predictor of SVR regardless of IL-28B type. In non-RVR patients, the CC IL-28B type was associated with a higher rate of SVR (Caucasians, 66% vs 31% and 24%; P<.0001). CONCLUSIONS: In treatment-naive HCV-1 patients treated with pegylated interferon and ribavirin, a polymorphism upstream of IL-28B is associated with increased on-treatment and sustained virologic response and effectively predicts treatment outcome.

2. Replicated association between an IL28B gene variant and a sustained response to pegylated interferon and ribavirin. McCarthy JJ, Li JH, Thompson A, Suchindran S, Lao XQ, Patel K, Tillmann HL, Muir AJ, McHutchison JG. Gastroenterology. 2010;138(7):2307. AIMS: Patients with chronic hepatitis C virus (HCV) infections are treated with pegylated interferon and ribavirin (PEG-IFN/RBV), which is effective in less than 50% of those infected with HCV genotype 1. Genome-wide association studies have linked response to PEG-IFN/RBV with common single nucleotide polymorphisms in the vicinity of interferon (IFN)-lambda genes on chromosome 19. We investigated the association between the polymorphism rs12979860 and treatment response in a diverse cohort of chronic HCV patients. METHODS: A cross-sectional study of 1021 consecutive patients enrolled in the Duke Hepatology Clinic Research Database and Biorepository. We analyzed DNA, clinical and demographic data, along with validated data of the response of 231 subjects to PEG-IFN/RBV. The study included Caucasians (n = 178), African Americans (n = 53), and HCV genotypes 1 (n = 186) and 2/3 (n = 45). The rs12979860 genotype was tested for an association with sustained virologic response, defined as undetectable levels of HCV RNA 24 weeks after treatment ended. RESULTS: The rs12979860 CC genotype (found in approximately 40% of Caucasians) predicted a sustained virologic response to therapy among Caucasians (odds ratio, 5.79; 95% confidence interval, 2.67-12.57; P = 9.0 x 10(-6)), independent of HCV genotype and other covariates. Rs12979860 CC predicted a sustained response with 78% specificity and 65% sensitivity in patients infected with HCV genotype 1). CONCLUSIONS: rs12979860 genotype is a significant independent predictor of response to PEG-IFN/RBV in patients with chronic HCV infection; tests for this genotype might be used to determine the best course of treatment for patients considering antiviral therapy.

3. An IL28B polymorphism determines treatment response of hepatitis C virus genotype 2 or 3 patients who do not achieve a rapid virologic response. Mangia A, Thompson AJ, Santoro R, Piazzolla V, Tillmann HL, Patel K, Shianna KV, Mottola L, Petruzzellis D, Bacca D, Carretta V, Minerva N, Goldstein DB, McHutchison JG. Gastroenterology. 2010;139(3):821. AIMS: Polymorphisms in the region of the interleukin (IL)-28B gene on chromosome 19 have been associated with peginterferon-alfa-induced clearance of genotype 1 hepatitis C virus (HCV); there are no data for patients with genotype 2 or 3 HCV. We evaluated the effects of IL-28B polymorphisms on response to treatment with peginterferon and ribavirin in a well-characterized cohort of genotype 2/3 patients. METHODS: DNA was analyzed from 268 patients (Caucasian: genotype 2, 213; genotype 3, 55). Patients were randomly assigned to groups that received standard duration (24 wk; n = 68) or variable durations of therapy. Patients who received variable durations (VD) and had a rapid virologic response (RVR) were treated for 12 weeks (VD12; n = 122); those without an RVR were treated for 24 weeks (VD24; n = 78). IL-28B genotypes (rs12979860) were analyzed for association with treatment response. RESULTS: The frequencies of the IL-28B genotypes were as follows: CC, 37%; CT, 48%; and TT, 15%; 82% of patients with the CC genotype achieved a sustained virologic response (SVR), compared with 75% with the CT and 58% with the TT genotypes (P = .0046). Differences between IL-28B genotypes were greatest among patients who failed to attain RVR (VD24 SVR rates: CC, 87%; CT, 67%; and TT, 29%; P = .0002). Among patients with RVRs (61%), the IL-28B genotype was not associated with SVR (>70% for all IL-28B genotypes). In a multivariable logistic regression model, IL-28B genotype predicted SVR (odds ratio, 1.76; 95% confidence interval, 1.16-2.7). CONCLUSIONS: An IL-28B polymorphism was associated with an SVR in patients infected with genotype 2/3 HCV who did not achieve a RVR. Analysis of IL-28B genotype might be used to guide treatment for these patients.

4. Interleukin-28B genetic variants and hepatitis virus infection by different viral genotypes. Montes-Cano MA, García-Lozano JR, Abad-Molina C, Romero-Gómez M, Barroso N, Aguilar-Reina J, Núñez-Roldán A, González-Escribano MF. Hepatology. 2010;52(1):33. Genetic host factors may modify the course of the hepatitis C virus (HCV) infection.


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Very recently, a genome-wide scan that reported association of the IL28B locus with response to treatment in HCV infection was published. The aim of the current study was to investigate the relationship of this locus with outcome of HCV infection in a cohort constituted by a total of 731 Spanish individuals. From these, 284 were subjects with persistent infection, 69 were individuals who naturally cleared the virus, and 378 were noninfected subjects. Genotyping of the rs12979860 (C>T) in the IL28B locus was performed using a TaqMan 5' allelic discrimination assay. The CC genotype was overrepresented among patients infected with viral genotypes non-1 (66.7% versus 39.1% in patients infected with viral genotype-1, P = 8.5 x 10(-5), odds ratio [OR]= 0.32, 95% confidence interval [CI]0.17-0.60); patients with spontaneous resolution of infection (72.5% versus 45.6% of the individuals with persistent infection, P = 6.2 x 10(-5), OR = 0.32; 95%CI, 0.18-0.57); and lastly, patients with sustained response (60.2% versus 32.1% found in patients with nonsustained response, P = 3.1 x 10(-5), OR = 0.31; 95%CI, 0.17-0.56). CONCLUSION: We have found different rates of viral genotype infection depending on the IL28B variant as well as an association of this locus with natural and treatment-mediated response.

5. IL28B genetic variation and treatment response in patients with hepatitis C virus genotype 3 infection. Moghaddam A, Melum E, Reinton N, Ring-Larsen H, Verbaan H, Bjøro K, Dalgard O. Hepatology. 2011;53(3):746.Polymorphisms nea r the IL28B gene, which code for interferon (IFN)-λ3, predict response to pegylated interferon-α(PEG-IFN) and ribavirin treatment in hepatitis C virus (HCV) genotype 1 infected patients. Follow-up studies of the effect of IL28B gene in HCV non-genotype 1 infected patients have almost always used predominantly HCV genotype 2-infected or mixed genotype 2/3-infected cohorts with results partly conflicting with HCV genotype 1. We performed a retrospective analysis of 281 patients infected with HCV genotype 3 for association of response to therapy with IL28B polymorphisms. We found that the HCV genotype 1 responder genotypes at rs12979860 and rs8099917 did not associate with sustained virological response to PEG-IFN/ribavirin therapy. However, the responder genotypes of both SNPs showed association with rapid viral response measured at 4 weeks (rs12979860, P = 3×10(-5) ; rs8099917, P = 3×10(-4) ). In multivariate analysis, age (<40 years), baseline viral load (<4×10(5) IU/mL) and the responder genotypes of SNPs rs12979860 or rs8099917 remained significant independent predictors of rapid viral response to therapy. Furthermore, we show that IL28B polymorphisms are associated with relapse in patients who achieve rapid viral response to PEG-IFN/ribavirin therapy. The responder genotypes also showed association with markers of stage and activity of liver disease, namely high aspartate aminotransferase platelet ratio index (APRI, rs12979860, P = 0.018; rs8099917, not significant) and high alanine aminotransferase (ALT, rs12979860, P = 0.002; rs8099917, P = 0.001), in addition to a high baseline viral load (rs12979860, P = 1.4×10(-5) ; rs8099917, P = 7.3×10(-6) ). CONCLUSION: Polymorphisms near the IL28B gene show association with rapid viral response but not sustained viral response to PEG-IFN/ribavirin therapy in HCV genotype 3-infected patients.

6. Interleukin 28B polymorphism predicts pegylated interferon plus ribavirin treatment outcome in chronic hepatitis C genotype 4. De Nicola S, Aghemo A, Rumi MG, Galmozzi E, Valenti L, Soffredini R, De Francesco R, Prati GM, D'Ambrosio R, Cheroni C, Donato MF, Colombo M. Hepatology. 2012;55(2):336. Single nucleotide polymorphisms (SNPs) near the interleukin 28B (IL28B) region are the strongest baseline predictors of a sustained virologic response (SVR) to peg-interferon (PegIFN) and ribavirin (Rbv) in patients with hepatitis C virus (HCV) genotype 1 infection. Whether this holds true for HCV-4 patients too is unknown. The aim was to investigate the predictive power of the rs12979860 IL28B SNP for a response to Peg-IFN and Rbv in HCV-4 patients. All HCV-4 patients consecutively treated between September 2004 and June 2010 with PegIFN and Rbv at two liver centers at the Maggiore Hospital Milan (Italy) underwent TaqMan SNP Genotyping assays for testing rs12979860 genotype. Of 112 treated patients (98 males, 75 of Egyptian descent, 26 with cirrhosis) 103 were included in the final analysis; five discontinued treatment for nonvirologic reasons and four did not consent to genetic testing. Twenty-four (23%) were genotype CC, 65 (63%) CT, and 14 (14%) TT. Overall, 50 (49%) achieved an SVR: 21 (88%) CC patients versus 29 (37%) CT/TT (P<0.0001). CC patients more often had a rapid virologic response (RVR) than CT/TT patients (12, 50% versus 23, 29%; P = 0.08), while also showing lower relapse rates (0% [0/21]versus 36% [16/45]P = 0.0013). In non-RVR patients, SVR rates were higher in CC than CT/TT patients (9 [75%]versus 13 [23%]P = 0.001). By logistic regression, the IL28B rs12979860 CC genotype was an independent predictor of SVR with an odds ratio of 8.0 (95% confidence interval 2.00-32.01; P = 0.003). CONCLUSION: The IL28B rs12979860 SNP may have an added value in the treatment algorithm of HCV-4 patients because it is the strongest predictor of an SVR to PegIFN/Rbv therapy.

7. IL28B polymorphism is associated with treatment response in patients with genotype 4 chronic hepatitis C. Asselah T, De Muynck S, Broët P, Masliah-Planchon J, Blanluet M, Bièche I, Lapalus M, Martinot-Peignoux M, Lada O, Estrabaud E, Zhang Q, El Ray A, Vidaud D, Ripault MP, Boyer N, Bedossa P, Valla D, Vidaud M, Marcellin P. J Hepatol. 2012;56(3):527. AIMS: Polymorphisms in the region of the interleukin (IL)28B gene have been associated with pegylated-interferon (PEG-IFN) and ribavirin treatment response mainly in genotype 1 HCV infections. However, there are few data on HCV genotype 4 (HCV-4) infection. We evaluated, in a unique well-characterized cohort of HCV-4 patients, the association of IL28B polymorphism with response to treatment or liver disease severity. METHODS: This study included 164 HCV-4 patients from different ethnic groups (Egyptian, European, and Sub-Saharan African). Among these patients, 82 were studied for response and 160 for disease severity. Free DNA extracted from all the 164 patient's serum samples was analyzed by direct sequencing of the SNP rs12979860 of IL28B. Genetic and bio-clinical features from patients having sustained virological response (43 SVR patients) and from those who did not respond to treatment or had a relapse after the end of the treatment (39 NR patients) were compared. IL28B polymorphism was compared between the 78 patients with mild fibrosis (Metavir score F0-F1) and the 82 with advanced fibrosis (F2-F4). RESULTS: Our data showed a better treatment response rate of the C allele of the IL28B gene SNP rs12979860 (p=0.0008). The response rates were 81.8%, 46.5%, and 29.4% for genotype CC, CT, and TT, respectively. No significant relationship was found between rs12979860 and the severity of the disease.


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CONCLUSIONS: The SNP rs12979860 is strongly associated with SVR in patients infected with HCV-4, but not with liver disease severity. Analysis of IL28B genotype might be used to guide treatment for these patients.

8. Impact of IL28B genotype on the early and sustained virologic response in treatment-naïve patients with chronic hepatitis C. Stättermayer AF, et al. Clin Gastroenterol Hepatol. 2011;9(4):344. AIMS: Single nucleotide polymorphisms (SNPs) in the gene that encodes interleukin (IL)-28B predict response of patients with chronic hepatitis C to antiviral therapy. We investigated the roles of polymorphisms rs12979860 and rs8099917 on the early virologic response of treatment-naïve patients. METHODS: SNPs were identified by real-time polymerase chain reaction analysis of samples from 682 patients (genotype [GT]1=372, GT2/3=208, GT4=102) who were treated with 180μg pegylated interferon-α2a and 400 or 800 mg (GT2/3, depending on the protocol) or 1000-1200 mg (GT1/4) ribavirin/day. The duration of treatment was 24 (GT2/3) or 24-72 weeks (GT1/4). RESULTS: Patients with C/C also had higher rates of rapid virologic response (RVR) (GT1, 38.3% vs 11.6%; GT4, 76.5% vs 23.5%; both P<.001) and sustained virologic responses (SVRs) (GT1, 79.1% vs 43.2%; GT4, 85.3% vs 44.1%; both P<.001). In patients with GT2/3, the RVR was more frequent in carriers of C/C (75.3% vs 52.6%, P<.01), but SVR rates were similar between those with C/C and T (80.5% vs 74.4%, P=.31). Results for rs8099917 were comparable. The positive predictive value of rs12979860 C/C for SVR was higher than that of rs8099917 T/T (80.5% vs 71.6%). Overall, RVR was the best predictor of SVR. In patients who did not have GT1, IL28B polymorphisms did not affect the SVR if RVR data were included in the multivariate analysis. CONCLUSIONS: An early virologic response to pegylated interferon and ribavirin is more likely among carriers of rs12979860 C/C and rs8099917 T/T, which might underlie their high rates of SVR. Determination of the IL28B genotype and whether patients have an RVR might be used in future studies of patients with hepatitis C virus genotype 1 or 4.

9. Association of IL28B variants with response to pegylated-interferon alpha plus ribavirin combination therapy reveals intersubgenotypic differences between genotypes 2a and 2b. Sakamoto N, Nakagawa M, Tanaka Y, Sekine-Osajima Y, Ueyama M, Kurosaki M, Nishida N, Tamori A, Yuki NS, Itsui Y, Azuma S, Kakinuma S, Hige S, Itoh Y, Tanaka E, Hiasa Y, Izumi N, Tokunaga K, Mizokami M, Watanabe M, Ochanomizu-Liver Conference Study Group. J Med Virol. 2011;83(5):871. Genetic polymorphisms of the interleukin 28B (IL28B) locus are associated closely with outcomes of pegylated-interferon (PEG-IFN) plus ribavirin (RBV) combination therapy. The aim of this study was to investigate the relationship between IL28B polymorphism and responses to therapy in patients infected with genotype 2. One hundred twenty-nine chronic hepatitis C patients infected with genotype 2, 77 patients with genotype 2a and 52 patients with genotype 2b, were analyzed. Clinical and laboratory parameters, including genetic variation near the IL28B gene (rs8099917), were assessed. Drug adherence was monitored in each patient. Univariate and multivariate statistical analyses of these parameters and clinical responses were carried out. Univariate analyses showed that a sustained virological response was correlated significantly with IL28B polymorphism, as well as age, white blood cell and neutrophil counts, adherence to RBV, and rapid virological response. Subgroup analysis revealed that patients infected with genotype 2b achieved significantly lower rapid virological response rates than those with genotype 2a. Patients with the IL28B-major allele showed higher virus clearance rates at each time point than those with the IL28B-minor allele, and the differences were more profound in patients infected with genotype 2b than those with genotype 2a. Furthermore, both rapid and sustained virological responses were associated significantly with IL28B alleles in patients with genotype 2b. IL28B polymorphism was predictive of PEG-IFN plus RBV combination treatment outcomes in patients infected with genotype 2 and, especially, with genotype 2b. In conclusion, IL-28B polymorphism affects responses to PEG-IFN-based treatment in difficult-to-treat HCV patients.

10. Screening for IL28B gene variants identifies predictors of hepatitis C therapy success. Doehring A, Hofmann WP, Schlecker C, Zeuzem S, Susser S, Geisslinger G, Sarrazin C, Lötsch J. Antivir Ther. 2010;15(8):1099. BACKGROUND: Recent research has shown that genetic variation in the IL28B gene predicts both chronicity of HCV infection and sustained virological response (SVR) to antiviral standard therapy. Because HCV affects 170 million people worldwide and is a leading cause of cirrhosis and hepatocellular carcinoma, screening for prognostic factors in routine clinical practice requires rapid and reliable assays. METHODS: The frequencies of gene polymorphisms IL28B rs8099917, rs12979860 and rs12980275 were investigated in two cohorts of 89 and 187 unrelated HCV-infected Caucasian patients and 195 non-infected participants. This was carried out by means of newly developed sensitive Pyrosequencing™screening assays. RESULTS: The minor alleles were more frequent in patients (n=276) than in controls (n=195), with odds ratios (recessive hereditary model) of 2.2-11.6, indicating a moderate to large genotype effect size. The positive predictive values of the minor alleles for chronicity of HCV infection were 68.3%, 64.8% and 65.8% for rs8099917, rs12979860 and rs12980275, respectively. The minor alleles were also more frequent in patients who had a non-SVR (n=49) than in SVR patients predictive values for non-SVR were 56.9%, 79.2% and 74% for rs8099917, rs12979860 and rs12980275, respectively. CONCLUSIONS: With the screening for IL28B polymorphisms rs12980275, rs8099917 and rs12979860, which are associated with HCV chronicity and with reduced SVR rates, an important prognostic factor of the therapy of chronic hepatitis C can be easily diagnosed.

11. IL28B SNP rs8099917 is strongly associated with pegylated interferon-αand ribavirin therapy treatment failure in HCV/HIV-1 coinfected patients. Aparicio E, Parera M, Franco S, Pérez-Alvarez N, Tural C, Clotet B, Martínez MA. PLoS One. 2010;5(10):e13771. Recent genome-wide association studies report that the SNP rs8099917, located 8.9 kb upstream of the start codon of IL28B, is associated with both disease chronicity and therapeutic response to pegIFN-αand RBV in patients infected with genotype 1 HCV. To determine the effect of rs8099917 variation on the response of HCV to therapy, we genotyped this variant in a cohort of 160 HCV/HIV-1 coinfected patients in our clinic unit who received combined peg-IFN-α/RBV therapy. The rs8099917 T/G or G/G genotypes were observed in 56 patients (35%). Treatment failure occurred in 80% of G-allele carriers versus 48% of non-carriers (P<0.0001). This


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result reveals that the G allele was strongly associated with treatment failure in this patient cohort. Importantly, a highly significant association was found between the G-allele and response to therapy in HCV genotype 1-infected patients (P<0.0001) but not in HCV genotype 3-infected patients. Multivariate analysis (odds ratio; 95% confidence interval; P value) indicated that the rs8099917 TT genotype was a strong predictor of treatment success (5.83; 1.26-26.92; P = 0.021), independent of baseline plasma HCV-RNA load less than 500 000 IU/ml (4.85; 1.18-19.95; P = 0.025) and absence of advanced liver fibrosis (5.24; 1.20-22.91; P = 0.025). These results reveal the high prevalence of the rs8099917 G allele in HCV/HIV-1 coinfected patients as well as its strong association with treatment failure in HCV genotype 1-infected patients. rs8099917 SNP genotyping may be a valid pre-treatment predictor of which patients are likely to respond to treatment in this group of difficult-to-treat HCV/HIV-infected patients.

12. Interleukin-28B genetic variants in identification of hepatitis C virus genotype 1 patients responding to 24 weeks peginterferon/ribavirin. Huang CF, Huang JF, Yang JF, Hsieh MY, Lin ZY, Chen SC, Wang LY, Juo SH, Chen KC, Chuang WL, Kuo HT, Dai CY, Yu ML. J Hepatol. 2012 Jan;56(1):34-40. Epub 2011 May 19. AIMS: A substantial proportion of hepatitis C virus genotype 1 (HCV-1) patients achieved a sustained virological response (SVR, HCV RNA seronegative throughout 24 weeks of post-treatment follow-up) after 24 weeks peginterferon/ribavirin therapy. We explored the role of interleukin-28B genotype in identifying patients who responded to the regimen. METHODS: Interleukin-28B rs8099917 genotype was determined in 226 HCV-1 patients with 24 weeks peginterferon/ribavirin. RESULTS: Compared to patients with rs8099917 TG/GG genotype, those with TT genotype had significantly higher rapid virological response (RVR, HCV RNA seronegative at treatment week 4, 54.0% vs. 17.9%, p<0.001) and SVR (64.7% vs. 25.6%, p<0.001) rates, and lower relapse rate (28.0% vs. 54.5%, p=0.01). Logistic regression analysis revealed that the strongest factor predictive of a RVR was the carriage of rs8099917 TT genotype (odds ratio/ 95% confidence intervals [OR/CI]: 6.24/2.34-16.63), followed by lower viral loads (OR/CI: 5.29/2.81-9.93) and age (OR/CI:0.94/0.91-9.97). The most important factor predictive of an SVR was the attainment of a RVR (OR/CI: 22.23/9.22-53.58), followed by the carriage of rs8099917 TT genotype (OR/CI: 3.38/1.18-9.65), lower viral loads (OR/CI: 2.23/1.00-4.93) and ribavirin exposure dose (OR/CI: 1.17/1.06-1.30). The determinant power of rs8099917 genotype on SVR was mainly restricted to non-RVR patients, particularly those with higher baseline viral loads. Combination of the two pretreatment predictors, interleukin-28B genotype and baseline viral loads, could predict treatment efficacy with a positive predictive value of 80% and a negative predictive value of 91%. CONCLUSIONS: Interleukin-28B genotype could help identifying patients who are or are not candidates for an abbreviated regimen before treatment.

13. Amino acid substitution in hepatitis C virus core region and genetic variation near the interleukin 28B gene predict viral response to telaprevir with peginterferon and ribavirin. Akuta N, et al. Hepatology. 2010;52(2):421. Genetic variation near the IL28B gene and substitution of amino acid (aa) 70 and 91 in the core region of hepatitis C virus (HCV) genotype 1b can predict the response to pegylated interferon (PEG-IFN)/ribavirin combination therapy, but its impact on triple therapy of telaprevir/PEG-IFN/ribavirin is not clear. The aims of this study were to investigate the predictive factors of sustained virological response to a 12-week or 24-week regimen of triple therapy in 72 of 81 Japanese adults infected with HCV genotype 1. Overall, sustained virological response and end-of-treatment response were achieved by 61% and 89%, respectively. Especially, the sustained virological response was achieved by 45% and 67% in the 12- and 24-week regimens, respectively. Multivariate analysis identified rs8099917 near the IL28B gene (genotype TT) and substitution at aa 70 (Arg70) as significant determinants of sustained virological response. Prediction of response to therapy based on a combination of these factors had high sensitivity, specificity, and positive and negative predictive values. The efficacy of triple therapy was high in the patients with genotype TT, who accomplished sustained virological response (84%), irrespective of substitution of core aa 70. In the patients having genotype non-TT, those of Arg70 gained high sustained virological response (50%), and sustained virological response (12%) was the worst in patients who possessed both genotype non-TT and Gln70(His70). Conclusion: This study identified genetic variation near the IL28B gene and aa substitution of the core region as predictors of sustained virological response to a triple therapy of telaprevir/PEG-IFN/ribavirin in Japanese patients infected with HCV genotype 1b.

14. Limited impact of IL28B genotype on response rates in telaprevir-treated patients with prior treatment failure. Pol S, Aerssens J, Zeuzem S, Andreone P, Lawitz EJ, Roberts S, Younossi Z, Foster GR, Focaccia R, Horban A, Pockros PJ, Van Heeswijk RP, De Meyer S, Luo D, Botfield M, Beumont M, Picchio G. J Hepatol. 2013 May;58(5):883-9. Epub 2013 Jan 12. AIMS: Nucleotide polymorphisms upstream of the interleukin 28B (IL28B) gene are strongly associated with hepatitis C virus (HCV) clearance in treatment-naïve patients treated with peginterferon/ribavirin (PegIFN/RBV). This subanalysis of the REALIZE study evaluated the impact of IL28B polymorphisms on sustained virologic response (SVR) in telaprevir-treated, HCV genotype 1-infected patients with prior PegIFN/RBV treatment failure. METHODS: Treatment-experienced patients were randomized to 12weeks of telaprevir (750mg every 8h) with/without a 4-week PegIFN/RBV lead-in, or placebo, each with PegIFN-α-2a (180μg/week) and ribavirin (1000-1200mg/day) for 48weeks overall. Data from telaprevir arms were pooled. RESULTS: Eighty percent (527/662) of patients consented to genetic testing and were included. Similar proportions of patients had IL28B CC, CT and TT genotypes across treatment arms; baseline characteristics were generally well balanced. SVR rates were higher in the pooled telaprevir versus placebo group for all IL28B genotypes; CC: 79% versus 29%, respectively; CT: 60% versus 16%, respectively; TT: 61% versus 13%, respectively. Within each prior response category (relapse, partial or null response), SVR and viral breakthrough rates with telaprevir-based treatment were comparable across IL28B genotypes. IL28B genotype did not significantly affect SVR (2-step multivariate analyses; p>0.16 in pairwise comparison among CC, TT, and CT). Variations in rapid virologic response and relapse rates were noted in certain patient subgroups. CONCLUSIONS: Our findings suggest that IL28B genotype has a limited impact on SVR rates


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with telaprevir-based therapy in treatment-experienced patients. IL28B genotyping may have limited utility in the baseline evaluation of similar patients considered for telaprevir-based therapy.

15. Genetic variation in IL28B and spontaneous clearance of hepatitis C virus. Thomas DL, Thio CL, Martin MP, Qi Y, Ge D, O'Huigin C, Kidd J, Kidd K, Khakoo SI, Alexander G, Goedert JJ, Kirk GD, Donfield SM, Rosen HR, Tobler LH, Busch MP, McHutchison JG, Goldstein DB, Carrington M. Nature. 2009;461(7265):798. Hepatitis C virus (HCV) infection is the most common blood-borne infection in the United States, with estimates of 4 million HCV-infected individuals in the United States and 170 million worldwide. Most (70-80%) HCV infections persist and about 30% of individuals with persistent infection develop chronic liver disease, including cirrhosis and hepatocellular carcinoma. Epidemiological, viral and host factors have been associated with the differences in HCV clearance or persistence, and studies have demonstrated that a strong host immune response against HCV favours viral clearance. Thus, variation in genes involved in the immune response may contribute to the ability to clear the virus. In a recent genome-wide association study, a single nucleotide polymorphism (rs12979860) 3 kilobases upstream of the IL28B gene, which encodes the type III interferon IFN-3, was shown to associate strongly with more than a twofold difference in response to HCV drug treatment. To determine the potential effect of rs12979860 variation on outcome to HCV infection in a natural history setting, we genotyped this variant in HCV cohorts comprised of individuals who spontaneously cleared the virus (n = 388) or had persistent infection (n = 620). We show that the C/C genotype strongly enhances resolution of HCV infection among individuals of both European and African ancestry. To our knowledge, this is the strongest and most significant genetic effect associated with natural clearance of HCV, and these results implicate a primary role for IL28B in resolution of HCV infection.

16. Association of genetic variation in IL28B with hepatitis C treatment-induced viral clearance in the Chinese Han population. Liao XW, Ling Y, Li XH, Han Y, Zhang SY, Gu LL, Yu DM, Yao BL, Zhang DH, Jin GD, Lu ZM, Gong QM, Zhang XX. Antivir Ther. 2011;16(2):141-7. BACKGROUND: Genome-wide association studies have recently shown that the rs12979860 polymorphism in IL28B is associated with the response to chronic hepatitis C treatment. The aim of this study was to investigate whether rs12979860 could be used as a predictive marker for end-of-treatment response (ETR) or sustained virological response (SVR) in the Chinese Han population. METHODS: The rs12979860 genotype was detected in 259 individuals infected with HCV by DNA sequencing. Among them, 120 patients were administered complete pegylated interferon-αand ribavirin combination therapy and 92 patients were followed for 24 weeks after the cessation of treatment and were divided into different groups according to outcomes of treatment. RESULTS: The rs12979860 genotype CC was the primary genotype (87.64%, 227/259) and genotype TT was found in only one individual within this cohort. The patients with the rs12979860 genotype CC had higher rates of ETR (P=0.0044) and SVR (P=0.0046) than the patients with N-CC (CT or TT). In multivariate analyses, the rs12979860 genotype CC was associated with a substantial difference in rates of achieving ETR (odds ratio [OR]8.983, 95% confidence interval [CI]2.173-37.145; P=0.0024) and SVR (OR 24.298, 95% CI 2.27-259.90; P=0.0083). CONCLUSIONS: This study demonstrated for the first time that the rs12979860 variation in IL28B could be a predictor of ETR and SVR in Chinese Han patients infected with HCV. The high frequency of the rs12979860 genotype CC might explain why the SVR rate is higher than that of the average global population.

17. Polymorphism near the IL28B gene in Korean hepatitis C virus-infected patients treated with peg-interferon plus ribavirin. Lyoo K, Song MJ, Hur W, Choi JE, Hong SW, Kim CW, Bae SH, Choi JY, Choi SW, Shin EC, Yoon SK. J Clin Virol. 2011 Dec;52(4):363-6. Epub 2011 Sep 9. BACKGROUND: Single nucleotide polymorphisms (SNPs) near the IL28B gene have recently been described as predictors of antiviral therapy responses in patients with hepatitis C virus (HCV) genotype-1. OBJECTIVES: The aim of this study was to investigate the association between genetic variation near the IL28B gene and treatment outcome prediction in Korean patients receiving peg-interferon (PEG-IFN) plus ribavirin therapy. STUDY DESIGN: The allelic discrimination assay by Taqman real-time PCR was developed to determine genotypes of SNPs, rs12979860 and rs8099917, which were analyzed in 65 Korean patients with HCV genotype-1. RESULTS: For rs12979860, the frequency of patients with sustained virological response (SVR) was 70.2% in those with the CC genotype and 25% in those with the CT genotype. Early virological response (EVR) inpatients with the CC genotype (84.2%) was higher than in those with the CT genotype (25.0%). For rs8099917, patients with the TT genotype showed significantly higher in SVR and EVR than those with the TG/GG genotype (69.6% vs 33.3% and 82.1% vs 44.4%, respectively). With regards to the genotype frequency of the SNPs, the homozygous genotypes for rs12979860 (CC) or rs8099917 (TT) in Korean patients showed a significantly higher frequency as compared with other ethnicities; Caucasians, African-American, Hispanic, and Japanese. CONCLUSIONS: These results demonstrate that the genotypes rs12979860 CC and rs8099917 TT were more frequently observed in Korean patients compared to other ethnicities, and suggest that the genetic characteristics of patients may be prognostic factor that predicts antiviral response to PEG-IFN therapy for chronic hepatitis C.

18. Association of IL28B SNP With Progression of Egyptian HCV Genotype 4 Patients to End Stage Liver Disease. El-Awady MK, et al. Hepat Mon. 2012 Apr;12(4):271-7. Epub 2012 Apr 30. BACKGROUND: IL28B single nucleotide polymorphisms (SNPs) play important roles in the management of hepatitis C virus (HCV) infections and are strongly associated with spontaneous and treatment-induced HCV clearance. OBJECTIVES: In the present study, the association between IL28B variants and the progression of HCV infection in Egyptian patients infected with type 4a virus will be examined. PATIENTS AND METHODS: Frequencies of the protective genotype C/C of SNP, rs12979860 were determined in healthy subjects, spontaneous resolvers, and chronic HCV type 4 patients with low F scores and in patients with end stage liver disease (ESLD). This study included a total of 404 subjects. Patients infected with HCV type 4a (n = 304) were divided into; chronic hepatitis C (CHC) with low F scores (CHC, n = 110), end stage liver disease (n = 110), liver cirrhosis (LC) (n = 35) and hepatocellular carcinoma (HCC) patients (n = 75),


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spontaneous resolvers of HCV infection (n = 84) were also included. A healthy group representing the Egyptian population (n = 100) was also included in the genotyping of IL28B. The later was typed via a polymerase chain reaction based restriction fragment length polymorphism (PCR-RFLP) assay analysis on purified genomic DNA extracted from all individuals. RESULTS: A significant increase (P<0.0005) was observed in frequencies of IL-28B rs12979860 C/C genotypes in the healthy population, than in the CHC, LC and HCC groups (C/C = 48%, 13%, 0%.and 0% respectively). On the other hand the C/C genotype was significantly higher (P<0.0005) in spontaneous resolvers than in healthy subjects. A comparable significant increase in the frequency of C/T allele accompanied by mild elevation of T/T allele frequency, were detected along the progression towards ESLD. CONCLUSIONS: Genotype C/C is associated with viral clearance during acute infection. The sharp decline in the C/C genotype from healthy to CHC subjects and the total absence of the C/C genotype in ESLD suggests a central role of this genotype against HCV disease progression.

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