Evolution And Resistance Pattern Character Of Hepatitis B Virus Quasispecies During Rescue Therapy After Lamivudine Resistance

AIM:To investigate the evolution and resistance pattern character of hepatitis B virus quasispecies during rescue therapy after lamivudine resistance.METHODS:seven lamivudine-resistant patients with different rescue therapy were chosen, a series of serum samples at different time during rescue therapy were collected. A series of serum samples from fifteen patients with suboptimal viral response to adefovir dipivoxil combination therapy or monotherapy after lamivudine resistance were collected. The reverse-transcriptase (RT) region of the HBV polymerase gene was amplified, cloned and sequenced, sequence mutation combination with clinical medicine was analyzed,the evolution of hepatitis B virus quasispecies during rescue therapy in seven patients and the hepatitis B virus resistance mutation pattern of fifteen patients with adefovir dipivoxil combination therapy or monotherapy were investigated.RESULTS:The results of evolution of hepatitis B virus quasispecies in seven patients were as following, Patient l,at week 22 during LAM+ADV combination therapy,the M204I+L80I mutation became predominant,being present in 60%(6/10) of clones,at week 10 during LdT therapy,the new mutation L180M was detected in 57%(4/7) of clones,at week 7,21 during ETV therapy,the M204I+L180M+L80I mutation was detected in 100%(5/5) and 33%(4/12) of clones respectively.Patient 2,the M204V+L180M mutation became predominant,being present in 80%(8/10) of clones at the time of LAM resistance,then with LAM+ADV combination therapy,at weekll,the M204V+L180M+G173L mutation was detected in 80%(8/10) of clones, at week 89,the mutation of M204V+L180M+T184S and M204V+L180M was detected in 25%(2/8) and 75%(6/8) of clones respectively, at week 119, ETV was given for rescue therapy,meanwhile,thymosin was given to regulate immunity therapy,all clones(10/10) were wild after 12 weeks of ETV therapy.Patient 3,all clones (10/10) were M204I+L80I at the time of LAM resistance,at week 20 during LAM+ADV combination therapy,all clones (9/9) were wild,but at week 36, the mutation was M204I+L80I(11/11) again, at week 54, the mutations was M204I+L80I (90%,9/10) and M204I+L80I+N236T(10%,1/10) respectively.Patient 4,the M204I mutation became predominant,being present in 90%(9/10)of clones at the time of LAM resistance,at week 22 during LAM+ADV combination therapy,the mutation of M204I+L80I+L180M and M204I+L80I was detected in 56%(5/9) and 44%(4/9) of clones respectively.Patient 5,the mutation of M204V+L180M+G173L and M204V+L180M was detected in 55%(6/11) and 45%(5/11) of clones respectively at week 8 during LAM+ETV 0.5mg/d combination therapy,then ETV lmg/d was given for further rescue therapy,at week 16,44 all clones were M204V+L180M+G173L (10/10,7/7).Patient 6 and 7, all clones were M204V+L180M+G173L(6/6,10/10) at the time of LAM resistance. Patient 6, virus rebounded after 70 weeks of ADV monotherapy, then the mutation M204V+L180M+G173L+N236T (60%,6/10) and M204V+L180M+G173L (40%, 4/10) was detected. After that ADV+ETV combination therapy was given, the mutation M204V+L180M+G173L (90%,9/10) and muti-resistance mutation M204V+L180M+T184G+N236T (10%,1/10) was detected. Patient 7 with ADV monotherapy, at week 15,there was M204V+L180M+G173L (90%,9/10) and 10% (1/10) double resistances mutation M204V+L180M+G173L+N236T. At week 48, there was M204V+L180M+G173L+N236T (40%,4/10) and M204V+L180V+G173L (60%,6/10). The results of the hepatitis B virus resistance mutation pattern of fifteen patients with adefovir dipivoxil rescue therapy were as following, the ADV resistance mutation pattern of A181V, A181T and A181T+N236T was selected in ADV monotherapy group. The LAM resistance mutation pattern of M204V+L180M+L229V, M204V+L180M, M204I+L80I, M204V+L180M+V2071 was detected in combination therapy group.Besides,20% of clones of three serum samples were double resistance to LAM and entecavir (ETV) in combination therapy group,which were M204V+L180M+G173L+S202G (2/10), M204V+L180M+T184S (2/10) and M204I+L80I+T184I (2/10), Known resistance mutation wasn't detected in two serum samples from two groups respectively, in which all clones were I269L, all clones were P109S in the one from monotherapy group.CONCLUSIONS:In seven lamivudine-resistant patients with different rescue therapy, HBV existed resistance mutation is not easy to disappear, morever it is easy to select cross-resistance or multi-resistance mutation.It affects the response to rescue therapy, morever it results in the failure of rescue therapy. In LAM resistance patients with suboptimal viral response to ADV therapy,ADV resistance mutation pattern of A181T+N236T,A181V,A181T was easily selected during ADV monotherapy, in combination ADV therapy group, the LAM resistance mutation pattern of M204V+L180M+L229V, M204V+L180M, M204I+L80I, M204V+L180M+V207I was predominant,the ETV resistance mutation T184I/S, S202G could be selected in combination therapy group for the sustainable use of LAM.The mutation I269L,P109S may affect the response to ADV therapy in some patients.