Naturally occurring hepatitis B virus reverse transcriptase mutations related to potential antiviral drug resistance and liver disease progression

Table 1 Distribution of preexisting RT mutation in 42 potential NAr regions in treatment naïve patients

Mutation	RT mutation type	Change in HBsAg3	Drug resistance	Genotype	Location	Ref.
Primary1	I169T		ETV	B, C	China	[20,53]
	A181T/V	sW172 stop	LMV, LdT, ADV, TNF	A, B, C, D	Canada, Italy, China, United States	[24,25,30,39,45,51,53]
	T184A/C/F/G/I/L/M/S		ETV, LMV	A, B, C, D	Canada, China, South Korea	[24,27,33]
	A194T	no change in HBsAg	ADV, TNF	B, C, D	Indonesia, Italy	[38,51]
	S202C/G/I		ETV, LMV	A, B, C, D	Canada, China, Spain, Italy	[24,27,30,57,59]
	M204I/V/S	V-sI195M, I- sW196S/L/Stop	LMV, LdT, ADV, TNF, ETV	A, B, C, D	Canada, Italy, South Korea, China, Indonesia, United States, Turkey	[24,25,27,30,33-36,38, 39,47,52,53]
	N236T		ADV, TNF	B, C	China	[27,30,47]
	M250I/L/V		ETV	A, B, C, D	Canada, Italy, China, United States	[24,25,27,45]
Secondary1	L80I/V	no change in HBsAg	LMV	A, B, C, D	Canada, Italy, South Korea, Indonesia, China	[24,25,33,38,47,53]
	V173L	sE164D	LMV	A, B, C, D	China, Canada, Italy	[20,24,25,30,39,47]
	L180M	no change in HBsAg	LMV, ETV, LdT	A, B, C, D	Canada, South Korea, China, Italy, Indonesia	[24,25,27,30,33,38, 39,47,53]
Putative2	S53N		LMV	B, C	China, South Korea	[21,33,58]
	T54N	sP46T	ADV	A, B, C, D	Italy	[83]
	L82M		LMV	C	South Korea	[33]
	V84M, S85A		ADV	B, C	South Korea, China	[33,53,127]
	I91L	no change in HBsAg	LMV	B, C, D	China, Indonesia, India	[21,38,54,58]
	Y126C/R/H	C-sT118A	ADV	A, B, C, D	South Korea, China, India	[33,54,58]
	T128I, T128N	N-sG145R, I-sP120S	LMV	B, C, D	Indonesia, China, India	[38,53,54,58]
	N139D/E/Q			A	India	[54]
	W153Q/R/E	Q- sP120T/ sG145R, E-sD144E/G145R	LMV	B, C, D	South Korea, Indonesia, China, India	[33,38,54,58]
	F166L	sF158Y	LMV
	V191I	sW182 stop	LMV, ADV	B, C, D	South Korea, China, Italy	[33,39,51,58]
	A200V	sL192F	LMV	B, C	South Korea, China	[27,33,42]
	V207I	sW199 stop, sM198I	LMV	A, B, C, D	Germany, China, Italy	[27,32,39,83]
	S213T		LMV, ETV	A, B, C, D	China, India	[39,40,42,58]
	V214A	T-sS204R	ADV	B, C, D	South Korea, Italy, China,	[30,33,53,83]
	Q215E/H/P/S		LMV, ADV	A, B, C, D	China, Italy, India, Turkey	[30,39,51,53-55]
	L217R, E218D, F221Y	D-sS210I/T	ADV	A, B, C, D	South Korea, China	[21,33,42,58]
	L229G/V/W		LMV	B, C	South Korea, China	[33,42,58]
	L229F	F-sC221L, V-sF220L	LMV	B, C, D	China, India	[54,58,128]
	I233V		ADV	A, C, D	Indonesia, Italy, India, China, South Korea, Germany	[25,38,40,58,78,79]
	P237H, N238D/S/T, Y245H	N/A	ADV	B, C	South Korea, China	[21,33,39,58]
	S/C256G		LMV, ETV	B, C	China, Indonesia	[21,38,58]
Pretreatment2	T38A, T38K	K-sQ30K		B, C	South Korea, China	[33,58]
	Y124H/D/N			B, C,	China, South Korea	[21,33,58]
	D134E/N	sI126S/N		B, C, D	China, South Korea, Indonesia	[21,33,38,58]
	N139K/H	K-sT131N, T-sT131P, H-sG139N		A, B, C	South Korea, India, Indonesia, China	[33,38,54,58]
	I224V	No change in HBsAg		B, C,	China, South Korea	[21,33,39,58]
	R242A

¹Well known NA resistance mutations (primary and secondary) with phenotypic data;

²Putative and pretreatment mutations relevant to NA resistance but not experimentally confirmed;

³Changes in HBsAg, reported in Sheldon et al[17], Liu et al[21], Locarnini et al[72], and Yang et al[77]. Rt139 is shared in both Categories 3 (N139D/E/Q) and 4 (N139K/H). Overall, 42 positions in the RT region were studied. ADV: Adefovir dipivoxil; ETV: Entecavir; Ldt: Telbivudine; LMV: Lamivudine; TNF: Tenofovir.

Table 2 Genotype-dependent amino acid polymorphic sites and reverse transcriptase mutations in treatment-naïve patients

RT position	Drug resistance	Mutations in RT region of four genotypes1				Polymorphism	Ref.
		A	B	C	D
38		T (4.4)4			T (14.0)4	A/T/T/A	[83]
53	LMV		D/T (1.8)4			I/N/S/N	[21]
54	ADV	N (2.2)2				T/T/T/H	[83]
84				I (0.5)2		V	[127]
				I (1.5)2			[21]
85				T (0.6)2		S	[127]
91	LMV		I (20.0)4	L (23.5)2	I (16.7)4	I/L/I/L	[83]
					I (14.3)2		[44]
103		I (100)3			I (1.67)3	V	[40]
122		H (47.0)2			H (6.66)2	F	[40]
					L/V/I(25.0)2		[44]
124		H (2.2)3	H (20.0)3	H (11.8)3		N/N/Y/H	[83]
			H (3.6)3	H (6.6)3			[21]
			D (5.5)3
126		H (6.7)4			R (23.7)4	Y/H/H/H	[83]
					R (17.9)4		[44]
					Y (1.4)4 Q (0.5)4		[83]
128	LMV	N (2.2)2			N (1.4)2 I (1.4)2	T	[83]
			I (1.9)2				[38]
129				L (60.0)2	L (21.4)2	M	[44]
			L (1.9)2	L (26.2)2			[38]
		L (100.0)2		L (9.0)2	L (3.3)2		[40]
134				N (40.5)2		D/N/D/D	[38,54]
				E (23.1)3	E (5.0)3		[21,54]
			I/S (1.8)3	E (5.8)3			[21]
139	LMV		K (3.7)2		K (11.9)2	Q/N/N/N	[38]
					K (2.3)3		[83]
				K (1.5)3			[21]
145			L (3.7)2	L (40.5)2		M	[83]
153	LMV	Q (100)2				W/R/R/R	[40]
191	LMV			V (8.3)2		V/I/I/V	[39]
		I (4.6)2		F (7.7)2			[54]
200	LMV	V (2.2)2				A	[83]
207	LMV		M (6.0)2 L (2.3)2			V	[26]
				I (5.9)2			[83]
				I/L/G (2.1)2			[39]
214	LMV/ADV			A (5.9)2	A (2.3)2	V	[83]
		A (0.5)2 I (0.5)2		I (0.6)2	A (0.8)2 E (0.7)2		[127]
215	ADV			E (7.7)2	H (5.0)2 S (5.0)2	Q	[54]
			H (0.9)2		H (3.0)2 S (4.3)2		[127]
					P (2.8)2 S (4.2)2		[83]
217		L (6.7)4			R (0.9)2	R/L/L/L	[83]
221	ADV			F (40.5)2		Y/Y//F/F	[38]
					Y (5.1)2		[83]
226			H/T (33.3)2	H/T (2.4)2		N	[38]
237					T (6.4)2	P	[127]
238	LMV, ETV	H(1.0)2 W(1.0)2	Q (3.9)2	T (8.7)2	H (2.3)2	N/H/N/N	[127]
			Q (1.82)2	S (2.19)2 T (0.73)2			[21]
				T (3.87)2			[39]
		D (2.2)2 T (2.2)2			D (1.4)2		[83]
245					H (1.4)2	Y	[83]
248	ADV		H (7.4)3	H (4.8)3		N	[38]
256	LMV			G (40.0)2	G (10.7)2	S/S/S/C	[44]
			G (20.0)2		G (3.7)2		[83]
			G (5.45)2				[21]
					G (3.7)2		[26]

A total of 29 reported genotype-dependent amino acid polymorphic sites in the RT region in treatment-naïve patients are shown. The first column contains the RT positions and the second column details the relationship between mutations and drug resistance. Column three to six indicate the prevalence of each mutation as percentages, according genotype. Consensus amino acids are presented in column seven.

¹Incidence (%) of mutations in the RT region;

²Putative mutation;

³Pretreatment mutation;

⁴Novel mutation. ADV: Adefovir dipivoxil; ETV: Entecavir; Ldt: Telbivudine; LMV: Lamivudine; TNF: Tenofovir.

Table 3 Positive relationships between HBeAg negative serostatus and preexisting reverse transcriptase mutation frequency in the treatment- naïve patients

HBeAg-positive		HBeAg-negative		HBV genotype (%)	Location	Ref.
Mutations1	HBV-DNA2	Mutations1	HBV-DNA2
3/14 (21.4)	7.8	11/14 (78.6)	5.7	B, C, E	California	[26]
6/24 (25.0)	5.5	18/24 (75.0)	3.9	B, C, B-C	China	[27]
0/4 (0.0)	7.2	4/4 (100.0)	4.7	A, B, C, D, F	California	[45]
3/6 (50.0)	8.0	3/6 (50.0)	3.2	D	Turkey	[36]
8/12 (67.0)	7.9	4/12 (33.0)	6.9	NA	Taiwan, China	[35]
27/43 (62.8)	5.7	16/43 (37.2)	4.7	B, C	China	[46]
8/13 (61.5)	6.3	5/13 (38.5)	5.4	B, C	China	[47]
0/5 (0.0)	NA	5/5 (100.0)	NA	NA	Japan	[34]
0/4 (0.0)	NA	4/4 (100.0)	NA	NA	Japan	[88]

¹Number of patients with RT mutation (%);

²HBV-DNA level (log₁₀ IU/mL).

Table 4 Variation in the prevalence of preexisting reverse transcriptase mutations according to mutation detection methods, genotype, and geographic distribution

Prevalence	Location	No. of cases	Genotype	HBV DNA loads (log10 IU/mL)	RT mutations prevalence	Mutation detecting methods	Ref.
HBV DNA RT mutations ≥ 5%	Italy	255	A, C, D	5.0	5.0% mutations overall	INNO-Lipa HBV DR v.3	[25]
	China	269	B, C, B-C	4.9	8.9% mutations overall	INNO-Lipa HBV DR v.3	[27]
	Canada	209	A, B, C, D	7.0	12% M204I/V, 10% L180M, 9% L80V/I, 3%V173L	INNO-Lipa HBV DR v.3	[24]
	Turkey	71	NA	NA	18.3% YMDD mutations	INNO-Lipa HBV DR v.1	[28]
	South Korea	131	C2	6.5	12.98% mutations overall	Direct Sequencing	[33]
	Turkey	77	D	7.3	7.8% YMDD mutations	Direct Sequencing	[36]
	China	213	B, C	6.2	6.1% mutations overall	Direct Sequencing	[47]
	China	104	B, C, B-C	4.5	26.9% YMDD mutations	Direct Sequencing	[93]
	Japan	18	NA	NA	27.8% YMDD mutations	Direct Sequencing	[34]
	Iran	325	D	NA	15.69% mutations overall	Direct Sequencing	[50]
	Taiwan, China	28	NA	7.5	57% YMDD mutations	Direct Sequencing	[35]
	China	357	B, C	6.3	16.8% mutations overall	Direct Sequencing	[39]
	Meta-analysis (China)	8156	B, C, D	NA	8.00% mutations overall	Record screening	[73]
	Japan	14	B, C	4.9	35.7% YMDD mutations	Ultra-deep sequencing	[94]
HBV DNA RT mutations < 5%	Iran	147	D	NA	None	Direct sequencing	[56]
	China	328	B, C	6.9	3.6% mutations overall	Direct sequencing	[53]
	Japan	20	NA	NA	None	Direct sequencing	[48]
	California	472	A, B, C, D, F	5.3	< 1% mutations overall	Direct sequencing	[45]
	Italy	100	NA	NA	None	Direct sequencing	[49]
	Italy	140	D	4.0	3.5% mutations overall	Direct sequencing	[51]
	Germany	96	A, D	NA	None	Direct sequencing	[32]
	Brazil	189	A, C, D, F	3.2	overall 6.0% in Northeast/ 0% in North	Direct sequencing	[41]
	California	198	B, C	4.2	1% mutations in polymerase	INNO-Lipa HBV DR v.3	[26]

Table 5 Relationship of preexisting reverse transcriptase mutations with disease severity

Type of mutation in RT	Chang in HBsAg	Genotype	Location	Disease progression	P value	Ref.
rtL80I2	NC	C	South Korea	HCC	0.036	[33]
rtD134N4	sI126S/N	B, C	China	HCC	0.007	[114]
rtN139K/T/H4	sT131N/P	C	South Korea	HCC	0.008	[33]
rtY141F5	sM307T	Ce	Taiwan	HCC	0.029	[37]
rtM204I/V1	sW196L/S/W	C	South Korea	HCC	0.021	[33]
rtF221Y3	NA	B,C,D	China	HCC, poor survival rate	0.028/0.004	[20,115]
rtI224V4	NC	C	China	HCC	0.005	[116]
rtM309K5	NA	C	China	HCC	0.007	[116]

HBV polymerase RT mutation;

¹Primary;

²Secondary;

³Putative;

⁴Pretreatment;

⁵Novel RT mutation. HCC: Hepatocellular carcinoma; NA: Not available; NC: Not changed; RT: Reverse transcriptase.

Table 6 Distribution of preexisting reverse transcriptase mutations among reverse transcriptase domains

Mutation frequency (%)1			P value5	Ref.
Domains2	A-B interdomain3	Non-A-B interdomains4
1.45	3.51	2.58	< 0.001	[38]
1.37	4.4	3.77	< 0.001	[20]
1.07	7.5	3.16	0.008	[33]
0.43	3.82	0.52	0.0014	[21]

¹Mutation frequency was calculated as the number of mutations found in a specific RT domain divided by the total number of sites in the domain;

²Domain including RT mutation sites; rt38, rt84, rt207, rt233, rt238, and rt256;

³A-B interdomain including RT mutations sites: rt53, rt191, rt213, rt218, rt229, and rt242;

⁴Non-A-B interdomains including RT mutation sites: rt124, rt126, rt128, rt134, rt139, and rt153;

⁵P-values of comparisons of mutation frequencies between A-B interdomain and other functional domains.