JOURNALOF 108-CLINICALMEDICINEANDPHARMACV Vol 7 - N°4/20I2
Nghien cdu xay dung quy trinh xac dinh tinh da hinh gene IL28B va budc d k khao sat gene IL28B d benh nhan viem gan C
Setting up a New Procedure for Identifying tlie Polymorphisms of IL28B Gene and Primary Result of IL28B Gene Polymorphism in Hepatitis C
Ngo Tat Trung', Oao Phuwng Giang*, Lc Huu Song* 'Bfnh vifn Tmng tmng Qudn dfim Phan Anh HoSng** Bfnh vifn QuanY 175
Hieu qui dieu trj viem gan C dupc cho l i cd lifin quan d i n tinh da hinh ciia gene IL28B, tuy nhifin hifn nay d Vift Nam van ehua cfi sfi lifu nao ve gene IL28B dupc cfing bfi. Mgc tiiu: (I) Xiy dpng quy trinh xac djnh d c t h i da hinh rs8099917, rsl2980275 v i rsl2979860 ciia gene IL28B; (li) Khio sit tinh da hinh gene iL28B tgl d c vj tri nfii trfin trong nhdm tsfnh nhin vifim gan C v i nhdm ngudi Vift Nam khde mgnh. Ddi tUOng: 38 bfnh nhin vifim gan C v i 32 ngudi khde manh dupc dUa vao nghifin cUu niy. Phuong phip: Tinh da hinh eua gene IL28B dupc xac dinh bing phucmg phap dpng hpc allele die hifu real-time PCR. Ket qui: D i hoin thifn ky thuft dfing hpc allele die hifu real-time PCR cho phfip phat hien dfing thdi 3 the da hinh rs8099917, rsl 2980275, rsl 2979860 eua gene OSB. 0/70 (0%) d t h i mang kilu allele hiem cQa 3 SNP. Tren ci 3 vj tri quan sit thi tJ lfi ca t h i mang kieu gene dfing hop tQ trpl chilm chii y l u vdi tJ If Iln luot la:
rs8099'»17 (90%), rsl2980275 (79%) v i rs12979860 (79%). Ty If d the mang gene dj hdp tQ Iln lUdt II rs8099917 (10%), rs12980275 (21%) va rs12979860 (21%). Kilu allele trfl T/T ciia SNP rs8099917 d nhdm viem gan C cao hdn cd y nghla so vdi nhdm ngUdi khde mgnh (100% so vdi 79%, p <0,01). Trong khi do khdng cd khic biet ve t i n suit allele d a 2 SNP rs12980275 va rsl 2979860 giQa 2 nhdm nghien cQu. Kit ludn: Ky thuft xic djnh tinh da hinh ge le OSB d l dUOC thilt Ifp thinh cdng va tan suit d c the da hinh gene OSB trfin cpng ddng din eU Vifit Nam d i budc dai dupc ghi nhgn.
Tif khoa: HCV, IL28B, dpng hoe allele dgc hifu real-time PCR.
Summary
Introduaion: Single nucleotide polymophisms (SNPs) near the gene OSB is considered as the most Important host factor that defines the success of Peg-INF alpha/ribavirln based regiment to eradicate HCV. A/m; i) To set up a protocol to screen for rs8099917, rs12980275 and rs12979S60 near OSB gene ii) and statistcally recoid the epWemologKs of IL28B SNP within Vietnamese healthy subjects and patients with C viral Hepatitis. Subjects: genomics DNA samprfe from 32 healthy donors, 38 patients with C viral Hepatitis. Methods: Allele specific keneties real-time PCR. primers sets optimized for rs8099917, rsl2980275 and rsl2979860 SNP were designed. Results: Allele specifie keneties real-time PCR was susessfully established, which allow us to simutaneously detect all three inspected SNP in one njn. Our primarily data revealed 0/70 (0%) carries homozygous minor alleles; 90%, 79% and 79% are homozygous major allele of rs8099917, rsl2980275 and rsl2979860 respeaively. The percentage of heterogenous allele cartiers Is rs8099917 (10%), rs1298027S (21%) and rs12979860 (21%). We also observed the frequency of rs8099917 major allele a/Tl in patients with C viral Hepatitis was significantly higher than that in healthy control group (100% vs 79%, p<0,01), whereas there was no allelk frequency difference of rsl2980275 and rsl2979860 between the two study groups. Conclusion: An income-appropriate- IL28B screen assay has recapitulated, which helped to record a statistical difference of SNP rs8099917, rs12980275, rsl 2979860 frequencies within Vietnamese healthy subjects and patients with C viral Hepatitis.
Keywords: HCV, OSB, kinetics allele specifie real-time PCR
Phin bifn khoa hpc: PGS.TS, TRINH TUAN DUNG 102
T i p 7-So 4/2012
L O i t v i n d e
Hifn nay, cd khoing 3% dan so t h i gidi, tUOng dng vdi khoing t r i n 170 trieu ngudi mang virus v i l m gan C (HCV). Tuy nhifin, chl khoing 20-30% trong sfi dfi cd the tu khfil bfnh, so cdn Igi se p h i t t r i l n thanh v i l m gan man tinh, xd gan va ung t h u gan [1]. Ngiy nay vifc sd dung Peg-INF alpha (beta) k i t hpp vdi ribavirin dupc col la mfit phac dfi chuan dupe c h i p n h f n rfing r i i trong d i l u trj benh nhin v i l m gan C [2]. TrQdc diy, ngudi ta cho ring hieu q u i d i l u tri phu thufic v i o k i l u gene HCV, trong dd, kilu gene 2,3 cd t i l n lupng d i p Qng tfit han so vdl k i l u gene 1,6. Gin day, ngUdi ta thay y l u to di truyin cua cd the chii, dgc b i f t I I gene 1L28B efi I n h hudng r i t ldn d i n hieu qui d i l u tri v i t i l n lUpng benh v i f m gan C [3-6].
Gene OSB cd d c diem da hinh ddn (SNP) nhU:
IS12980275, rs8105790. rsl 1881222, rs8103142, rs28416B13, rs4803219, rsS099917 v l rs7248668 [3-6]. Nghiin cdu cho thay tiiy thupc v i o ele chQng tfic ngucfl khic nhau m i t i n suat ciia d e SNP nfii tren cung khic nhau. Tuy nhien, i n h hufing ciia chung tdi d i p dng d i l u trj eua bfinh nhin mang HCV vdi phle do Peg-INF alpha /ribavirin la tUdng dfii giong nhau: eac benh nhin mang dfing hpp td t h i h i l m (homozygous minor allele) dUOc tien iupng dap Qng dieu tri kem hole khdng d i p Qtig d i l u trj vdi Peg-INF alpha /ribavirin. NgUpe lai, b l n h nhan mang ddng hop t d the trdi (homozygous major allele) lai cd d i p iing rat tfit vdi phic do Peg-INF alpha /ribavirin va bfinh nhin dj hop tQ (heterozygous allele) se co d i p dng vda p h l i vdl phic do nay. Trong eac t h i da hinh cua O S B ndi trfin thi 2 SNP rs8099917 v i rsl 2980275 cd lien quan chat che vdi dap dng dieu trj d bfnh nhan chiu A. Trting khi dd SNP rsl2979860 lai CO Inh hUdng tdi k h i n i n g d i o thai t y nhifin HCV cua benh nhin goc chau Au, chau Phi v i Ble My [3,6].
Do dd, cd dupe thdng tin ro ring v l d c t h i da hinh gene IL28B mdt mat giup eho d c bac sy l i m sing v l benh nhin d n nhic nguy ca v i lpi Ich cua vifc sd dung Peg-INF alpha /ribavirin trong dieu trj HCV, mat khie eung tifin luong kha n i n g t h i n h cfing cho tdng b l n h n h i n rifing bift {7]. O i l u dfi da dUpc thong nhat t h i n h van b i n dong thuan cua Hifip hpi n g h l l n cdu gan chau Au trong chi djnh xet nghilm t h i da hinh gene IL28B trudc khi d n ' nhac d e pha do d i l u trj cho benh nhan vifim gan C [8].
d Vift Nam chUa cfi mdt nghien cdu he thfing n i o j md t i tan suit d e the da hinh gene IL2SB, eung nhu chua , CO nghifn cdu n i o de c f p d i n I n h hudng ciia d c the da r hinh tren gene IL2BB doi vdi k h i nang t y d i o t h i i HCV hoac d e mdc d i p yng dieu trj vdi p h i c do Peg-INF alpha /ribavirin. Vl vay, chiing tfii tien n g h l l n cdu n i y n h i m tim hieu tan suit xuat h i f n 3 the da hinh chQ y l u cQa gene IL28B rs8099917, rsl 2980275 v l rsl 2979860 t r i n q u i n thi ngudi khfie manh v l benh n h i n viem gan C.
2. Ofii tupng v i phUdng p h i p nghien cdu 2.1. Ddi tuang: 38 bfnh nhan viem gan C v i 32 ngudi khde mgnh.
2.2. Vdt lieu: 2X Sybr green I master mix, cic bd mfii die hifu cho d c allele ciia tdng the da hinh rs8099917, rsl 2979860 dUpc tiep thu tQ nghlfin cdu cua Thomas R.
O'Brien v i cdng sy [9], chung tdi thiet k l thfim bd moi die hifu cho rsl2980275 nhQ sau: rs12980275 G allele Fon 5'- AGAAGTCAAATTCCTAGAAACG-3'; rsl 2980275 A allele For:
5'- GAGAAGTCAAATTCaAGAAACA-3'; rsl 2980275- Common.Re 5'-GAATaaGTACCCATTAAACAACAAa-3'.
Ngoai ra, cdn cd d c hda chit d i m bio dp tinh sach phuc vy d c ky t h u i t sinh hpc phin tQ.
2.3. Phuang phdp: Tinh da hinh ciia gene IL28B dUpc xle djnh theo nguyfin IJ ddng hoc allele dac hieu real-time PCR. Phuang phip niy l i do Ssren Germer [10] thilt lap Iln d i u tifn vio nam 2000 v i dQpc Thomas R. O'Brien ciing cdng sQ sQ dyng trong phin tich t h i da hinh gene IL28B [9]. Tuy nhiln, Thomas R. O'Brien va cfing su chi sQ dung no cho 2 vj tri da hinh la rs8099917 va rs12979860, cdn chung tdi thilt ke thfim d e primer die hifiu eho d vj tri rsl 2980275. Day l i vj tri dac trung cho quan the chiu A, do do rit eIn thiet dUdc trien khai tgi Viet Nam [5].
Nguyen IJ ciia phUdng phap n i y dUpc tdm t i t nhu sau: 2 bp moi d i e h i f u cho tQng allele diiac thiet k l sao eho dau 3' cQa moi xudt (forward primer) bat cap ehinh xac vdi hai allele rieng b i f t tgi vi tri cfi SNP, trong khi dfi moi ngUpe (reverse primer) sfi dUpc dung chung cho d 2 allele.
Tgi mfii mfit vi tri SNP, sfi ton tai 1 trong 3 trinh ty nucleotide nhU sau: dong hpp tQ allele trdi (homozygous major alleles), dj hpp tQ (heterozygous) va ddng hpp tQ allele hiem (homozygous minor alleles).
Khi t i l n h i n h ehan doan SNP. tien hinh dong thdi 2 p h i n Ung real time PCR (sd dyng Sybr green I d l nhin b i l t sin pham PCR) bat d p tUOng Ung vdi 2 allele tgl vi tri nghi ngd cfi SNP. Luc dd sfi xiy ra 2 k h i nang: i) neu mau DNA l i dfing hpp td, tQ tin hifu huynh quang (tren Iy thuyet) chi duoc ghi nhin d duy nhit 1 allele tuong Qng.
Tuy nhien, trong thyc hinh se xiy ra hifn tupng bat cap lech (mismatch) eiia primer, do do ngay e l khi m l u DNA I I dong hpp tQ thi tin hifu huynh quang van xuat hifn d c l 2 allele trong dd tin hifiu gta se ehim hon tin hifiu that khoing t d 3 d i n IS chu ky (hinh 1- panel trai) [9,10], ii) trong trudng hpp mau DNA la dj hop td, tin hifiu huJnh quang se cd mat dfing deu fi e l hai alen, dudng cong pho huJnh quang eua 2 allele se gan trung kbit nhau hole chl khic nhau dUdi 1 chu ky (fiCt s l ) - (hinh 1) [9,10].
103
)UURNAI.O^ 1U» I LINKAL Ml n u INt ANDI'HARMACY Vol 7-N'4/20I2
pooled DNA 1 I Allele, Allele2 speciflc specific PCRs PCRs
Specific signal / , Mismatch
signal
2 d 6 a 10 i ; H 16 IB 20 22 24 26 28 30 32 M 36 38 40 42 44 Cycle number
Hinh 1: Co id ly thuyet ciia phuang phap dong hoc allele dgc hiiu real-time PCR do Soren Germer di xudng 19. W]: dau 3' cua moi xuoi (forward primer) bdt ccip chinh xdc vdi nucleotide tai vi tri SNP. De kiem tra sued mat cua nio! allele tgi moi vi tri SNP, ihuc hien dong thai 2 phdn i/ng real - time PCR (tuang Cfng voi 2 bq boi allele 1 specific primer, allele 2 specific primer va nhgn duoc 2 dudng tin hieu huynh quang). Trong irUong hop dong hop til, se xuot hien 2 pho huynh quang rieng biet. dudng sdm (specific signal] mo la tin hieu dac hieu, duang muon mo la tin hieu bat cap lech primer (mismatch signal). Trong tracing hgp di hap til 2 duang cong tin hieu huynh quang se gan nhtf trimg khit len nhau.
3 . K e t q u a
3.7. Hinh dnh cua phu'ang phdp dong hoc allele dgc hieu real-time PCR {kinetics allele specific real-time PCR)
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H i n d 2 : Ket qua xay dang phamg phdp dong hoc allele ddc hieu real-time PCR.
Nhan xet: So v d i ly t h u y e t h i n h a n h c u a c h i i n g t o i t h u d u a c h o a n t o a n p h u h o p va c h o t i n h i e u r a t ro r a n g .
Tap 7-sd 4/2012
C62 5 C/C Homozygous C817 C/T- heterozygous
+ Hmh 3; Kef qua yiai tnnh tugene tai tiNP rsl 29/9860 tren cdc mdu C625 vdCBI7. r Nhdn xet Ket q u i g i i i trlnh tU gene h o i n t o i n phu htfp vdi phuong phip ddng hpc allele dac hieu real-time PCR.
Mau benh pham Cd25 duoc chdn d o i n la ddng hop t d C/C (hinh 2 d i u mui ten panel trii) va m i u C817 duoc chin doan la dj hop t d C/T (hinh 2 d i u mui tdn panel phii) ddi vdi SNP rsl 2979860
3.2. TSn suAalhle cdo cac th^ga hinh rs80999I7, rs12S80275 vd rsl2979a60 iquin thenguii Viet Nam Bing 1: T i n suat xudt hien cdc the' allele SNP cda gene IL28B d quan thd ngudi Vidt Nam
Vi tri gene rs8099917(n=70) rsl2980275(n=70)
rsl2979860(n=70) Allele T/G A/G
G/A
Allele hiem G G
A
Tan suit cic the da hinh T/r(n.%) T/G(n,%) G/G (n,%) 63 (90)
A/A (n.%) 55 (79) G/G (n,%) 55 (79)
7(10) A/G (n,%) 15(21) G/A (n,%) 15(21)
0(0) G/G (n,%) 0(0) A/A (n,%) 0(0)
Nhdn xet: Trong quan the nghien cdu khfing cfi e l the nao mang allele hiem ciia e l 3 vj tri gene khio sat. Trfn d 3 VI tri quan sat thi tJ lfi ca the mang kieu gene dfing hop t d trdi chilm chii y l u vdi tJ I I Ian lupt la: rs8099917 (90%), rsl2980275 (79%) v i rsl2979860 (79%).
3.3. So sdnh tdn sudt allele 3 thi da hinh rs8099917, rs1298027S vd rs 12979860 cda gene IL28B giOfa benh nhdn viim gan C vd ngUcdkhde mgnh
Bing 2: Tan suat xuat hiln cic allele 3 the da hinh cua gene IL28B tren benh nhin viem gan C va ngu'di khfie manh.
Vl tri gene rs8099917(n,%)
rsl 2980275 (n,%)
rsl 2979860 (n,%) Allele T/G
A/G G/A
Nhdm chdng T/T
30(79) A/A 29(76) G/G 29(76)
T/G 8(21) A/G 9(24) G/A 9(24)
G/G 0(0) G/G 0(0) A/A 0(0)
Benh nhan Viem gan C T/T
32(100) A/A 26(83) G/G 26(83)
T/G 0(0) A/G 6(17) G/A 6(17)
G/G 0(0) G/G 0(0) A/A 0(0)
P
<0,01
>0,05
>0,05 Nhdnxet: Kieu allele trfii T/T ciia SNP rs8099917 d nhdm v i l m gan C cao hdn cd J nghia so vdi nhdm ngudi khfie manh. Trong khi dd khfing cfi khac b i f t ve t i n s u i t allele cQa 2 SNP rsl 2980275 va rsl 2979860 giQa 2 nhfim nghien cQu.
3.4. Mdi tuang quan gida kii'u allele tgi cdc vj tri rs8099917,rs12980275vd rsl 2979860
Trong c i nhdm ehdng v i nhfim b f n h n h i n HCV e l t h i nao da mang rsl2980275 thi cung cd rsl2979860 v i ngupe lai. Ogc b i f t , cfi 7/8 trUdng hpp m l u ngUdi khfie manh cd mang SNP d c l 3 vj tri rs8099917, rsl 2980275 v i rsl 2979860 (sfi l i f u khfing trinh biy).
Chdng tfi efi mfit sy tUdng dfing gida d c allele cOa gene nay tgi cic v| tri k h i o sit.
4. Ban l u i n
Thinh cfing cug mfit phic dfi d i l u trj virus nfii chung trong dfi cfi virus v i l m gan C khfing chi phy thufic v i o die diem di truyen hpc dudi l o i i eCia virus (kilu gene virus), cic dot b i l n khing thufic x u i t hifn trong q u i trinh dieu trj, nd cfin phy thufic r i t Idn v i o d c y l u tfi di truyin hpc cCia b i n t h i n c l t h i nhiem virus. G i n d i y nhilu cdng trinh nghifin cdu di truyin hpe q u i n the d l chdng minh moi lien quan chit che giQa t h i da hinh t h i i gene IL28B vdi dap Qng d i l u trj b f n h n h i n v i l m gan C b i n g phle dfi k i t hop Peg-INF alpha /ribavirin [3-6]. CIc nghiin cQu n i y CO ehung k i t luan ring b l n h n h i n mang dfing hpp tQ allele h i l m ciia gene IL2BB sfi efi d i p Qng d i l u trj k i m hon benh nhin mang gene d| hpp tQ v i kfim han nhilu lan so vdi d c b l n h nhin mang dfing hop tQ allele trfii [3- 6]. Vi the vifc co dupc cic thdng tin ro ring l i l n quan d i n t h i da hinh gene IL28B sfi giup cho c l bic sy d i l u trj va bfinh n h i n c i n nhic lpi ich va nguy co t i l m I n khi lua chpn phac do Peg-INF alpha /ribavirin trong dieu trj HCV.
Mae dil ed rat nhieu the da hinh gene IL28B da dupe cdng bo [3-6], trong sfi do cic vj tri rs8099917, rsl 2980275 dUpc eho I I cd lien quan chgt che n h i t tdi d i p dng dieu tri cua bfinh nhan HCV c h i u A vdi p h i c dfi Peg-INF alpha /ribavirin, con rsl 2979860 lai cfi g i i trj t i l n iupng k h i n i n g t y d i o t h l i HCV khdi eo the b f n h n h i n goc Au, Phi. Ble My.
Tuy nhifin. tan suit xuat hifn d e t h i da hinh IL286 tren d e cpng dong d i n cU l i khac nhau v l d i e bift chua cd con sfi thfing kfi d chdng tfic ngudi Vift v l b f n h n h i n mang HCV ngUdi Vift. Oilu n i y khuyen khfch chung tdi thyc hien vifc dieu tra tan suat cQa d c t h i da hinh IL28B t r i n cpng dfing d i n cU Viet Nam va b f n h nhan nhiem HCV tai Viet nam.
Mpt khd khan d i t ra vdi chung tdi la t h i l t lap dupc mdt phuong phap d i n g tin ciy nhUng r l tiln, phu hpp vdi viec trien khai thudng quy xet nghifm the da hinh IL28B trong cpng dfing. Sau khi tim hieu v l uu nhUpc diem cQa cac ky thuat d l dupe thyc hien trude d i y t r i n t h i gidi, chung tfii tien h i n h thiet lap phUdng phap
"Ofing hpc allele dfc h i f u real-time PCR - kinetics allele specific real-time PCR". Phddng p h i p n i y l i do Ssren Germer t h i l t Ifp I l n d i u t i l n v i o n i m 2000 [10]. Nguyen ly ciia nd dUpc mfi phfing d hinh 1. So vdi d c phuong p h i p xle djnh k i l u gene (genotyping) truyen thfing nhU:
d t enzyme gidi hgn (RFLP) hay dpc trinh t y (Sanger sequencing), phuong p h i p m i chiing tfii sd dyng efi cic uu t h i I I : (i) Quy trinh x i t n g h i f m ddn g i i n : n l u nhu RFLP bao gfim 3 bUdc chfnh l i chgy PCR, d t enzyme v i chgy d i f n di agarose cfin dpc trinh t y (Sanger sequencing) phSi d n tdi 2 Iln chgy PCR v i p h i n tich sfi l i f u sgu glli trinh ty. thi kinetics gllele specific real-time PCR chi cd mft budc chgy RT-qPCR; (ii) Rl t i l n hdn m l e dii tfnh trung blnh mfit p h i n dng real-time PCR sfi cfi g l l t h i n h cao hon p h i n iifng PCR thfing thddng. nhUng (RFLP) dfii hfii thfim budc e l t enzyme gidi hgn v i chay d i f n di agarose, nen thyc t l phuong p h i p kinetics allele specific real-time PCR v i n kinh t l hOn.
Mac du kJ t h u f t dfing hpc allele die hifu real-time PCR da dupc Seren Germer t h i l t l i p I l n d i u t i l n v i o nam 2000 [10], tuy n h i l n chua mfit cd sd n i o d Vift Nam lim chii v l dua v i o Qng dung ky thuat niy. vl t h i sau khi da i p dyng t h i n h cdng dUdc phUdng p h i p niy, chung toi t i l n h i n h s i n g Ipc 3 t h i da hinh rsB099917, rsl29S0275 v i rsl 2979860 cua gene IL28B tren ngUdi khfie manh va b l n h n h i n v i l m gan C. K i t q u i dupe trinh b i y d bing 1 v i 2, cho thay vdi 70 dfii tuong ngudi V i l t Nam khfing co - e l t h i nao mang allele hiem cua c i 3 vj tri gene khio sat.
Trin c l 3 vj tri quan sat thi tJ I I c l t h i mang kilu gene dfing hpp t d trfii chiem chii y l u vdi tJ I I Iln lupt la:
rs8099917 (90%), rsl2980275 (79%) v i rsl2979860 (79%).
O l y I I mfit k i t q u i d i u tiln dupe ghi n h i n d cpng dong ngudi Vift Nam. Nfi sfi l i cd sd d d l i f u eho d c nghifn cQu tilp theo khdng chl trong vtf m gan C m i trong c l viem gan B v i d c b f n h do n h i l m virus khic. So vdi cic cpng dong khie tren t h i gidi nhU N h i t Bin, chung tfii thay ty If dfing hpp t d trfii I I tgi v| tri rs8099917 eiia ehung tfil cao hdn (90% so vdi 77,7%) trong khi dd tai vj tri rsl 2979860 thi khfing cfi khic bifit (79% so vdi 76.8%) [11]. Mot n g h i i n cdu tgi Trung Quoc t r i n 56 b f n h n h i n cho thay tJ I I c l t h i mang kieu qene dong hop t d allele trpl tai vi tri rs8099917 ehilm 80.4% (45/56) v l chi efi 19.6% (11/56) e l the cfi allele dj hpp t d v l cung gifing nhU quin the nghien cdu cQa chung to! khfing efi c l the n i o mang kieu allele dfing hpp h i l m [12], Nhu vay, so vdi nhdng nghifn cdu khic trong viing nhd Trung Quoc v i Nhat Bin thi chiing tfii cung cfi k i t q u i khfing khic b i f t nhilu.
CIc n g h i f n ciifu trong khu vdc chi quan t i m din 1 hole 2 vj trf SNP cOa gene IL1BB nhu Nhgt Bin la 106
noMINLi lUa Tap 7-Sfi 4/2012
rs8099917 va rsl 2979860 [11], Trung Quoc chl khio sit 1 dilm [12]. Trong nghifn cQu niy, chung tfii t i l n h i n h phin tich dong thdi tai 3 diem dfing thdi cung tiln h i n h so sinh tfnh da hlnhcQa gene IL28B t r i n 2 nhfim: b f n h nhan v i l m gan C va ngUdI khde manh. K i t q u i eho t h i y kieu allele trdi T/T cua SNP rs8099917 6 nhdm v i l m gan C cao han cd J nghTa so vdl nhfim ngddi khde mgnh. Trong khi dd khdng efi khic b i f t v l t i n s u i t allele cOa 2 SNP rsl2980275 v l rsl2979860 glQa 2 nhfim n g h i i n cQu. Do 100% benh n h i n viem gan C efi k i l u allele dfing hpp tQ trpl v l so iQpng bfinh n h f n cfin ft nfin chQng tfil khfing tiln hinh p h i n tfch mfii lifin quan gida tinh da hinh t h l i kieu gene IL28B vdi h i l u q u i d i l u trj. Trong nghlln cdu ciia cae tac g i i Trung Qufic cung nhu nhilu nghiin cQu khac d i u cho thay hieu q u i d i l u trt v i l m gan C cao hOn d bfinh nhan mang kieu allele dfing hpp t d trfii tgi vj trf rs8099917 [12]. Td nhdng k i t q u i dd d l p h i n n i o g i i i thich mac du kilu gene 1 v i 6 cOa virus v i f m gan C l i nhQng kieu gene pho bien tgi Vift Nam nhung h i f u q u i dieu trj v i n tUOng dfii cao so vdi cic khi vyc khic.
Nhu tren da trinh bay, gene IL28B co n h i l u diem da hinh don (SNP) nhU: rs12980275, rsB105790, rs11S81222, rs8103142, rs28416813, rs4803219. rs8099917 v l rs7248668 [3-6]. Tren b l n h nhan vifim gan C, ngudi ta d i chifng minh 3 d i l m rsl2980275, rsl2979860 v i rs8099917 la cd l i l n quan d i n tiln lUpng, hifu q u i dieu trj. Kit q u i nghien cdu cQa chung tfii cho t h i y cd mpt sy tuong dfing v l tan suat xuat hien cua the da hinh tai hai vi tri rsl 2980275 v l rsl 2979860, do dfi gpi J efi lfi khong nfin t i l n h i n h xet nghiem dfing thdi the da hinh tai 3 vi tri rs12980275. rsl2979860 v l rs8099917 ma chi d n 2 trong so do l i rs*2979860 v i rs8099917. Tuy nhifin, d l cfi k i t luan chung tdi sfi t i l n hanh n g h i i n cdu t r i n so iQong benh nhin Idn hon.
5. Kit l u i n
-Phuang p h i p danh g i i da hinh gene II28B b i n g ky thuat 'Ofing hpc allele d i e hieu real-time PCR - kinetics allele specific real-time PCR" da dupe x i y dyng t h i n h cdng.' -Gida nhfim bfinh n h i n viem gan C va nhfim ngddi khde manh, t i n suit x u i t h i f n da hinh gene II28B tai vj tri rs8099917llkhlcnhau.
- 2 t h i da hinh rsl 2980275 v l rsl 2979860 cung x u i t hien dong thdi t r i n quan t h i nghien cdu.
T i l l i f u tham k h i o
1 Hoofnagle, J,H.. Course and outcome of hepatitis C.
Hepatology, 2002, 36(5 Suppl 1): p. S21-9.
2 Manns, M.P., et al., Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet, 2001.358(9286): p, 958-65.
3 Ge, D.. et al,, Genetic variation in OSB predicts hepatitis C treatment-induced viral clearance. Nature, 2009.
461 (7262): p, 399-401.
4, Suppiah, v., et ai., IL28B is associated with response to chronic hepatitis C interferon-alpha and ribavirin therapy. Nat Genet, 2009.41 (10): p. 1100-4, 5 Tanaka, Y„ et al.. Genome-wide assodation of IL28B
with response to pegylated interi^eron-alpha and ribavirin therapy for chronic hepatitis C. Nat Genet, 2009.41(10): p. 1105-9.
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Clark, PJ., AJ. Thompson, and J.G, McHutchison, OSB genomlc-based treatment paradigms for patients with chronic hepatitis C infection: the future of personalized HCV therapies. Am J Gastroenterol, 2011.106(1): p. 38-45.
8. Lange, CM. and 5. Zeuzem. OSB single nucleotide polymorphisms in the treatment of hepatitis C. J Hepatol. 2011.55(3): p. 692-701.
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PLoS One, 2011.6(7): p. e20904.
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n . Kobayashi, M., et al.. Association of two polymorphisms of the IL28B gene with viral factors and treatment response in 1,518 patients infected with hepatitis C virus. J Gastroenterol.
12. Li, W., et al., Predicting sustained viral response to hepatitis C using a rapid and simple IL28B rs8099917 genotyping assay. Antiviral Res. 94(1): p. 54-56.
