SNP rsl2979860 related spontaneous clearance of hepatitis C virus infection in HCV/HIV-1 coinfected patients - Epidemiological Review

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17DGHXV]:RMFLHFKàDSLĔVNL1_{Joanna Pogorzelska,} 2_{Oksana Kowalczuk,} 2-DFHN1LNOLĔVNL1_{Robert Flisiak}

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1_{Department of Infectious Diseases and Hepatology, Medical University of Bialystok} _{Department of Clinical Molecular Biology, Medical University of Bialystok}

ABSTRACT

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7+($,0RIWKLVDQDO\VLVZDVHYDOXDWLRQRILPSDFWRIUVSRO\PRUSKLVPRQ+,9RU+&9YLUDOORDG &'&'DQG&'FRXQWDVZHOODV+&9FOHDUDQFHDPRQJ+&9+,9FRLQIHFWHGSDWLHQWV

0$7(5,$/6$1'0(7+2'6The study included 41 consecutive HCV/HIV coinfected patients. HIV RNA, +&951$+&9JHQRW\SHDQGUVSRO\PRUSKLVPVHTXHQFHDVZHOODV&'&'DQG&'FHOOVFRXQW were analyzed in all patients.

5(68/76&&JHQRW\SHUVZDVLGHQWLILHGLQIURPSDWLHQWV'XULQJDWOHDVW\HDUVIROORZXS ILYHJHQRW\SH&&SDWLHQWVEHFDPH+&951$XQGHWHFWDEOHWKDWZDVQRWDFDVHLQ&7DQG77SDWLHQWV 1RVWDWLVWLFDOGLIIHUHQFHVLQ+,9YLUDOORDGDQGWKHQXPEHURI&'&'DQG&'UHODWHGWRUVSRO\-PRUSKLVPZHUHREVHUYHG7KHEDVHOLQHOHYHORI+&951$LQSDWLHQWVZLWK&&JHQRW\SHZDVVLJQLILFDQWO\ORZHU FRPSDUHGWRSDWLHQWVZLWKQRQ&&JHQRW\SHV“YV“,8P/ &21&/86,21&&JHQRW\SHUHODWHGWR613UVFDQDIIHFWWKHORZHUOHYHORI+&9YLUDOORDGFRPSDUHG to patients with CT and TT genotypes and promotes spontaneous clearance of HCV RNA in HCV/HIV coinfected patients.

.H\ZRUGVHIV/HCV coinfection, SNP rs12979860, HCV clearance

INTRODUCTION

+&9LQIHFWLRQLVHVWLPDWHGWRDIIHFWRIWKH ZRUOGZLGHSRSXODWLRQ2QHRIWKHPRVWLPSRU-tant issue in era of new anti-HCV agents is prediction of virologic response or HCV spontaneous clearance EDVHGRQEDVHOLQHRURQWUHDWPHQWHYLGHQFHV)UH-quent spontaneous elimination of HCV is associated with single nucleotyde polymorphism (SNP) at site UVRIFKURPRVRPH+&9PRQRLQIHFWHG patients with CC genotype demonstrate spontaneous HCV clearance more often than those with genotype CT or TT (4). Higher efficacy of treatment with pegylated interferon alfa (PegIFND DQG ULEDYLULQ 5%9 ZDV GHPRQVWUDWHG LQ ERWK +&9 PRQRLQIHFWHG DQG +,9 HCV coinfected genotype CC patients compared to non-CC patients (5).

The aim of this analysis was evaluation of SNP

UVSRO\PRUSKLVPRQVSRQWDQHRXV+&9FOHDU-ance, HCV and HIV viral load, as well as CD3, CD4 DQG&'FRXQWLQ+&9+,9FRLQIHFWHGSDWLHQWV

MATERIALS AND METHODS

The study included 41 consecutive patients (10 IHPDOHVDQGPDOHVDJHGIURPWR\HDUVZKR were diagnosed as infected with HIV and positive for anti-HCV. Patients who did not received anti-HCV or ART (Antiretroviral Therapy) treatment were followed for at least 4 (mean: 5,6 r) years. HIV RNA and HCV RNA concentrations were measured, HCV geno-type was determined as well as lymphocytes CD3, CD4 DQG&'FRXQWVZHUHDVVHVVHGDWEDVHOLQH 'LDJQRVLVRI+,9LQIHFWLRQZDVEDVHGRQGHWHFWLRQ WKH+,9DQWLERGLHVZLWK(/,6$DQG:HVWHUQEORWWHVWLQJ &DPEULGJH%LRWHFK&RUSRUDWLRQ86$+,9YLUDOORDG ZDVGHWHUPLQHGE\573&5XVLQJWKH&REDV$PSOLFRU +,68OWUD6HQVLWLYH6XESRSXODWLRQVRI&'&'

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 No 3

DQG&'LQEORRGZHUHDQDOL]HGE\IORZF\WRPHWU\XV-LQJ%HFWRQ'LFNLQVRQLQVWUXPHQW+&951$OHYHO and its genotype were determinated with RT-nested-3&56\QJHQ%LRWHFK86$ZLWKVHQVLWLYLW\RI,8 P/5VSROLPRUSKLVPVHTXHQFHGHWHFWLRQZDV FDUULHGRXWE\GLUHFW3&5SURGXFWVHTXHQFLQJ'1$ZDV LVRODWHGIURPWKHSHULSKHULDOEORRGXVLQJWKHUHDJHQWVHW and automatic device for nucleic acid isolation EasyMag (Biomereueix, France) according to the manufacturer’s protocol. Briefly, 50 PORIZKROHEORRGZDVPL[HGZLWK 500 Pl of the Lysis Buffer (Biomereueix, France) and LQFXEDWHGDW0_{C for 1 h with 30} Pl of Proteinaza K solution (10 mg/ml, Sigma). Proteolysis was followed E\'1$ELQGLQJWRWKHDGGHGPDJQHWLFEHDGV6LOLFD ((Biomereueix, France), washing and eluting into the 50 Pl of the Washing Buffer 3 ((Biomereueix, France). $OOWKHVWHSVRISURWHRO\]HGEORRGSURFHVVLQJZHUHSHU-formed automatically in EasyMag machine. A portion RI,/%JHQHIODQNLQJWKHSROLPRUSKLFVLWHRILQWHU-HVWZDVDPSOLILHGLQ3&5UHDFWLRQLQPl of reaction PL[WXUHFRQWDLQLQJ[3&5EXIIHUPM dNTPs, 0,5 PM each of sense and antysense primers, 1 U TaqRed '1$3RO\PHUDVH(85[3RODQG*GDQVNDQGDERXW 50 ng of DNA. The primer sequence was as follows: sense 5’-GCTTATCGCATACGGCTAGG-3’, antisense 5’-AGGCTCAGGGTCAATCACAG-3’. Cycle condi-tions for PCR reaction were as follows: after initial hold for 5 minutes at 950_{C, the samples were cycled} 40 times at 950_{C for 15 seconds, 60}0_{C for 1 minute and} 0&IRUVHFIROORZHGE\WKHWHUPLQDO3&5SURGXFWV H[WHQVLRQIRUPLQDW0_{C. PCR products were} ana-lyzed on agarose gel, excited from the gel and purified with the “Gel-Out” set of reagents (A&A Biotechnol-ogy, Poland, Gdansk). PCR products sequencing were performed with “Big Dye Terminator v3.1 Sequencing Standard Kit” and 3500 Genetic Analyzer (Applied Biosystems, Foster City, USA).

7KHVWXG\ZDVDSSURYHGE\WKH(WKLFDO&RPPLWWHH RI0HGLFDO8QLYHUVLW\LQ%LDá\VWRN,QIRUPHGFRQVHQW LQFOXGLQJDSSURYDOIRUJHQHWLFVWXGLHVZDVREWDLQHG from each patient.

Statistical analysis was performed with Mann-:KLWQH\ WHVW9DOXHV RI S ZHUH FRQVLGHUHG DV statistically significant. The Statistics 10.0 for Windows was applied to carry out the analysis (Statsoft Inc., Tulsa, USA).

RESULTS

CC genotype was identified in 16 patients, while LQ&7DQGLQ77UVSRO\PRUSKLVPZDV REVHUYHGTwenty-two patients were infected with genotype 1 HCV, seven with genotype 4, and seven with genotype 3. During follow-up period HCV-RNA

EHFDPH XQGHWHFWDEOH LQ ILYH JHQRW\SH && SDWLHQWV (31%). Among the five patients who cleared HCV RNA, genotype was not specified. In contrast HCV RNA FOHDUDQFHZDVQRWREVHUYHGLQDQ\QRQ&&SDWLHQWV %DVHOLQH+&951$OHYHOLQ&&SDWLHQWV“ IU/mL) was significantly lower than in non-CC patients “,8P/S $VVKRZQLQWDEOH, mean HIV RNA was more than ten times lower in CC FRPSDUHGWRQRQ&&SDWLHQWVEXWWKHGLIIHUHQFHZDVQRW VWDWLVWLFDOO\VLJQLILFDQW“YV“ IU/mL) (Fig1). There were no significant differences EHWZHHQ&&YHUVXVQRQ&&SDWLHQWVUHJDUGLQJ&' &'DQG&'FRXQW7DE

7DEOH, Mean (rSE) BMI, HIV RNA, HCV RNA and CD3, &'DQG&'O\PSKRF\WHVVXEVHWVLQSDWLHQWVZLWK CC versus non-CC (CT or TT) genotypes.

Parameter 613UVJHQRW\SHV p &&Q  QRQ&&Q 

BMI r r 

HIV RNA, (IU/mL) r r  HCV RNA (IU/mL) r r  CD3 Pl) 1010 r151 1161 r194  CD4 Pl) r r46  &'Pl) r r  DISCUSSION 7KHILUVWSXEOLFDWLRQVGHPRQVWUDWLQJEHQHILFLDOHI-IHFWVRI&&JHQRW\SHRIUVSRO\PRUSKLVPLQ spontaneous clearance of hepatitis C virus and efficacy of HCV genotype 1 treatment in patients with HIV/HCV FRLQIHFLRQZDVSXEOLVKHGLQ+RZHYHUSXEOLFD-tions on spontaneous clearance of HCV infection among HCV/HIV infected patients are sporadic. SajadiHWDO SRLQWHGRXWODFNRIDVVRFLDWLRQEHWZHHQ+,9YLUDOORDG DQGUVJHQRW\SHVDQGFRQVLGHUHGSRVVLELOLW\ of spontaneous HCV clearances among HCV/HIV co-infected patients, as related to CC genotype. Results in WKLVSDSHUDUHLQOLQHZLWKWKHVHREVHUYDWLRQV:HGHP-onstrated nine times lower HCV viral load in CC patients FRPSDUHGWRQRQ&&WKDWVXJJHVWSRVVLEOHWUHQGWRYLUDO clearence. Therefore we can assume that CC genotype RIUVSRO\PRUSKLVPPD\EHDQXVHIXOEDVHOLQH predictor of HCV clearance in HCV/HIV coinfected pa-WLHQWV+RZHYHUWRFRQILUPWKLVFRQFHSWSURVSHFWLYH VWXGLHVVKRXOGEHFDUULHGRXWLQWKLVSRSXODWLRQLunge et DOREVHUYHGVSRQWDQHRXVFOHDUDQFHVRI+&9LQ Brazilians living with HIV/HCV, that was significantly more frequent in genotype CC patients. Clausen et al. GHVFULEHGIRUWKHILUVWWLPHLQHXURSHDQVIUHTXHQW spontaneous HCV clearances in HCV/HIV co-infected SDWLHQWVZLWK&&JHQRW\SHV

SNP rs12979860 related spontaneous clearance of HCV ₄₀₉

No 3

CONCLUSION

7KH && JHQRW\SH RI UV SRO\PRUSKLVP may affect the HCV viral load lower compared to pa-tients of the CT and TT genotypes, and is conducive to a spontaneous elimination of HCV infection in patients ZLWK +,9  +&9 FRLQIHFWHG 5V SRO\PRU-phisms is not associated with BMI, HIV viral load and O\PSKRF\WHVVXESRSXODWLRQVFRXQW

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 6DMDGL006KDNHUL17DOZDQL5HWDO,/%JHQR-type does not correlate with HIV control in African $PHULFDQV&OLQ7UDQVO6FL>3XE0HG @

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 /XQJH95GD5RFKD'%HWDO,/%SRO\PRUSKLVP associated with spontaneous clearance of hepatitis C infection in a Southern Brazilian HIV type 1 popula-WLRQ$,'65HV+XP5HWURYLUXVHV >3XE0HG@

 &ODXVHQ/1:HLV1$VWYDG.HWDO,QWHUOHXNLQ% polymorphisms are associated with hepatitis C virus clearance and viral load in a HIV-1-infected cohort. J 9LUDO+HSDWH>3XE0HG@ 5HFHLYHG $FFHSWHGIRUSXEOLFDWLRQ $GGUHVVIRUFRUUHVSRQGHQFH: 7DGHXV]:RMFLHFKàDSLĔVNL %LDá\VWRNXOĩXUDZLD e-mail twlapinski@wp.pl