DRUG COMBINATION CONTAINING TLR7 AGONIST

Abstract

A drug combination containing a TLR7 agonist. Specifically, a drug combination jointly using the compound of formula I acting as a TLR7 agonist and entecavir for the treatment of hepatitis B virus infection and a use thereof, the drug combination having a good anti-hepatitis B virus infection effect.

##STR00001##

Claims

1. A pharmaceutical combination, comprising a compound of formula I or a pharmaceutically acceptable salt thereof and entecavir or a pharmaceutically acceptable salt or solvate thereof, wherein the compound of formula I is shown as follows: ##STR00055## wherein, L.sub.1 is selected from —O—; L.sub.2 is selected from —CH.sub.2—, wherein the —CH.sub.2— is optionally substituted with R.sub.4; R.sub.1 is selected from the group consisting of hydrogen and C.sub.1-10 alkyl, wherein the C.sub.1-10 alkyl is optionally substituted with R.sub.5; R.sub.2 is selected from the group consisting of hydrogen, cyano, —COOH and —CONH.sub.2, wherein the —COOH and —CONH.sub.2 are optionally substituted with R.sub.6; B is selected from the group consisting of 6-10 membered aryl and 5-10 membered heteroaryl; L.sub.3 is selected from the group consisting of C.sub.0-6 alkylene and imino, wherein the C.sub.0-6 alkylene and imino are optionally substituted with R.sub.7; R.sub.3 is selected from the group consisting of hydrogen, amino, C.sub.1-10 alkyl, C.sub.3-10 cyclohydrocarbyl, 3-10 membered heterocyclohydrocarbyl, 6-10 membered aryl and 5-10 membered heteroaryl, wherein the amino, C.sub.1-10 alkyl, C.sub.3-10 cyclohydrocarbyl, 3-10 membered heterocyclohydrocarbyl, 6-10 membered aryl and 5-10 membered heteroaryl are optionally substituted with R.sub.8, or R.sub.3 and L.sub.3, together with an ortho atom on ring B, form a saturated or unsaturated 5-8 membered ring, wherein the 5-8 membered ring is optionally substituted with R.sub.9; n is 0, 1, 2, 3, 4 or 5; R.sub.4, R.sub.5, R.sub.6, R.sub.7, R.sub.8 and R.sub.9 are each independently selected from halogen, cyano, hydroxy, sulfydryl, amino, —R, —OR, ═O, —SR, —NHR and —NR.sub.2; R is independently selected from the group consisting of C.sub.1-8 alkyl, C.sub.3-8 cyclohydrocarbyl, 3-8 membered heterocyclohydrocarbyl, 6-8 membered aryl and 5-8 membered heteroaryl.

2-15. (canceled)

16. The pharmaceutical combination according to claim 1, wherein the compound of formula I or the pharmaceutically acceptable salt thereof is selected from the group consisting of: 2-butoxy-7-(3-((4-methylpiperazin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(3-(morpholinomethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 7-(3-(aminomethyl)benzyl)-2-butoxy-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(3-(pyrrolidin-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((3,3-difluoropyrrolidin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((3-fluoropyrrolidin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 1-(4-((4-amino-2-butoxy-5H-pyrrolo[3,2-d]pyrimidin-7-yl)methyl)benzyl)pyrrolidin-3-ol; 2-butoxy-7-(4-(piperidin-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(morpholinomethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((4-methylpiperazin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((dimethylamino)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((diethylamino)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((dipropylamino)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 7-(4-(azetidin-1-ylmethyl)benzyl)-2-butoxy-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((3-methoxylazetidin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(4-methyl-1,4-diazepan-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((2,6-dimethylmorpholino)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 7-(4-((1S,4S)-2-oxa-5-azabicyclo[2.2.1]heptan-5-ylmethyl)benzyl)-2-butoxy-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((4-methoxypiperidin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-((4-isopropylpiperazin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(pyrrolidin-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((6-(pyrrolidin-1-ylmethyl)pyridin-3-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(3-(2-(pyrrolidin-1-yl)ethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(1-(pyrrolidin-1-yl)ethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(1-methylpiperidin-4-yl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-(4-(1-methylpyrrolidin-2-yl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 1-(4-((4-amino-2-butoxy-5H-pyrrolo[3,2-d]pyrimidin-7-yl)methyl)phenyl)-4-methylpiperazin-2-one; 7-benzyl-2-(2-methoxyethoxy)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-(2-methoxyethoxy)-7-((6-methylpyridin-3-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 7-((5-chloropyridin-2-yl)methyl)-2-(2-methoxyethoxy)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-(2-methoxyethoxy)-7-((6-(pyrrolidin-1-ylmethyl)pyridin-3-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 1-(4-((4-amino-2-(2-methoxyethoxy)-5H-pyrrolo[3,2-d]pyrimidin-7-yl)methyl)phenyl)-4-methylpiperazin-2-one; 2-butoxy-7-((5-(pyrrolidin-1-ylmethyl)pyridin-2-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 4-amino-2-butoxy-7-((6-(pyrrolidin-1-ylmethyl)pyridin-3-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-6-carbonitrile; 4-amino-2-butoxy-7-(4-(pyrrolidin-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-6-carbonitrile; 4-amino-2-butoxy-7-(4-(morpholinomethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-6-carbonitrile; 4-amino-2-butoxy-7-(4-((4-methylpiperazin-1-yl)methyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-6-carbonitrile; 4-amino-2-butoxy-7-(4-(pyrrolidin-1-ylmethyl)benzyl)-5H-pyrrolo[3,2-d]pyrimidin-6-carboxamide; 2-butoxy-7-((1,2,3,4-tetrahydroisoquinolin-7-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((2-ethyl-1,2,3,4-tetrahydroisoquinolin-7-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((2-isopropyl-1,2,3,4-tetrahydroisoquinolin-7-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((1,2,3,4-tetrahydroisoquinolin-6-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((2-methyl-1,2,3,4-tetrahydroisoquinolin-6-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; 2-butoxy-7-((2-ethyl-1,2,3,4-tetrahydroisoquinolin-6-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine; and 2-butoxy-7-((2-(pyrrolidin-1-ylmethyl)thiazol-5-yl)methyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine, or a pharmaceutically acceptable salt thereof.

17. The pharmaceutical combination according to claim 1, wherein the compound of formula I in the pharmaceutical combination is selected from a compound of formula A: ##STR00056##

18. The pharmaceutical combination according to claim 1, wherein entecavir or the pharmaceutically acceptable salt or solvate thereof is selected from the group consisting of entecavir maleate, entecavir monomaleate, entecavir hydrate, entecavir 0.5-2 hydrate, and entecavir monohydrate.

19. The pharmaceutical combination according to claim 18, wherein entecavir or the pharmaceutically acceptable salt or solvate thereof is selected from entecavir monomaleate monohydrate.

20. The pharmaceutical combination according to claim 1, wherein an average daily dose ratio of the compound of formula I or the pharmaceutically acceptable salt thereof to entecavir or the pharmaceutically acceptable salt or solvate thereof in the pharmaceutical combination is selected from 10:1 to 1:10.

21. The pharmaceutical combination according to claim 20, wherein the average daily dose ratio is selected from the group consisting of 1:1.5 to 1:4, 1:1.6 to 1:3.8, 1:1.8 to 1:3.8, 1:1.8 to 1:3.6 and 1:2 to 1:3.5.

22. The pharmaceutical combination according to claim 1, wherein the compound of formula I or the pharmaceutically acceptable salt thereof in the pharmaceutical combination is administered thrice every day, twice every day, once every day, once every two days, once every three days, once every four days, once every five days, once every six days, once every week, once every two weeks or once every three weeks.

23. The pharmaceutical combination according to claim 1, wherein the compound of formula I or the pharmaceutically acceptable salt thereof in the pharmaceutical combination is administered at a dose of 0.1 to 10.0 mg each time.

24. The pharmaceutical combination according to claim 23, wherein the compound of formula I or the pharmaceutically acceptable salt thereof is administered at a dose of 0.2 to 5.0 mg, 0.4 to 4.0 mg, 0.5 to 3.0 mg, 0.6 to 2.6 mg, 0.8 to 2.2 mg, 0.8 to 1.8 mg, 1.0 to 2.0 mg, 1.0 to 1.8 mg, or 1.0 to 1.6 mg each time.

25. The pharmaceutical combination according to claim 1, wherein entecavir or the pharmaceutically acceptable salt or solvate thereof in the pharmaceutical combination is administered thrice every day, twice every day, once every day, once every two days, once every three days, once every four days, once every five days, once every six days, once every week, once every two weeks or once every three weeks.

26. The pharmaceutical combination according to claim 1, wherein entecavir or the pharmaceutically acceptable salt or solvate thereof in the pharmaceutical combination is administered at a dose of 0.005 mg to 5.0 mg each time.

27. The pharmaceutical combination according to claim 26, wherein entecavir or the pharmaceutically acceptable salt or solvate thereof is administered at a dose of 0.05 mg to 5.0 mg, 0.10 mg to 2.0 mg, 0.25 mg to 2.0 mg, or 0.5 mg to 1.0 mg each time.

28. The pharmaceutical combination according to claim 1, wherein the pharmaceutical combination is a fixed combination; or, the fixed combination is in the form of a solid pharmaceutical composition; or, the compound of formula I or the pharmaceutically acceptable salt thereof and entecavir or the pharmaceutically acceptable salt or solvate thereof in the fixed combination are present in the same solid pharmaceutical composition.

29. The pharmaceutical combination according to claim 28, wherein the solid pharmaceutical composition is selected from the group consisting of a tablet and a capsule.

30. The pharmaceutical combination according to claim 1, wherein the pharmaceutical combination is a non-fixed combination; or, the compound of formula I or the pharmaceutically acceptable salt thereof and entecavir or the pharmaceutically acceptable salt or solvate thereof in the non-fixed combination are each in the form of a solid pharmaceutical composition; or, the compound of formula I or the pharmaceutically acceptable salt thereof and entecavir or the pharmaceutically acceptable salt or solvate thereof in the non-fixed combination are each in the form of a solid pharmaceutical composition, and the solid pharmaceutical composition of the compound of formula I or the pharmaceutically acceptable salt thereof and the solid pharmaceutical composition of entecavir or the pharmaceutically acceptable salt or solvate thereof are present in the same sachet; or, the compound of formula I or the pharmaceutically acceptable salt thereof and entecavir or the pharmaceutically acceptable salt or solvate thereof in the non-fixed combination are each in the form of a solid pharmaceutical composition, and the solid pharmaceutical composition of the compound of formula I or the pharmaceutically acceptable salt thereof and the solid pharmaceutical composition of entecavir or the pharmaceutically acceptable salt or solvate thereof are not present in the same sachet.

31. The pharmaceutical combination according to claim 30, wherein the solid pharmaceutical composition is selected from the group consisting of a tablet and a capsule.

32. A method of treating hepatitis B virus infection, comprising administering to a subject in need thereof an effective amount of the pharmaceutical combination according to claim 1.

33. A kit for treating hepatitis B virus infection, comprising: (a) a first pharmaceutical composition comprising a compound of formula I or the pharmaceutically acceptable salt thereof as active ingredient; (b) a second pharmaceutical composition comprising entecavir or the solvate thereof as active ingredient; and optionally, (c) a package insert for combined use of the compound of formula I or the pharmaceutically acceptable salt thereof and entecavir or the pharmaceutically acceptable salt or solvate thereof, ##STR00057## wherein, L.sub.1 is selected from —O—; L.sub.2 is selected from —CH.sub.2—, wherein the —CH.sub.2— is optionally substituted with R.sub.4; R.sub.1 is selected from the group consisting of hydrogen and C.sub.1-10 alkyl, wherein the C.sub.1-10 alkyl is optionally substituted with R.sub.5; R.sub.2 is selected from the group consisting of hydrogen, cyano, —COOH and —CONH.sub.2, wherein the —COOH and —CONH.sub.2 are optionally substituted with R.sub.6; B is selected from the group consisting of 6-10 membered aryl and 5-10 membered heteroaryl; L.sub.3 is selected from the group consisting of C.sub.0-6 alkylene and imino, wherein the C.sub.0-6 alkylene and imino are optionally substituted with R.sub.7; R.sub.3 is selected from the group consisting of hydrogen, amino, C.sub.1-10 alkyl, C.sub.3-10 cyclohydrocarbyl, 3-10 membered heterocyclohydrocarbyl, 6-10 membered aryl and 5-10 membered heteroaryl, wherein the amino, C.sub.1-10 alkyl, C.sub.3-10 cyclohydrocarbyl, 3-10 membered heterocyclohydrocarbyl, 6-10 membered aryl and 5-10 membered heteroaryl are optionally substituted with R.sub.8, or R.sub.3 and L.sub.3, together with an ortho atom on ring B, form a saturated or unsaturated 5-8 membered ring, wherein the 5-8 membered ring is optionally substituted with R.sub.9; n is 0, 1, 2, 3, 4 or 5; R.sub.4, R.sub.5, R.sub.6, R.sub.7, R.sub.8 and R.sub.9 are each independently selected from halogen, cyano, hydroxy, sulfydryl, amino, —R, —OR, ═O, —SR, —NHR and —NR.sub.2; R is independently selected from the group consisting of C.sub.1-8 alkyl, C.sub.3-8 cyclohydrocarbyl, 3-8 membered heterocyclohydrocarbyl, 6-8 membered aryl and 5-8 membered heteroaryl.

Description

DRAWING OF THE SPECIFICATION

[0168] FIG. 1 illustrates effect of a TLR7 agonist on serum HBV DNA replication levels in AAV mice.

DETAILED DESCRIPTION

[0169] For the sake of clarity, the present application is further illustrated by examples, which are, however, not intended to limit the scope of the present application. All reagents are commercially available and can be used without further purification.

Example 1. AAV Mice Study

[0170] 1.1. Test Compounds

[0171] In this example, the TLR7 agonist was

##STR00053##

(2-butoxy-7-(4-(pyrrolidin-1-ylmethyl) benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine); see WO2016023511 for the preparation method.

[0172] In this example, the entecavir used is a monohydrate thereof.

[0173] 1.2. Method and Grouping

[0174] Male C57BL/6 mice (Shanghai Slac Laboratory Animal Co., Ltd.) aged 6-8 weeks were taken, and rAAV8-1.3HBV virus (FivePlus Institute of Molecular Medicine, Beijing) was injected into the C57BL/6 mice via tail veins at a dose of 1×10.sup.11 vg. Blood was collected from the orbit at week 2 and week 4 after the virus was injected. Serum was separated, and HBV DNA copy number in serum were measured to determine whether the model was successfully constructed or not. Mice were randomized according to the HBV DNA copy number into vehicle control group, TLR7 agonist group (20 mg/kg, tiw), entecavir (ETV) group (0.0032 (week 1, 1 W) & 0.001 (week 2-6, 2-6 W) mg/kg, qd), and TLR7 agonist and ETV combination group, each group containing 6 mice. The mice in each group were administered intragastrically for 6 weeks, and the administration was stopped for 3 week. Blood was collected from the orbit at weeks 1, 2, 3, 4, 5 and 6 during the administration period and weeks 1, 2 and 3 during the withdrawal period, and serum was separated. The HBV DNA copy number was determined by fluorescence quantitative PCR.

[0175] Groups:

[0176] i. Vehicle control group;

[0177] ii. Monotherapy group: entecavir;

[0178] iii. Monotherapy group: TLR7 agonist;

[0179] iv. Combination group (combo): TLR7 agonist and entecavir;

TABLE-US-00002 TABLE 1-1 Dosing regimen Administration Administration Administration Group Dosage (mg/kg) Route interval cycle i.   i.g. qd 6 w ii. 0.0032 (1W) & 0.001 (2-6W) i.g. qd 6 w iii. 20 (TLR7 agonist) i.g. tiw 6 w 20 (TLR7 agonist) i.g. tiw 6 w iv. 0.0032 (1W) & 0.001 (2-6W) i.g. qd 6 w (entecavir) Note: i.g.: intragastric; qd: once daily; tiw: once every three days; 6 w: six weeks. The administration of the combination group was consistent with that of the monotherapy groups.

[0180] 1.3. Method for Evaluating Pharmaceutical Combination

[0181] The test results were evaluated by Jin equation. The Jin equation is a modified method based on the Burgi method, and is also referred to the probability addition method:

[0182] The combination efficacy is calculated according to the following equation: q=E.sub.A+B/(E.sub.A+E.sub.B−E.sub.A×E.sub.B)

[0183] wherein E.sub.A denotes the efficacy of drug A monotherapy; E.sub.B denotes the efficacy of drug B monotherapy; E.sub.A+B denotes the efficacy of the combination of drugs A and B.

[0184] The significance of q: 0.85 to 1.15 indicates simple addition; 1.15 to 20 indicates enhancement; greater than 20 indicates significant enhancement; 0.85 to 0.55 indicates antagonism; less than 0.55 indicates significant antagonism. The q value takes the absolute value.

[0185] 1.4. Results

[0186] The results are shown in Table 1-2, Table 1-3, Table 1-4 and FIG. 1.

TABLE-US-00003 TABLE 1-2 Efficacy of different therapies on HBV DNA replication level (logic lU/mL) in mouse serum (treatment period) Group Treatment period (d) 0 7 14 21 28 35 42 i. 7.57 ± 0.17 7.31 ± 0.23 7.31 ± 0.31 6.79 ± 0.47 6.72 ± 0.53 6.95 ± 0.58 7.23 ± 0.67 ii. 7.53 ± 0.15 5.27 ± 0.18 5.66 ± 0.14 5.96 ± 0.28 5.34 ± 0.43 5.62 ± 0.36 6.44 ± 0.46 *** *** *** *** *** *** iii. 7.53 ± 0.15 6.64 ± 1.33 4.55 ± 1.41 4.13 ± 0.67 3.64 ± 0.21 3.61 ± 0.24 3.41 ± 0.33 *** *** *** *** *** iv. 7.50 ± 0.13 4.15 ± 0.76 2.48 ± 0.81 2.62 ± 0.74 1.82 ± 0.54 2.32 ± 0.16 2.42 ± 0.31 *** *** *** *** *** *** Note: comparison with the vehicle control group, where *P < 0.05, **P < 0.01, ***, P < 0.001.

TABLE-US-00004 TABLE 1-3 Efficacy of different therapies on HBV DNA replication level (log.sub.10 IU/mL) in mouse serum (withdrawal period) Withdrawal period (d) Group 49 56 63 i. 7.40 ± 0.49 7.51 ± 0.48 7.54 ± 0.34 ii. 6.97 ± 0.72 7.32 ± 0.73 7.33 ± 0.80 iii. 4.37 ± 0.48*** 4.86 ± 0.41*** 5.26 ± 0.39*** iv. 3.60 ± 0.32*** 3.96 ± 0.55*** 5.71 ± 0.68*** Note: comparison with the vehicle control group, where *P < 0.05, **P < 0.01, ***P < 0.001

TABLE-US-00005 TABLE 1-4 Evaluation of combination therapy by Jin equation Treatment period (d) 7 14 21 28 35 42 q value 2.35 33.54 3.25 23.38 20.32 3.02

[0187] Over a 42-day treatment period, HBV DNA was significantly reduced in the monotherapy groups of entecavir and TLR7 agonist (groups ii and iii); the combination group (group iv) also demonstrated significantly reduced HBV DNA, and q-value results indicated that the combination therapy exhibited superior synergic effect.

Example 2

[0188] 1. Test Compounds

[0189] In this example, the TLR7 agonist was

##STR00054##

(2-butoxy-7-(4-(pyrrolidin-1-ylmethyl) benzyl)-5H-pyrrolo[3,2-d]pyrimidin-4-amine); see WO2016023511 for the preparation method. The compound was prepared into tablets with strength of 0.2 mg or 0.5 mg.

[0190] Entecavir: commercially available entecavir tablets with strength of 0.5 mg. Commercially available entecavir tablets include, but are not limited to, entecavir dispersible tablets (Runzhong).

[0191] Placebo: a placebo for TLR7 agonist.

[0192] 2. Enrolled Subjects

[0193] The subject should meet all of the following criteria:

[0194] 1) Male patients or non-pregnant, non-lactating female patients aged 18-65 years (inclusive);

[0195] 2) Serum virology: persistent positive serum HBsAg for 6 months or more, or with evidence of chronic hepatitis B for 6 months;

[0196] 3) Previously treated patients: historical HBV DNA inhibition less than the lower limit of normal 24 weeks before enrollment, and HBV DNA inhibition defined as <69 IU/mL determined by a Roche reagent during screening; Fibroscan ≤9.0 Kpa (fasting), and ALT≤5×ULN;

[0197] 4) Previously untreated patients: HBV DNA >10.sup.5 copies/mL (or >20000 IU/mL) for HBeAg-positive chronic hepatitis B patients or HBV DNA >10.sup.4 copies/mL (or >2000 IU/mL) for HBeAg-negative patients as determined by Roche's Cobas Taqman real-time quantitative PCR test v2.0, with the lower limit of detection being 20 IU/mL; Fibroscan ≤12.4 Kpa (fasting), and 1×ULN≤ALT≤5×ULN. The previously untreated patients mean that the patients did not receive antiviral therapies against HBV or participate in related clinical trials.

[0198] 3. Dosage and Administration

[0199] TLR7 agonist: once weekly.

[0200] Entecavir: 0.5 mg/day, once daily.

[0201] The first group included 12 subjects, of which 8 received 1.2 mg of the TLR7 agonist in combination with entecavir and 4 received placebo in combination with entecavir for 24 weeks.

[0202] The second, third and fourth groups each included 36 subjects randomly subgrouped in a ratio of TLR7 agonist:placebo=5:1. The three groups respectively received 1.0 mg of TLR7 agonist combined with entecavir or placebo combined with entecavir (the second group), 1.5 mg of TLR7 agonist combined with entecavir or placebo combined with entecavir (the third group), and 1.8 mg of TLR7 agonist combined with entecavir or placebo combined with entecavir (the fourth group). The groups were treated for 24 weeks.

[0203] 4. Efficacy Endpoints

[0204] Efficacy endpoints: serum HBsAg, HBeAg, or HBV DNA, etc.

[0205] Exploratory endpoints: HBV RNA and HBcrAg, etc.

[0206] The mean, standard deviation, median, quartile, minimum and maximum values were used to describe the average content of the index parameters at each detection time point in each group and the change from baseline; the mixed-effects model for repeated measures (MMRM) was used, with group, baseline, detection time point, and the interaction term of group and detection time point as fixed effects and the subject and the intercept term as random effects, the changes from baseline of the index parameters at each detection time point in treatment group were compared with those of the placebo groups, respectively, and corrected by the Dunnett's test.

[0207] 5. Results

[0208] All of the TLR7 agonist and entecavir combination groups showed significantly inhibition on HBV DNA replication. All of the TLR7 agonist and placebo combinations achieved inhibition of HBV DNA replication. The entecavir and placebo combinations also had inhibitory effects on HBV DNA replication.