Antiviral Therapy 2017; 22:61 70 (doi: /IMP3085)

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1 Antivirl Therpy 217; 22:61 7 (doi: /IMP385) Originl rticle Comprison of the Abbott RelTime HCV nd Roche COBAS Ampliprep/COBAS TqMn HCV ssys for the monitoring of sofosbuvir-bsed therpy Eiichi Ogw 1 *, Norihiro Furusyo 1, Msyuki Murt 1, Motohiro Shimizu 1, Kzuhiro Toyod 1, Teko Hott 2, Tkeshi Uchiumi 2, Jun Hyshi 3 1 Deprtment of Generl Internl Medicine, Kyushu University Hospitl, Fukuok, Jpn 2 Deprtment of Clinicl Chemistry nd Lbortory of Medicine, Kyushu University Hospitl, Fukuok, Jpn 3 Kyushu Generl Internl Medicine Center, Hrdoi Hospitl, Fukuok, Jpn *Corresponding uthor e-mil: eogw@gim.med.kyushu-u.c.jp Bckground: On-tretment HCV kinetics ply n invluble role in evluting the efficcy of interferon-bsed therpies. However, the importnce of HCV RNA monitoring hs not been well discussed concerning tretment with sofosbuvir (SOF)-bsed regimens, especilly for the utility of the Abbott RelTime HCV (ART) ssy. Methods: This study consisted of 151 ptients infected with HCV genotype-1 or -2, including ptients with prior tretment-experience or cirrhosis. HCV genotype-1 ptients were treted with SOF/ledipsvir nd genotype-2 ptients with SOF/ribvirin, both for 12 weeks. Seril mesurements of HCV RNA were performed with both the ART nd COBAS AmpliPrep/COBAS TqMn v2. (CAP/ CTM) ssys simultneously t weeks, 1, 2, 4, 6, 8, 1 nd 12 of tretment. Results: The rtes of HCV RNA trget not detected (TND) by ART were significntly lower thn those by CAP/CTM between weeks 2 nd 12 (end of tretment [EOT]), irrespective of prior tretment-experience or cirrhosis. 11 (11.6%) genotype-1 nd 8 (14.3%) genotype-2 ptients did not chieve HCV RNA TND by ART t EOT, in contrst to ll hving HCV RNA TND by CAP/CTM; however, ll chieved sustined virologicl response. The time t which HCV RNA becme TND or unquntifible ws not ssocited with tretment outcome by either the ART or CAP/CTM ssy. Conclusions: Over 1% of the ptients continued to hve detectble HCV RNA by ART t EOT, irrespective of HCV genotype, prior tretment-experience nd/or cirrhosis. However, prolonged residul HCV RNA ws not ssocited with tretment filure. Introduction An estimted 17 million people worldwide hve chronic HCV infection [1], which is mjor cuse of liver cirrhosis nd heptocellulr crcinom [2]. Of the rpidly evolving tretments for HCV, vrious interferonfree regimens bsed on direct-cting ntivirl gents (DAAs) hve become the gold stndrd [3,4], lthough mny ptients continue to be treted with pegylted interferon-lph (PEG-IFN-) nd ribvirin (RBV) in Asin nd Africn countries, which hve reltively high rtes of HCV prevlence [5]. On-tretment HCV kinetics ply n invluble role in evluting the efficcy of ntivirl therpies tht include PEG-IFN- nd RBV for chronic HCV infection [6 9]. In prticulr, it contributes to the prediction of tretment outcome nd to decisions on the tretment durtion. Rel-time PCR-bsed ssys with differing sensitivity nd ccurcy re commonly used in clinicl prctice. Among them is the most widely used HCV ssy, the Roche COBAS AmpliPrep/ COBAS TqMn HCV test (CAP/CTM). The utility of more sensitive ssy for very low viremi, the Abbott RelTime HCV (ART) ssy, hs been reported for DAA-bsed triple therpy [1 13]. The mjor difference between these ssys is the discordnce in the detection of HCV RNA t the lower limits of quntifiction, which could influence tretment outcome. Among the IFN-free therpies, regimens contining sofosbuvir (SOF; nucleotide NS5B polymerse inhibitor) re currently used in Europe, North Americ nd Jpn. According to rel-world cohort study, the sustined virologicl response (SVR) rte is over 9% for SOF nd ledipsvir (LDV; NS5A inhibitor) therpy, 217 Interntionl Medicl Press (print) (online) 61

2 E Ogw et l. with or without RBV, due to good tolerbility nd the ntivirl effect [14,15]. The importnce of on-tretment HCV RNA mesurement hs not been well discussed, nd there re questions bout the necessity of predicting tretment outcome becuse lmost ll ptients chieve rpid virologicl response. Recently, comprison of the effect of the CAP/CTM nd ART ssys for SOFbsed regimens ws reported [16,17]; however, there is limited dt on SOF/LDV for ptients with genotype-1, especilly for tretment-experienced nd/or cirrhotic ptients. The purpose of this study is to nlyse HCV kinetics during tretment with SOF/LDV for ptients with HCV genotype-1 nd with SOF/RBV for ptients with genotype-2, using the ART nd CAP/CTM v2. ssys to determine the ssocition of HCV RNA levels during tretment with tretment outcome. Methods Ptients This study consisted of 152 Jpnese ptients with confirmed chronic HCV genotype-1 or -2 infection, including ptients with prior tretment-experience or cirrhosis, who initited SOF-bsed therpy between June 215 nd Jnury 216. Eligible ptients were ged 2 yers nd older who hd HCV monoinfection. Exclusion criteri included lost to follow up (n=1), clinicl or biochemicl evidence of heptic decompenstion (Child Pugh B or C; n=), nd ny non-hcvrelted liver disese, such s utoimmune heptitis or primry biliry cholngitis (n=). After exclusions, the dt of 151 ptients ws vilble for nlysis. The study ws conducted in ccordnce with the ethicl principles of the 28 Declrtion of Helsinki, ws pproved by the Ethics Committee of Kyushu University Hospitl, nd is registered s clinicl study on the University Hospitl Medicl Informtion Network (ID 23115). Clinicl nd lbortory ssessment Clinicl prmeters were mesured by stndrd lbortory techniques t commercil lbortory (SRL Lbortory, Tokyo, Jpn). Blood smples were obtined t weeks, 2, 4, 6, 8, 1 nd 12 (end of tretment [EOT]), then t 4-week intervls until 12 weeks fter the end of tretment. Heptic fibrosis sttus ws ssessed by biopsy or trnsient elstogrphy (FibroScn). Liver biopsy ws performed within 1 month before the initition of tretment. The minimum length of the liver biopsy ws 15 mm, nd t lest 1 complete portl trcts were necessry for inclusion. For ech specimen, the stge of fibrosis ws estblished ccording to the METAVIR score [18]. We defined cirrhosis s METAVIR F4 or FibroScn >14.9 kp [19], which ws used if the META- VIR score ws unvilble. Genotyping of the interleukin-28b (IL28B; rs899917) gene ws performed using the ABI TqMn llelic discrimintion kit (75 RelTime PCR System; Applied Biosystems, Crlsbd, CA, USA). The llele of the IL28B single nucleoside polymorphism ws determined for ll studied ptients. Determintion of the HCV RNA level All serum smples were tested simultneously for HCV RNA level by the CAP/CTM v2. nd ART ssys. Testing ws performed t Kyushu University Hospitl ccording to the mnufcturer s instructions. Performnce chrcteristics of the two ssys hve been described in detil previously [13]. CAP/CTM ssy results re reported s trget not detected (TND), trget detected but lower limittion of quntifiction (TD/<LLOQ; <15 IU/ml) or bsolute number (quntifible results: 15 IU/ml). The rnge for liner quntifiction is between 15 nd IU/ml. ART ssy results re reported s TND, TD/<LLOQ (<12 IU/ml) or bsolute number (quntifible results: 12 IU/ml). The rnge of liner quntifiction is between 12 nd 1 8 IU/ml. Antivirl tretment nd tretment outcome All HCV genotype-1 ptients received combintion of SOF/LDV (Hrvoni; Giled Sciences KK, Tokyo, Jpn) for 12 weeks. All HCV genotype-2 ptients received SOF (Sovldi; Giled Sciences KK) nd RBV (Rebetol; MSD KK, Tokyo, Jpn; bsed on bodyweight, orlly) for 12 weeks. SVR ws defined s HCV RNA <LLOQ 12 weeks fter the end of tretment (SVR12) for both ssys. A virologicl relpse ws defined s recurrence of quntifible HCV RNA fter the end of tretment. Sttisticl nlysis Sttisticl nlyses were conducted using SPSS Sttistics version 22. (IBM SPSS Inc., Chicgo, IL, USA). Bseline continuous dt re expressed s medin (first third qurtiles) nd ctegoricl vribles re reported s frequencies nd percentges. Univrite nlysis ws done using the c 2 or Fisher s exct test, s pproprite. The positive predictive vlue (PPV) of undetectble nd unquntifible HCV RNA is the proportion of ptients with HCV RNA TND nd TND or TD/<LLOQ who chieved SVR12. The negtive predictive vlue (NPV) of detectble nd quntifible HCV RNA is the proportion of ptients with HCV RNA TD/<LLOQ or LLOQ nd LLOQ who filed tretment. These vlues were clculted for both ssys for ech tretment regimen t weeks 2, 4, 6, 8, 1 nd 12. A P-vlue less thn.5 ws regrded s sttisticlly significnt in ll nlyses Interntionl Medicl Press

3 HCV RNA monitoring of sofosbuvir-bsed therpy Results Ptient chrcteristics nd tretment outcome for HCV genotype-1 The bseline demogrphics nd clinicl chrcteristics of the 151 ptients included in this study re shown in Tble 1. Of the 95 (62.9%) infected with HCV genotype-1, lmost ll (n=94, 98.9%) were infected with HCV genotype-1b. Medin ge ws 69, with 87 (91.6%) of the ptients over 6, 52 (54.7%) tretmentexperienced nd 35 (36.8%) with cirrhosis. Of the tretment-experienced ptients, 2 (38.5%) hd been treted with DAA, such s triple therpy with telprevir (n=2) or simeprevir (SMV; n=15) or n IFN-free regimen with dcltsvir (DCV)/sunprevir (n=3). Tble 1. Bseline chrcteristics Genotype-1 Genotype-2 Number Mle 4 (42.1) 29 (51.8) Age, yers 69 (64 76) 64 (5 71) Body mss index, kg/m ( ) 22.4 ( ) Serum lbumin, g/l 4 (38 43) 41 (37 44) Alnine minotrnsferse, U/l 34 (24 5) 32 (18 6) g-glutmyl trnspeptidse, U/l 29 (19 48) 27 (14 5) egfr, ml/min/1.73 m 2 62 (52 72) 76 (66 86) -fetoprotein, ng/ml 5.5 ( ) 3. ( ) Prothrombin time, % 9 (75 98) 91 (78 12) Hemoglobin, g/l 136 ( ) 136 ( ) Pltelet count, 1 9 /l 147 (12 199) 166 (119 26) HCV RNA level, log 1 IU/ml ART, log 1 IU/ml 5.86 ( ) 6.3 ( ) CAP/CTM, v2., log 1 IU/ml 6.1 ( ) 6.3 ( ) HCV genotype 1 1 (1.1) 1b 94 (98.9) 2 27 (48.2) 2b 29 (51.8) Liver cirrhosis 35 (36.8) 15 (26.8) Dibetes mellitus 1 (1.6) 13 (23.2) IL28B SNPs (rs899917) TT 58 (61.1) 39 (69.6) TG/GG 37 (38.9) 17 (3.4) Tretment-nive 43 (45.3) 4 (71.4) Tretment-experienced PEG-IFN/RBV 32 (33.7) 16 (28.6) Telprevir-triple 2 (2.1) Simeprevir-triple 15 (15.8) DCV/ASV 3 (3.2) SVR12 93 (97.9) 5 (89.3) Dt re expressed s n (%) or medin (first third qurtiles). Triple therpy in combintion with PEG-IFN/RBV. ART, Abbott RelTime HCV ssy; CAP/CTM, Roche COBAS AmpliPrep/COBAS TqMn HCV ssy; DCV/ASV, dcltsvir/sunprevir; egfr, estimted glomerulr filtrtion rte; IL28B, interleukin-28b; PEG-IFN, pegylted interferon; RBV, ribvirin; SNP, single nucleoside polymorphism; SVR12, sustined virologicl response 12 weeks fter the end of tretment. All HCV genotype-1 ptients were treted with SOF/ LDV for 12 weeks without temporry interruption or discontinution nd 97.9% (93/95) chieved SVR12. The two non-svr ptients hd evidence of liver cirrhosis nd hd relpse fter the end of tretment. Ptient chrcteristics nd tretment outcome for HCV genotype-2 Of the 56 HCV genotype-2 ptients, 27 (48.2%) were infected with HCV genotype-2 nd 29 (51.8%) with -2b. Medin ge ws 64, with 34 (6.7%) of the ptients over 6, 16 tretment-experienced (28.6%) nd 15 with cirrhosis (26.8%). All tretment-experienced ptients hd been treted with PEG-IFN-/RBV. All HCV genotype-2 ptients were treted with SOF/RBV for 12 weeks without temporry interruption or discontinution nd 89.3% (5/56) chieved SVR12. None experienced virl brekthrough during tretment, in common with SOF/LDV for HCV genotype-1. Of the six non-svr ptients, five were infected with HCV genotype-2, three were tretment-experienced nd two hd evidence of liver cirrhosis. HCV kinetics during SOF/LDV tretment for HCV genotype-1 SOF/LDV tretment brought rpid HCV RNA reduction in the erly stge (medin HCV RNA level t week 1: 2.1 log IU/ml by CAP/CTM nd 1.91 log IU/ml by ART). Overll, the rtes of HCV RNA TND by ART were significntly lower thn those by CAP/CTM between weeks 2 nd 12, s shown in Figure 1A. Of note, there ws n extreme difference between the ssys in the HCV RNA TND rtes t weeks 4 nd 6 (82.1% nd 96.8% by CAP/CTM nd 33.7% nd 57.9% by ART, respectively; both P<.1). Similr results were found for the tretment-experienced nd cirrhotic groups, s shown in Figure 1B nd 1C. Moreover, 11 (11.6%) did not chieve HCV RNA TND by ART t EOT, in contrst to ll hving HCV RNA TND by CAP/CTM. HCV kinetics during SOF/RBV tretment for HCV genotype-2 SOF/RBV tretment lso brought rpid HCV RNA reduction in the erly stge (medin HCV RNA level t week 1: 1.9 log IU/ml by CAP/CTM nd 1.81 log IU/ ml by ART), similr to SOF/LDV tretment. Overll, the HCV RNA TND rtes by ART were significntly lower thn those by CAP/CTM between weeks 2 nd 12, s shown in Figure 2A. Of note, there ws n extreme difference between the ssys in the HCV RNA TND rtes t weeks 2, 4 nd 6 (44.6%, 82.1% nd 96.8% by CAP/CTM nd 12.5%, 39.3% nd 64.3% by ART, respectively; ll P<.1). Similr results were found in the tretment-experienced nd cirrhotic groups, s shown in Figure 2B nd 2C. The sme pttern ws seen Antivirl Therpy

4 E Ogw et l. Figure 1. Rtes of undetectble (trget not detected) HCV RNA during SOF/LDV tretment for HCV genotype-1 ptients A Genotype-1, SOF/LDV, overll B Genotype-1, SOF/LDV, tretment-experienced b c Rtes of undetectble HCV RNA (trget not detected), % w 2w 4w 6w 8w 1w 12w Rtes of undetectble HCV RNA (trget not detected), % b w 2w 4w 6w 8w 1w 12w C Rtes of undetectble HCV RNA (trget not detected), % b Genotype-1, SOF/LDV, cirrhosis w 2w 4w 6w 8w 1w 12w b c CAP/CTM ART (A) Overll (n=95), (B) tretment-experienced (n=52) nd (C) cirrhosis (n=35). P<.1; b P<.1; c P<.5. ART, Abbott RelTime HCV ssy; CAP/CTM, Roche COBAS AmpliPrep/COBAS TqMn HCV ssy; EOT, end of tretment; SOF/LDV, sofosbuvir/ledipsvir. s for SOF/LDV tretment, 8 (14.3%) did not chieve HCV RNA TND by ART t EOT, in contrst to ll hving HCV RNA TND by CAP/CTM. Moreover, residul HCV RNA (TD/<LLOQ) ws detected in two cses t week 4 fter the end of tretment, despite the ptients hving chieved SVR12. Predictive vlue of HCV RNA during SOF/LDV tretment for HCV genotype-1 The predictive vlues of undetectble (TND) nd unquntifible (TND or TD/<LLOQ) HCV RNA by ART during SOF/LDV tretment for HCV genotype-1 re shown in Tbles 2, 3, 4 nd 5. Of the Interntionl Medicl Press

5 HCV RNA monitoring of sofosbuvir-bsed therpy Figure 2. Rtes of undetectble (trget not detected) HCV RNA during SOF/RBV tretment for HCV genotype-2 ptients A Genotype-2, SOF/RBV, overll B Genotype-2, SOF/RBV, tretment-experienced 98.2 b 93.8 Rtes of undetectble HCV RNA (trget not detected), % w 2w 4w 6w 8w 1w 12w 85.7 Rtes of undetectble HCV RNA (trget not detected), % c w 2w 4w 6w 8w 1w 12w C Rtes of undetectble HCV RNA (trget not detected), % c 73.3 Genotype-2, SOF/RBV, cirrhosis w 2w 4w 6w 8w 1w 12w CAP/CTM ART (A) Overll (n=56), (B) tretment-experienced (n=16) nd (C) cirrhosis (n=15). P<.1; b P<.1; c P<.5. ART, Abbott RelTime HCV ssy; CAP/CTM, Roche COBAS AmpliPrep/COBAS TqMn HCV ssy; EOT, end of tretment; SOF/RBV, sofosbuvir/ribvirin. ptients treted with SOF/LDV, only two (2.1%) did not chieve SVR12. Both hd HCV RNA TND by ART t week 8, lthough they hd HCV RNA TND by CAP/CTM t week 2. Therefore, ll ptients with HCV RNA TND by ART t weeks 2, 4 nd 6 chieved SVR12 (PPV %). Otherwise, one (1.1%) hd HCV RNA LLOQ by ART t weeks 8 nd 1, but chieved SVR12. It should be noted tht 11 (11.6%) ptients hd HCV RNA TD/<LLOQ t EOT, but ll of them chieved SVR4 nd SVR12. We lso evluted the predictive vlues of HCV RNA by CAP/CTM, s shown in Additionl file 1. Most of the Antivirl Therpy

6 E Ogw et l. Tble 2. Predictive vlues of undetectble HCV RNA by ART during SOF/LDV therpy for HCV genotype-1 ptients (week 2 to 6) Week 2 Week 4 Week 6 TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ Number Sensitivity, % (n/totl n) 8.6 (8/93) 91.4 (85/93) 34.4 (32/93) 65.6 (61/93) 59.1 (55/93) 4.9 (38/93) Specificity, % (n/totl n) (2/2) (/2) (2/2) (/2) (2/2) (/2) PPV, % (n/totl n) (8/8) 97.7 (85/87) (32/32) 96.8 (61/63) (55/55) 95. (38/4) NPV, % (n/totl n) 2.3 (2/87) (/8) 3.2 (2/63) (/32) 5. (2/4) (/55) ART, Abbott RelTime HCV ssy; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/LDV, sofosbuvir/ledipsvir; TD, trget detected; TND, trget not detected. Tble 3. Predictive vlues of undetectble HCV RNA by ART during SOF/LDV therpy for HCV genotype-1 ptients (week 8 to 12) Week 8 Week 1 Week 12 TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ Number Sensitivity, % (n/totl n) 73.1 (68/93) 26.9 (25/93) 8.6 (75/93) 19.4 (18/93) 88.2 (82/93) 11.8 (11/93) Specificity, % (n/totl n) (/2) (2/2) (/2) (2/2) (/2) (2/2) PPV, % (n/totl n) 97.1 (68/7) (25/25) 97.4 (75/77) (18/18) 97.6 (82/84) (11/11) NPV, % (n/totl n) (/25) 2.9 (2/7) (/18) 2.6 (2/77) (/11) 2.4 (2/84) ART, Abbott RelTime HCV ssy; EOT, end of tretment; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/LDV, sofosbuvir/ledipsvir; TD, trget detected; TND, trget not detected. Tble 4. Predictive vlues of unquntifible HCV RNA by ART during SOF/LDV therpy for HCV genotype-1 ptients (week 2 to 6) Week 2 Week 4 Week 6 TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ Number Sensitivity, % (n/totl n) 4.9 (38/93) 59.1 (55/93) 8.6 (75/93) 19.4 (18/93) 95.7 (89/93) 4.3 (4/93) Specificity, % (n/totl n) 5. (1/2) 5. (1/2) (/2) (2/2) (/2) (2/2) PPV, % (n/totl n) 97.4 (38/39) 98.2 (55/56) 97.4 (75/77) (18/18) 97.8 (89/91) (4/4) NPV, % (n/totl n) 1.8 (1/56) 2.6 (1/39) (/18) 2.6 (2/77) (/4) 2.2 (2/91) ART, Abbott RelTime HCV ssy; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/LDV, sofosbuvir/ledipsvir; TD, trget detected; TND, trget not detected. Tble 5. Predictive vlues of unquntifible HCV RNA by ART during SOF/LDV therpy for HCV genotype-1 ptients (week 8 to 12) Week 8 Week 1 Week 12 TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ Number Sensitivity, % (n/totl n) 98.9 (92/93) 1.1 (1/93) 98.9 (92/93) 1.1 (1/93) (93/93) NA Specificity, % (n/totl n) (/2) (2/2) (/2) (2/2) (/2) NA PPV, % (n/totl n) 97.8 (89/91) (1/1) 97.8 (89/91) (1/1) 97.9 (93/95) NA NPV, % (n/totl n) (/4) 2.1 (2/94) (/4) 2.1 (2/94) NA NA ART, Abbott RelTime HCV ssy; EOT, end of tretment; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/LDV, sofosbuvir/ledipsvir; TD, trget detected; TND, trget not detected Interntionl Medicl Press

7 HCV RNA monitoring of sofosbuvir-bsed therpy ptients hd undetectble (n=78, 82.1%) nd unquntifible (n=92, 96.8%) HCV RNA t week 4, nd ll hd undetectble HCV RNA t week 8. High PPV nd low NPV vlues t ll points during tretment were seen for both the ART nd CAP/CTM ssys. Predictive vlue of HCV RNA during SOF/RBV tretment for HCV genotype-2 The predictive vlues of undetectble nd unquntifible HCV RNA by ART during SOF/RBV tretment for HCV genotype-2 re shown in Tbles 6, 7, 8 nd 9. Rpid virologicl response (HCV RNA TND) ws found for 7 (12.5%) ptients t week 2 nd for 22 (39.3%) ptients t week 4, but one (PPV 85.7%) nd five (PPV 89.8%) relpsed t the respective time points. Only one ptient (1.8%) hd HCV RNA LLOQ t weeks 8 nd 1, but chieved SVR12. However, s in the cse with SOF/LDV tretment, 8 (14.3%) ptients hd HCV RNA TD/<LLOQ t EOT, two of whom did not chieve SVR4 but did chieve SVR12. The predictive vlues of HCV RNA results by CAP/ CTM for genotype 2 ptients re shown in Additionl file 1. The CAP/CTM ssy ws less sensitive, indicting tht fewer ptients hd quntifible HCV RNA t weeks 2 nd 4 thn were identified by ART. Of the 15 ptients (26.8%) who hd HCV RNA LLOQ t week 2, 12 chieved SVR12 (PPV 8.%). This PPV vlue ws not significnt but ws lower thn tht of ptients who hd unquntifible HCV RNA t week 2 (PPV 92.7%, 38/41). Tken together, the dt indicte tht HCV RNA kinetics were not ssocited with tretment outcome, even with the more sensitive ART ssy, for either SOF/LDV or SOF/RBV tretment. Discussion Mesurement of HCV RNA during ntivirl tretment with PEG-IFN-/RBV hs been shown to be cliniclly useful for predicting tretment outcome nd for mking decisions on tretment durtion [6 9]. Furthermore, monitoring virl kinetics in the erly phse hs been reported to be beneficil for regimens tht include DAA in combintion with PEG-IFN-/ RBV [11 13,2,21]. However, with the pprovl of the IFN-free regimen with SOF, lmost ll ptients chieve SVR, regrdless of erly virl response t week 4. In this study, we followed the virl kinetics in 12-week regimens of SOF/LDV for genotype-1 nd SOF/RBV for genotype-2, using the ART nd CAP/ CTM v2. ssys. More thn 1% of the ptients hd HCV RNA TD/<LLOQ by ART t EOT for both tretments but ll chieved SVR12, indicting tht detectble residul HCV RNA t the end of SOF/LDV or SOF/RBV tretment ws not ssocited with tretment filure. Some studies used the CAP/CTM nd ART ssys to follow virl kinetics during DAA (NS3/4A protese inhibitor)-bsed triple therpy [1 13]. The time of HCV RNA TND ws significntly lter by ART thn by CAP/CTM; however, lte response by ART ws not relted to tretment filure, especilly in fvourble groups with mild fibrosis or prior relpse. ART ws useful for predicting tretment outcome only for difficult-to-tret groups, such s prior null response or dvnced fibrosis/cirrhosis [13]. Another merit of ART is tht HCV RNA TD/<LLOQ during the tretment stge with only PEG-IFN-/RBV (without DAA fter 12 weeks of triple therpy) indicted relpse, irrespective of HCV RNA TND by CAP/CTM [13]. The SOF/LDV regimen hs resulted in remrkbly high SVR rte, over 9% for ptients with genotype-1, even in rel-world clinicl prctice nd subgroups of ptients such s those with prior tretment-experience or who re cirrhotic [14,15]. The vlue of HCV RNA mesurement t specific points during the new tretments is in ensuring ptient dherence: high SVR rtes hve mde it less importnt to predict tretment outcome from on-tretment virl kinetics. However, it continues to be importnt to discuss the utility nd sensitivity of the HCV RNA ssys. In this study, we simultneously determined the HCV RNA level by both CAP/CTM nd ART t 2-week intervls during tretment nd rised three importnt findings in the detiled nlysis of our dt. First, HCV RNA remined detectble by ART for longer durtion during SOF-bsed tretment, irrespective of the HCV genotype. Notbly, the rtes of HCV RNA TND t week 4 were extremely different between the two ssys (genotype-1, CAP/CTM 82.1% nd ART 31.5%: genotype-2, CAP/CTM 85.7% nd ART 35.%; both P<.1). Msoumy et l. [17] provided similr results, but their rtes of HCV RNA TND t week 4 of SOF/DCV ± RBV (CAP/CTM 51% nd ART 1%) nd SOF/SMV ± RBV (CAP/CTM 55% nd ART 14%) therpy were lower thn ours for SOF/LDV. Nevertheless, the HCV RNA TND rtes t weeks 8 nd 12 were similr to our results. Second, detectble residul HCV RNA (TD/<LLOQ) t EOT (week 12) ws found for 11.6% of our genotype-1 ptients nd for 14.3% of those with genotype-2, irrespective of prior tretment experience or cirrhosis; however, ll of them chieved SVR12: this ws not ssocited with relpse by either SOF/LDV or SOF/RBV therpy. In two genotype-2 ptients, residul HCV RNA remined detectble t week 4 fter the end of tretment, but both chieved SVR12. Recently, similr results were reported by Sidhrthn et l. [16] for 6-week SOF-bsed tretment of tretment-nive ptients. This is mjor difference from IFN-bsed regimens in tht detectble residul HCV RNA t EOT ws, Antivirl Therpy

8 E Ogw et l. Tble 6. Predictive vlues of undetectble HCV RNA by ART during SOF/RBV therpy for HCV genotype-2 ptients (week 2 to 6) Week 2 Week 4 Week 6 TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ Number Sensitivity, % (n/totl n) 12. (6/5) 88. (44/5) 36. (18/5) 64. (32/5) 64. (32/5) 36. (18/5) Specificity, % (n/totl n) 83.3 (5/6) 16.7 (1/6) 33.3 (2/6) 66.7 (4/6) 33.3 (2/6) 66.7 (4/6) PPV, % (n/totl n) 85.7 (6/7) 89.8 (44/49) 81.8 (18/22) 94.1 (32/34) 88.9 (32/36) 9. (18/2) NPV, % (n/totl n) 1.2 (5/49) 14.3 (1/7) 5.9 (2/34) 18.2 (4/22) 1. (2/2) 11.1 (4/36) ART, Abbott RelTime HCV ssy; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/RBV, sofosbuvir/ribvirin; TD, trget detected; TND, trget not detected. Tble 7. Predictive vlues of undetectble HCV RNA by ART during SOF/RBV therpy for HCV genotype-2 ptients (week 8 to 12) Week 8 Week 1 Week 12 TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ TND TD/<LLOQ, LLOQ Number Sensitivity, % (n/totl n) 72. (36/5) 28. (14/5) 76. (38/5) 2. (1/5) 84. (42/5) 16. (8/5) Specificity, % (n/totl n) 16.7 (1/6) 83.3 (5/6) (/6) (6/6) (/6) (6/6) PPV, % (n/totl n) 87.8 (36/41) 93.3 (14/15) 87. (4/46) (1/1) 87.5 (42/48) (8/8) NPV, % (n/totl n) 6.7 (1/15) 12.2 (5/41) (/1) 13. (6/46) (/8) 12.5 (6/48) ART, Abbott RelTime HCV ssy; EOT, end of tretment; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/RBV, sofosbuvir/ribvirin; TD, trget detected; TND, trget not detected. Tble 8. Predictive vlues of unquntifible HCV RNA by ART during SOF/RBV therpy for HCV genotype-2 ptients (week 2 to 6) Week 2 Week 4 Week 6 TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ Number Sensitivity, % (n/totl n) 54. (27/5) 46. (23/5) 82. (41/5) 18. (9/5) 94. (47/5) 6. (3/5) Specificity, % (n/totl n) 66.7 (4/6) 33.3 (2/6) 16.7 (1/6) 83.3 (5/6) 16.7 (1/6) 83.3 (5/6) PPV, % (n/totl n) 93.1 (27/29) 85.2 (23/27) 89.1 (41/46) 9. (9/1) 9.4 (47/52) 75. (3/4) NPV, % (n/totl n) 14.8 (4/27) 6.9 (2/29) 1. (1/1) 1.9 (5/46) 25. (1/4) 9.6 (5/52) ART, Abbott RelTime HCV ssy; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/RBV, sofosbuvir/ribvirin; TD, trget detected; TND, trget not detected. Tble 9. Predictive vlues of unquntifible HCV RNA by ART during SOF/RBV therpy for HCV genotype-2 ptients (week 8 to 12) Week 8 Week 1 Week 12 TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ TND, TD/<LLOQ LLOQ Number Sensitivity, % (n/totl n) 98. (49/5) 2. (1/5) (5/5) NA (5/5) NA Specificity, % (n/totl n) (/6) (6/6) (/6) NA (/6) NA PPV, % (n/totl n) 89.1 (49/55) (1/1) 89.3 (5/56) NA 89.3 (5/56) NA NPV, % (n/totl n) (/1) 1.9 (6/55) NA NA NA NA ART, Abbott RelTime HCV ssy; EOT, end of tretment; LLOQ, lower limit of quntifiction; NPV, negtive predictive vlue; PPV, positive predictive vlue; SOF/RBV, sofosbuvir/ribvirin; TD, trget detected; TND, trget not detected Interntionl Medicl Press

9 HCV RNA monitoring of sofosbuvir-bsed therpy without exception, relpse sign in DAA-bsed triple therpy. A possible reson for this phenomenon cn be explined by the improvement of the host immune system due to the decline of the HCV RNA level [22,23]. Additionlly, the CAP/CTM ssy hs AmpErse (urcil-n-glycosylse) enzyme, which removes ny potentil contminting DNA from previous mplifictions; ny non-specific product is destroyed by the AmpErse enzyme, thus improving sensitivity nd specificity. Third, the time t which HCV RNA becme undetectble or unquntifible ws not ssocited with tretment outcome by either the ART or CAP/CTM ssy. In fct, PPV ws very high by both ssys t ll points during tretment. There is lck of dt evluting whether or not tretment durtion cn be individulized using on-tretment HCV kinetics, especilly for IFN-free regimens. However, frequent HCV RNA monitoring during SOF-bsed tretment my not be useful in predicting tretment outcome or for the determintion of tretment durtion in response-guided therpy, including for ptients with cirrhosis nd/or those with tretment experience, irrespective of the sensitivity of the HCV ssy. We were ble to provide detiled dt on the kinetics of HCV by the use of two rel-time ssys. Moreover, ll studied ptients were treted with the sme SOF-bsed regimens for fixed tretment durtion, ccording to their HCV genotype. However, the limittions of this study include its reltively smll popultion with tretment filure. We were ble to highlight the importnce of HCV RNA monitoring, but further exmintion in lrger trils will be needed to evlute the clinicl utility. In conclusion, the period of HCV RNA TD/<LLOQ ws significntly longer when mesured by ART thn by CAP/CTM for ptients treted with SOF/LDV or SOF/ RBV, irrespective of tretment-experience or cirrhosis. Furthermore, over 1% of the ptients remined HCV RNA TD/<LLOQ t EOT; however, prolonged HCV RNA TD/<LLOQ ws not ssocited with tretment filure. Contrry to tretment regimens with PEG-IFN/RBV, becuse of the high success rtes frequent monitoring of the HCV RNA level of ptients treted with DAAs my hve limited utility in predicting tretment outcome nd for guiding tretment durtion, irrespective of the sensitivity of the HCV ssy used. Acknowledgements We re grteful to Tkeo Hyshi, Fujiko Mitsumoto- Kseid, Koji Tkym, Kzuy Ur nd Sho Ymski for their ssistnce with dt collection nd to Yoshitk Etoh for his excellent lb work from the Deprtment of Generl Internl Medicine, Kyushu University Hospitl. We re lso grteful to Dongchon Kng from the Deprtment of Clinicl Chemistry nd Lbortory of Medicine, Kyushu University Hospitl. Disclosure sttement NF hs received grnts from Tisho Toym Phrmceuticl Co., Diichi Snkyo Co., Chugi Phrmceuticl Co., Mitsubishi Tnbe Phrm Co., Bristol Myers Squibb nd Jnssen Phrmceuticl KK. The other uthors declre tht they hve no conflicts of interest. This work ws supported by JSPS KAKENHI Grnt Number 15K Additionl file Additionl file 1: Supplementry informtion cn be found t documents/3912_ogw_addfile1.pdf References 1. Perz JF, Armstrong GL, Frrington LA, et l. The contributions of heptitis B virus nd heptitis C virus infections to cirrhosis nd primry liver cncer worldwide. J Heptol 26; 45: Alter HJ, Seeff LB. Recovery, persistence, nd sequele in heptitis C virus infection: perspective on long-term outcome. Semin Liver Dis 2; 2: AASLD/IDSA HCV Guidnce Pnel. Heptitis C guidnce: AASLD-IDSA recommendtions for testing, mnging, nd treting dults infected with heptitis C virus. Heptology 215; 62: Europen Assocition for Study of Liver. 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10 E Ogw et l. 13. Ogw E, Furusyo N, Murt M, et l. Impct of HCV kinetics on tretment outcome differs by the type of reltime HCV ssy in NS3/4A protese inhibitor-bsed triple therpy. Antivirl Res 216; 126: Bckus LI, Belperio PS, Shhoumin TA, et l. Rel world effectiveness of ledipsvir/sofosbuvir in 4365 tretmentnive genotype 1 heptitis C infected ptients. Heptology 216; 64: Ionnou GN, Beste LA, Chng MF, et l. Effectiveness of sofosbuvir, ledipsvir/sofosbuvir, or pritprevir/ritonvir/ ombitsvir nd dsbuvir regimens for tretment of ptients with heptitis C in the Veterns Affirs Ntionl Helthcre System. Gstroenterology 216; 151: Sidhrthn S, Kohli A, Sims Z, et l. Utility of heptitis C virl lod monitoring on direct-cting ntivirl therpy. Clin Infect Dis 215; 6: Msoumy B, Vermehren J, Welker MW, et l. Clinicl vlue of on-tretment HCV RNA levels during different sofosbuvir-bsed ntivirl regimens. J Heptol 216; 65: The French METAVIR Coopertive Study Group. Introbserver nd interobserver vritions in liver biopsy interprettion in ptients with chronic heptitis C. Heptology 1994; 2: Ogw E, Furusyo N, Toyod K, et l. The longitudinl quntittive ssessment by trnsient elstogrphy of chronic heptitis C ptients treted with pegylted interferon lph-2b nd ribvirin. Antivirl Res 29; 83: Furusyo N, Ogw E, Murt M, et l. Therpeutic drug monitoring of telprevir in chronic heptitis C ptients receiving telprevir-bsed triple therpy is useful for predicting virologicl response. J Antimicrob Chemother 214; 69: Ogw E, Furusyo N, Kjiwr E, et l. Comprtive effectiveness nd sfety study of triple therpy with simeprevir or telprevir for non-cirrhotic ptients with chronic heptitis C virus genotype 1b infection. J Gstroenterol Heptol 215; 3: Meissner EG, Wu D, Osinusi A, et l. Endogenous intrheptic IFNs nd ssocition with IFN-free HCV tretment outcome. J Clin Invest 214; 124: Mrtin B, Hennecke N, Lohmnn V, et l. Restortion of HCV-specific CD8 + T cell function by interferon-free therpy. J Heptol 214; 61: Accepted 7 September 216; published online 15 September Interntionl Medicl Press