WO2017016921A1

WO2017016921A1 - New crystalline forms of (6s)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylic acid

Info

Publication number: WO2017016921A1
Application number: PCT/EP2016/067103
Authority: WO
Inventors: Xinhui HU; Jing XIONG
Original assignee: F. Hoffmann-La Roche Ag; Hoffmann-La Roche Inc.
Priority date: 2015-07-24
Filing date: 2016-07-19
Publication date: 2017-02-02

Abstract

The present invention relates to a new crystalline form of compound (I), (6S)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3- carboxylic acid and pharmaceutical compositions comprising the crystalline forms thereof disclosed herein, which may be used for the treatment or prophylaxis of a viral disease in a patient relating to hepatitis B infection or a disease caused by hepatitis B infection.

Description

New crystalline forms of (6S)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2- dihydrobenzo[a] quinolizine-3-carboxylic acid

The present invention relates to new crystalline forms of compound (I),

(65^,)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3- carboxylic acid and pharmaceutical compositions comprising the crystalline forms thereof disclosed herein, which may be used for the treatment or prophylaxis of a viral disease in a patient relating to hepatitis B infection or a disease caused by hepatitis B infection.

BACKGROUAND OF THE INVENTION

Hepatitis B, which is caused by Hepatitis B virus infection, is recognized as a chronic viral disease of the liver which is characterized by liver disease. HBsAg quantification is a significant biomarker for prognosis and treatment response in chronic hepatitis B . However the achievement of HBsAg loss and seroconversion is rarely observed in chronically infected patients but remains the ultimate goal of therapy. Compound (I), (6S)-10-methoxy-6-isopropyl-9-(3- methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylic acid can be used as inhibitor of HBsAg production or secretion in the treatment or prophylaxis of HBV infection .

Crystalline form is a term to denote polymorphs and pseudo-polymorphs of a crystalline solid. It has fundamental influences on the physicochemical properties such as solubility, chemical stability, physical stability, photo -stability, powder particulate properties, etc. To find solid forms which can enhance the developability of compound (I) fundamentally, comprehensive studies were conducted and as a result, achieved this invention.

The present invention relates generally to solid crystalline forms of compound (I) with sufficient stability, physical stability and photo-stability, and methods to make the forms. SUMMARY OF THE INVENTION

The present invention relates to crystalline forms, and methods for the synthesis of selective production of crystalline forms of (6S)-10-methoxy-6-isopropyl-9-(3- methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylic acid or a hydrate thereof.

In one aspect, provided herein is a crystalline form of compound (I) or a hydrate thereof. In another aspect, the crystalline form of compound (I) is Form A, Form B, Form C, Form

E, Form K, Form F, Form G, Form H, Form J or a combination thereof.

In another embodiment, the crystalline form is Form A that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.54°+0.10°, 10.17°+0.10°, 15.62°+0.10°, 18.05°+0.10°, 20.46°+0.10° and 23.62°+0.10°. In a further embodiment, the crystalline form is Form A that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.54°+0.10°, 9.35°+0.10°, 10.17°+0.10°, 10.83°+0.10°, 13.63°+0.10°, 14.59°+0.10°, 15.62°+0.10°,

18.05°+0.10°, 20.46°+0.10°, 21.64°+0.10°, 22.61°+0.10°, 22.75°+0.10°, 23.62°+0.10° and 26.47°+0.10°. In a further embodiment, the crystalline form is Form A that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [%]

6.40°+0.10° 8.79 16.64°+0.10° 5.64 27.51°+0.10° 3.00

7.54°+0.10° 96.11 18.05°+0.10° 63.83 28.77°+0.10° 6.18

9.35°+0.10° 18.66 18.80°+0.10° 5.15 29.56°+0.10° 0.97

10.17°+0.10° 100.00 19.36°+0.10° 5.89 30.22°+0.10° 4.51

10.83°+0.10° 20.86 20.12°+0.10° 8.53 32.27°+0.10° 2.47 11.20°+0.10° 1.86 20.46°+0.10° 32.92 33.17°+0.10° 0.64

11.88°+0.10° 0.94 21.64°+0.10° 23.33 34.92°+0.10° 0.88

13.14°+0.10° 7.66 22.61°+0.10° 17.59 36.07°+0.10° 0.61

13.63°+0.10° 9.95 22.75°+0.10° 18.09 37.18°+0.10° 0.38

14.59°+0.10° 18.89 23.62°+0.10° 57.41 37.90°+0.10° 0.73

15.62°+0.10° 30.73 25.61°+0.10° 1.78 38.81°+0.10° 0.52

16.10°+0.10° 3.83 26.47°+0.10° 14.16

In a further embodiment, the crystalline form is Form A that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 5.

In a further embodiment, the crystalline form is Form A with a differential scanning calorimetry (DSC) thermogram comprising endothermic peak with onset temperature at 172°C +3°C.

In a further embodiment, the crystalline form is Form A with a differential scanning calorimetry (DSC) thermogram comprising endothermic peak substantially the same as shown in FIG. 6.

In a further embodiment, the crystalline form is Form A which is an anhydrous form of compound (I).

In another embodiment, the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.46 °+0.10°, 11.84 °+0.10°, 15.62 °+0.10°, 17.51 °+0.10°, 20.19 °+0.10°, 21.85 °+0.10°, 23.07 °+0.10° and 26.43°+0.10°. In a further embodiment, the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.46 °+0.10°, 11.84 °+0.10°, 11.99 °+0.10°, 14.79 °+0.10°, 15.62 °+0.10°, 17.51 °+0.10°, 20.19 °+0.10°, 20.43 °+0.10°, 21.70 °+0.10°, 21.85 °+0.10°, 23.07 °+0.10°, 24.13 °+0.10°, 26.43 °+0.10°, 26.90 °+0.10°, 29.35 °+0.10° and 30.27°+0.10°.

In a further embodiment, the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at: Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [%]

9.46 °+0.10° 60.18 21.85 °+0.10° 43.10 29.68 °+0.10° 6.00

11.84 °+0.10° 37.96 23.07 °+0.10° 90.00 30.27 °+0.10° 12.04

11.99 °+0.10° 29.47 24.13 °+0.10° 25.77 31.16 °+0.10° 4.25

13.78 °+0.10° 1.56 25.23 °+0.10° 3.20 31.85 °+0.10° 5.81

14.79 °+0.10° 25.63 25.42 °+0.10° 5.23 33.22 °+0.10° 3.81

15.62 °+0.10° 95.44 25.66 °+0.10° 4.17 33.99 °+0.10° 2.36

17.51 °+0.10° 68.02 26.43 °+0.10° 100.00 34.41 °+0.10° 2.88

18.71 °+0.10° 5.13 26.90 °+0.10° 13.37 35.68 °+0.10° 1.71

19.56 °+0.10° 4.22 27.80 °+0.10° 1.21 36.26 °+0.10° 3.07

20.19 °+0.10° 49.29 28.78 °+0.10° 4.25 37.05 °+0.10° 0.79

20.43 °+0.10° 17.80 29.19 °+0.10° 8.26 38.38 °+0.10° 3.09

21.70 °+0.10° 21.74 29.35 °+0.10° 11.54

In a further embodiment, the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 1.

In a further embodiment, the crystalline form is Form B with a differential scanning calorimetry (DSC) thermogram comprising two endothermic peaks, corresponding to dehydration of Form B and melting of Form A respectively.

In a further embodiment, the crystalline form is Form B with a differential scanning calorimetry (DSC) thermogram comprising endothermic peaks substantially the same as shown in FIG. 2.

In a further embodiment, the crystalline form is Form B which is a monohydrate form of compound (I).

In a further embodiment, the crystalline form is Form B with a X-ray crystal structure shown in FIG 4.

In another embodiment, the crystalline form is Form C that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.76°±0.10' 8.48°+0.10°, 10.34°+0.10°, 16.73°+0.10°, 17.43°+0.10° and 26.42°+0.10°. In a further embodiment, the crystalline form is Form C that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.76°+0.10°, 8.48°+0.10°, 10.34°+0.10°, 10.69°+0.10°, 14.05°+0.10°, 14.57°+0.10°, 15.21°+0.10°,

16.73°+0.10°, 17.43°+0.10°, 18.07°+0.10°, 19.39°+0.10°, 20.1°+0.10°, 21.57°+0.10°,

21.97°+0.10°, 25.84°+0.10° and 26.42°+0.10°.

In a further embodiment, the crystalline form is Form C that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

In a further embodiment, the crystalline form is Form C that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 7. In a further embodiment, the crystalline form is Form C with a differential scanning calorimetry (DSC) thermogram comprising two endothermic peaks and an exothermic peak, corresponding to melting of form C with onset temperature at 160°C ±3°C, recrystallization of Form A and melting of form A in chronological order. In a further embodiment, the crystalline form is Form C with a differential scanning calorimetry (DSC) thermogram comprising endothermic peaks and an exothermic peak substantially the same as shown in FIG. 9. In a further embodiment, the crystalline form is Form C which is an anhydrous form of compound (I).

In another embodiment, the crystalline form is Form E that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.58°+0.10°, 8.82°+0.10°, 17.00°+0.10°, 17.29°+0.10°, 17.71°+0.10° and 26.74°+0.10°.

In a further embodiment, the crystalline form is Form E that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.58°+0.10°, 8.82°+0.10°, 9.20°+0.10°, 10.41°+0.10°, 11.06°+0.10°, 12.70°+0.10°, 14.01°+0.10°, 15.07°+0.10°, 16.43°+0.10°, 17.00°+0.10°, 17.29°+0.10°, 17.71°+0.10°, 19.23°+0.10°, 21.92°+0.10°,

22.27°+0.10°, 22.70°+0.10°, 24.09°+0.10° and 26.74°+0.10°.

In a further embodiment, the crystalline form is Form E that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

In a further embodiment, the crystalline form is Form E that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 10. In a further embodiment, the crystalline form is Form E with a differential scanning calorimetry (DSC) thermogram comprising two endothermic peaks and an exothermic peak, corresponding to melting of Form E with onset temperature at 79°C ±3°C, recrystallization of Form A and melting of Form A in chronological order. In a further embodiment, the crystalline form is Form E with a differential scanning calorimetry (DSC) thermogram comprising endothermic peaks and an exothermic peak substantially the same as shown in FIG. 11.

In a further embodiment, the crystalline form is Form E which is an anhydrous form of compound (I). In another embodiment, the crystalline form is Form K that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.48 °+0.10°, 9.09 °+0.10°, 10.40 °+0.10°, 16.71 °+0.10°, 18.23 °+0.10° and 21.97 °+0.10°.

In a further embodiment, the crystalline form is Form K that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 5.19 °+0.10°, 8.48 °+0.10°, 9.09 °+0.10°, 10.40 °+0.10°, 12.98 °+0.10°, 15.51 °+0.10°, 16.71 °+0.10°, 18.23 °+0.10°, 21.40 °+0.10°, 21.97 °+0.10°, 22.37 °+0.10°, 22.59 ° +0.10°, 24.50 °+0.10° and

26.23°+0.10°.

In a further embodiment, the crystalline form is Form K that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

5.19°+0.10° 9.38 15.51°+0.10° 18.43 22.59°+0.10° 22.25

8.48°+0.10° 44.71 16.71°+0.10° 48.48 23.08°+0.10° 7.90

9.09°+0.10° 100.00 17.23°+0.10° 6.31 24.11°+0.10° 2.81

9.87°+0.10° 7.02 17.61°+0.10° 7.91 24.50°+0.10° 13.64

10.40°+0.10° 35.19 18.23°+0.10° 65.45 26.23°+0.10° 11.83

10.99°+0.10° 5.54 19.73°+0.10° 5.01 27.56°+0.10° 4.85

12.98°+0.10° 12.51 20.13°+0.10° 3.95 28.81°+0.10° 0.80

13.64°+0.10° 2.88 21.40°+0.10° 22.05 30.87°+0.10° 1.89

14.20°+0.10° 3.97 21.97°+0.10° 24.82 37.08°+0.10° 0.78

14.78°+0.10° 7.57 22.37°+0.10° 13.14 In a further embodiment, the crystalline form is Form K that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 16.

In another embodiment, the crystalline form is Form F that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.51°+0.10°, 15.61°+0.10°, 17.48°+0.10° , 21.91°+0.10°, 23.13°+0.10° and 26.44°+0.10°.

In a further embodiment, the crystalline form is Form F that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.51°+0.10°, 11.83 °+0.10°, 12.02 °+0.10°, 15.61°+0.10°, 17.48°+0.10°, 20.10°+0.10°, 21.91°+0.10°,

23.13°+0.10° and 26.44°+0.10°.

In a further embodiment, the crystalline form is Form F that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

In a further embodiment, the crystalline form is Form F that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 12.

In another embodiment, the crystalline form is Form G that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.56°±0.10°, 15.57°+0.10°, 17.38°+0.10°, 23.11°+0.10°, 23.18°+0.10° and 26.47°+0.10°.

In a further embodiment, the crystalline form is Form G that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.56°+0.10°, 11.76°+0.10°, 12.09°+0.10°, 14.91°+0.10°, 15.57°+0.10°, 17.38°+0.10°, 19.97°+0.10°,

21.89°+0.10°, 23.11°+0.10°, 23.18°+0.10°, 24.34°+0.10° and 26.47°+0.10°. In a further embodiment, the crystalline form is Form G that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

In a further embodiment, the crystalline form is Form G that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 13. In another embodiment, the crystalline form is Form H that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.48°+0.10°, 9.74°+0.10°, 12.03°+0.10°, 15.42°+0.10°, 23.09°+0.10° and 26.32°+0.10°.

In a further embodiment, the crystalline form is Form H that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.48 °+0.10°, 9.74 °+0.10°, 11.66 °+0.10°, 12.03 °+0.10°, 14.23 °+0.10°, 15.01 °+0.10°, 15.42 °+0.10°, 16.97 °+0.10°, 17.30 °+0.10°, 19.79 °+0.10°, 21.92 °+0.10°, 22.00 °+0.10°, 23.09 °+0.10°, 23.60 °+0.10°, 24.30 °+0.10° and 26.32 °+0.10°.

In a further embodiment, the crystalline form is Form H that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

7.09°+0.10° 0.76 17.3°+0.10° 41.40 25.78°+0.10° 6.24

7.53°+0.10° 7.49 18.53°+0.10° 2.91 26.32°+0.10° 53.49

9.48°+0.10° 86.82 19.79°+0.10° 26.11 27.60°+0.10° 4.91

9.74°+0.10° 100.00 20.85°+0.10° 5.41 28.55°+0.10° 3.19

11.66°+0.10° 31.38 21.92°+0.10° 45.66 29.26°+0.10° 5.16 12.03°+0.10° 57.60 22.00°+0.10° 48.93 29.93°+0.10° 4.36

13.66°+0.10° 2.49 22.55°+0.10° 7.06 32.25°+0.10° 3.48

14.23°+0.10° 13.19 23.09°+0.10° 92.19 33.37°+0.10° 0.61

15.01°+0.10° 13.83 23.60°+0.10° 14.15 34.64°+0.10° 1.91

15.42°+0.10° 83.70 24.30°+0.10° 17.23 35.68°+0.10° 1.49

16.97°+0.10° 22.94 25.20°+0.10° 2.74 38.35°+0.10° 1.47

In a further embodiment, the crystalline form is Form H that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 14.

In another embodiment, the crystalline form is Form J that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.61°+0.10°, 9.63°+0.10°, 9.81°+0.10°, 10.21°+0.10°, 23.17°+0.10° and 23.61°+0.10°.

In a further embodiment, the crystalline form is Form J that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.61°+0.10° 9.63 °+0.10°, 9.81 °+0.10°, 10.21 °+0.10°, 10.82 °+0.10°, 11.67 °+0.10°, 12.12 °+0.10°, 15.45 °+0.10°, 15.63 °+0.10°, 17.36 °+0.10°, 18.05 °+0.10°, 20.50 °+0.10°, 21.63 °+0.10°, 22.01 °+0.10°, 22.60 °+0.10°, 23.17 °+0.10°, 23.61 °+0.10° and 26.33 °+0.10°.

In a further embodiment, the crystalline form is Form J that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at:

Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [%] Pos. [°2Th.] Rel. Int. [¼]

6.43 °+0.10° 6.14 15.45 °+0.10° 33.46 22.6 °+0.10° 19.39

7.61 °+0.10° 100.00 15.63 °+0.10° 24.21 23.17 °+0.10° 41.64

9.63 °+0.10° 41.68 16.69 °+0.10° 5.10 23.61 °+0.10° 46.04

9.81 °+0.10° 38.53 16.99 °+0.10° 7.95 24.37 °+0.10° 7.31

10.21 °+0.10° 56.99 17.36 °+0.10° 11.16 25.63 °+0.10° 3.25

10.82 °+0.10° 10.35 18.05 °+0.10° 17.65 26.33 °+0.10° 24.88

11.67 °+0.10° 11.09 18.82 °+0.10° 1.82 27.53 °+0.10° 3.11

12.12 °+0.10° 19.19 19.36 °+0.10° 4.03 28.77 °+0.10° 5.72

13.13 °+0.10° 4.51 19.76 °+0.10° 9.50 30.27 °+0.10° 3.81

13.67 °+0.10° 6.04 20.5 °+0.10° 20.00 34.75 °+0.10° 1.20

14.23 °+0.10° 4.15 21.63 °+0.10° 14.12

14.59 °+0.10° 9.31 22.01 °+0.10° 23.36 In a further embodiment, the crystalline form is Form J that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 15.

In another aspect, provided herein is a method of preparing the crystalline form disclosed herein. In another embodiment is a method of preparing the crystalline polymorph Form B which comprises a step of recrystallization of (65^,)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2- oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylic acid in a solvent comprising water.

The recrystallization comprises the steps of:

a) Dissolution of compound (I) in a mixture solvent of water and an organic solvent to form a clear solution; and

b) Introducing precipitation by evaporation slowly at room temperature to form Form B.

The suitable organic solvent is selected from IPA, ethanol, methanol, acetone and acetonitrile, particularly the organic solvent is IPA.

In another embodiment is a method of preparing the crystalline polymorph Form A which comprises a step of dehydration of Form B.

The dehydration is performed by storing Form B at a temperature between 60 °C and 120°C for a period between 1 day and 4 days, particularly at a temperature between 75 °C and 85 °C for a period between 2.5 days and 3.5 days, more particularly at 80 °C for a period between 3 days.

In another embodiment is a method of preparing the crystalline polymorph Form C which comprises a step of shaking of slurry of Form A in an organic solvent overnight.

The suitable organic solvent is selected from ethanol or ethyl acetate, particularly the organic solvent is ethanol.

The suitable time is from 6 hours to 40 hours, particularly the time is 24 hours.

In another embodiment is a method of preparing the crystalline polymorph Form E which comprises the steps of: a) Melting Compound (I) above melting point of Form A but below the decomposition temperature in a variable temperature chamber; and

b) Slowly cooling the above sample down to room temperature to form Form E.

The suitable high temperature is between 172 °C and 303.6 °C, particularly the temperature isl80 °C.

In step b), the suitable cooling rate is 5 to 20 °C/ min, particularly the cooling rate is 10 °C/ min.

In another aspect, provided herein is a pharmaceutical composition comprising the crystalline form disclosed herein; and a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle or a combination thereof.

In another aspect, provided herein is the use of the crystalline form disclosed herein or the pharmaceutical composition for the manufacture of a medicament for the treatment or prophylaxis of a viral disease in a patient.

In another aspect, the viral disease disclosed herein is hepatitis B infection or a disease caused by hepatitis B infection.

In another aspect, provided herein is a method for the treatment or prophylaxis of hepatitis B infection or a disease caused by hepatitis B infection, which method comprises administering a therapeutically effective amount of the crystalline form or the pharmaceutical composition disclosed herein.

The term "XRPD" denotes the analytical method of X-Ray Powder Diffraction. Bragg's law describes the diffraction of crystalline material with the equation "2d * sin(theta) = n * lambda", wherein "d" denotes perpendicular distance (in Angstroms) between pairs of adjacent planes in a crystal ("d-spacing"), "theta" denotes the Bragg angle, "lambda" denotes the wavelength and "n" is an integer. When Bragg's law is fulfilled, the reflected beams are in phase and interfere constructively so that Bragg diffraction peaks are observed in the X-ray diffraction pattern. At angles of incidence other than the Bragg angle, reflected beams are out of phase and destructive interference or cancellation occurs. The relative peak height of X-ray powder diffraction (XRPD) pattern depends on many factors related to sample preparation and geometric shapes of the instrument, however peak position is insensitive to experimental details. In some embodiments, the crystalline form disclosed herein characterized by XRPD pattern with some listed peak positions essentially can also be characterized by XRPD pattern provided in the appended drawings of the present invention. According to the state of the instrument for the experiment, the error margin in 2-theta of the characteristic peaks is ±0.10°.

The term "DSC" denotes the analytical method of Differential Scanning Calorimetry. The term "onset" when used in connection with DSC thermograms denotes the intersection point of the baseline before transition and the interflection tangent.

Similarly, the relative peak height of differential scanning calorimetry (DSC) depends on many factors related to sample preparation and geometric shapes of the instrument, however peak position is insensitive to experimental details. In some embodiments, the crystalline form disclosed herein characterized by DSC thermogram with some listed peak positions essentially can also be characterized by DSC thermogram provided in the appended drawings of the present invention. According to the state of the instrument for the experiment, the error margin in the melting peaks is ±3°C.

The term "TGA" denotes the analytical method of thermo gravimetric analysis wherein changes in weight in relation to change in temperature are determined. Similarly, the weight loss and corresponding temperature of thermal gravimetric analysis

(TGA) depends on many factors related to sample preparation and geometric shapes of the instrument. In some embodiments, the crystalline form disclosed herein characterized by TGA thermogram with some listed weight losses essentially can also be characterized by TGA thermogram provided in the appended drawings of the present invention. Whenever a number disclosed in the invention with "±" stands for a numerical range with a lower limit and an upper limit, any number falling within the rage between lower limit and upper limit is disclosed. As used herein, the term "relative intensity" refers to the intensity of a peak with respect to the intensity of the strongest peak in the XRPD pattern which is regarded as 100%.

As used herein, the term "combination" refers to a crystalline form containing one or more other crystalline forms.

As used herein, the term "peak" refers to a feature, in a spectrum and/or data presented in a graph, that one skilled in the art would recognize as not attributable to background noise.

The terms "pharmaceutical composition" is used interchangeably and denote a mixture or solution comprising a therapeutically effective amount of an active pharmaceutical ingredient together with pharmaceutically acceptable excipients to be administered to a mammal, e.g., a human in need thereof.

The term "therapeutically effective amount" denotes an amount of a compound or molecule of the present invention that, when administered to a subject, (i) treats or prevents the particular disease or condition, (ii) attenuates, ameliorates or eliminates one or more symptoms of the particular disease or condition or (iii) prevents or delays the onset of one or more symptoms of the particular disease or condition described herein. The therapeutically effective amount will vary depending on the compound, the disease state being treated, the severity of the disease treated, the age and relative health of the subject, the route and form of administration, the judgement of the attending medical or veterinary practitioner, and other factors.

Pharmaceutical composition or medicament contain the crystalline forms of the compound (I) of the invention and a therapeutically inert carrier, diluent or excipient, as well as methods of using the crystalline forms of compound (I) of the invention to prepare such compositions and medicaments.

The crystalline forms of compound (I) of the invention may be administered by any suitable means, including oral, topical (including buccal and sublingual), rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal, intrapulmonary, intradermal, intrathecal and epidural and intranasal, and, if desired for local treatment, intralesional administration. Parenteral infusions include intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. The crystalline forms of compound (I) of the present invention may be administered in any convenient administrative form, e.g., tablets, powders, capsules, solutions, dispersions, suspensions, syrups, sprays, suppositories, gels, emulsions, patches, etc. Such compositions may contain components conventional in pharmaceutical preparations, e.g., diluents, carriers, pH modifiers, sweeteners, bulking agents, and further active agents.

An embodiment, therefore, includes a pharmaceutical composition comprising a crystalline form of compound (I). In a further embodiment includes a pharmaceutical composition comprising compound (I), or a stereoisomer or pharmaceutically acceptable salt thereof, together with a pharmaceutically acceptable carrier or excipient.

ABBREVIATIONS

IPA Isopropyl alcohol

Pos. Position

Rel. Int. Relative Intensity

RH Relative humidity

Cts counts

d- spacing calculated lattice d- spacing

TRS Total relative substance

LUX Lumen/m²

DESCRIPTION OF THE FIGURES

FIG. 1 X-ray powder diffraction pattern of Form B

FIG. 2 DSC thermogram of Form B

FIG. 3 TGA curve of Form B

FIG. 4 X-ray crystal structure of Form B

FIG. 5 X-ray powder diffraction pattern of Form A

FIG. 6 DSC thermogram of Form A

FIG. 7 TGA curve of Form A

FIG. 8 X-ray powder diffraction pattern of Form C

FIG. 9 DSC thermogram of Form C

FIG. 10 X-ray powder diffraction pattern of Form E FIG. 11 DSC thermogram of Form E

FIG. 12 X-ray powder diffraction pattern of Form F

FIG. 13 X-ray powder diffraction pattern of Form G

FIG. 14 X-ray powder diffraction pattern of Form H

FIG. 15 X-ray powder diffraction pattern of Form J

FIG. 16 X-ray powder diffraction pattern of Form K

EXAMPLES

The invention will be more fully understood by reference to the following examples. They should not, however, be construed as limiting the scope of the invention.

Example 1: preparation of (6S)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo- 6,7-dihydrobenzo[a] quinolizine-3-carboxylic acid

Step 1: Preparation of 4-bromo-l-methoxy-2-(3-methoxypropoxy)benzene

A 250 mL round-bottomed flask was charged with 5-bromo-2-methoxy-phenol (15.5 g, 76.4 mmol), l-bromo-3-methoxy-propane (12.9 g, 84 mmol), K₂CO₃ (22 g, 2153 mmol) and DMF (50 mL). The resultant mixture was stirred at 50 °C for 3 hours, and then ethyl acetate and water was added. The organic phase was separated, and then dried over anhydrous Na₂S0₄ and then concentrated to give 4-bromo-l-methoxy-2-(3-methoxypropoxy)benzene (23 g).

Step 2: Preparation of l-[4-methoxy-3-(3-methoxypropoxy)phenyl]-3-methyl-butan-2- one

A mixture of 4-bromo-l-methoxy-2-(3-methoxypropoxy)benzene (20 g, 73 mmol), 3- methylbutan-2-one (19 g, 219 mmol), Pd₂(dba)₃ (1 g, 1.2 mmol), Xantphos (1.3 g, 2.4 mmol) and t-BuONa (23 g, 241 mol) in 500 mL of THF was stirred at 70 °C overnight. Then ethyl acetate and water were added. The separated organic phase was washed with brine, and then dried over anhydrous Na₂S0₄ and then concentrated. The residue was purified by column chromatography to give l-[4-methoxy-3-(3-methoxypropoxy)phenyl]-3-methyl-butan-2-one (19 g)

Step 3: Preparation of l-[4-methoxy-3-(3-methoxypropoxy)phenyl]-3-methyl-butan-2- amine

l-[4-Methoxy-3-(3-methoxypropoxy)phenyl]-3-methyl-butan-2-one (19 g, 73 mmol) was dissolved in MeOH (150 mL). Then NH₄OAc (84 g, 1.1 mol) and NaBH₃CN (9.2 g, 146 mmol) were added. The mixture was stirred at room temperature overnight. 20% NaOH aqueous solution (100 mL) was added to the mixture. The reaction mixture was stirred for 20 minutes. The mixture was extracted with ethyl acetate, and the organic layer was dried over anhydrous Na₂S0₄ and then concentrated to afford l-[4-methoxy-3-(3-methoxypropoxy)phenyl]-3-methyl- butan-2-amine (8 g) which was used in the next step without further purification.

Step 4: Preparation of N-[l-[[4-methoxy-3-(3-methoxypropoxy)phenyl]methyl]-2- methyl-propyl]formamide

A mixture of l-(4-methoxy-3-ethoxy-phenyl)butan-2-amine (73 mmol) and formic acid (40 mL) in dioxane (150 mL) was refluxed for 16 hours and then concentrated under reduced pressure to afford crude N-[l-[[4-methoxy-3-(3-methoxypropoxy)phenyl]methyl]-2-methyl- propyl]formamide which was used in the next step without purification.

Step 5: Preparation of 7-methoxy-3-isopropyl-6-(3-methoxypropoxy)-3,4- dihydroisoquinoline

To a solution ofN-[l-[[4-methoxy-3-(3-methoxypropoxy)phenyl]methyl]-2-methyl- propyl]formamide (64.7 mmol) in acetonitrile (150 mL) was added POCI₃ (10.1 g, 64.7 mmol) dropwise at 0-5 °C. The resultant mixture was refluxed for 4 hours and then concentrated. Ethyl acetate was added, followed by addition of ammonia water to adjust the pH of the aqueous solution to around 11. The aqueous layer was extracted with ethyl acetate. The organic layers were combined and then concentrated. The residue was purified by column chromatography to give 7-methoxy-3-isopropyl-6-(3-methoxypropoxy)-3,4-dihydroisoquinoline (16 g).

Step 6: Preparation of ethyl 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo- l,6,7,llb-tetrahydrobenzo[a]quinolizine-3-carboxylate

A mixture of 7-methoxy-3-isopropyl-6-(3-methoxypropoxy)-3,4-dihydroisoquinoline (16 g, 55 mmol) and ethyl 2-(ethoxymethylene)-3-oxo-butanoate (30 g, 165 mmol) in EtOH (150 mL) was stirred at 100 °C overnight. The mixture was concentrated to give crude ethyl 10-methoxy-6- isopropyl-9-(3-methoxypropoxy)-2-oxo- 1,6,7, 1 lb-tetrahydrobenzo[a]quinolizine-3-carboxylate as dark brown oil which was used in the next step without purification.

Step 7: Preparation of ethyl 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo- 6,7-dihydrobenzo[a]quinolizine-3-carboxylate

A mixture of crude ethyl 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-l,6,7,l lb- tetrahydrobenzo[a]quinolizine-3-carboxylate (55 mmol) and p-chloranil (13.4 g, 55 mmol) in DME (100 mL) was refluxed for 2 hours. After being cooled to room temperature, the mixture was concentrated under vacuum to give crude ethyl 10-methoxy-6-isopropyl-9-(3- methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylate as brown oil.

Step 8: Preparation of 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a] quinolizine-3-carboxylic acid

To a solution of crude ethyl 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a]quinolizine-3-carboxylate from Step 7 in EtOH (100 mL) was added 10% NaOH aqueous solution dropwise at room temperature. The resultant mixture was stirred for 2 hours, and then acidified to pH 1-2 with 2M hydrochloric acid. The mixture was extracted with DCM, and the combined organic layers were washed with brine, and then dried over anhydrous Na₂S0₄ and then concentrated. The residue was purified by column chromatography and recrystallization from EtOH/ethyl ether to afford 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a]quinolizine-3-carboxylic acid (8.7 g) as a white solid. 1H NMR (400 MHz, DMSO-J₆): δ 8.76 (s, 1H), 7.52 (s, 1H), 7.45 (s, 1H), 7.09 (s, 1H), 4.43 (dd, 1H), 4.08 (m, 2H), 3.88 (s, 3H), 3.48 (t, 2H), 3.13-3.17 (m, 2H), 2.01 (m, 2H), 1.61-1.66 (m, 1H), 0.88 (d, 3H), 0.71 (d, 3H). MS obsd. (ESI⁺) [(M+H)⁺]: 402.

Step 9: Preparation of (6S)-10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a] quinolizine-3-carboxylic acid

Separation of the racemic 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a]quinolizine-3-carboxylic acid (2.0 g) by chiral HPLC afforded (e^- lO-methoxy- 6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7-dihydrobenzo[a]quinolizine-3-carboxylic acid (840 mg) and (6R)- 10-methoxy-6-isopropyl-9-(3-methoxypropoxy)-2-oxo-6,7- dihydrobenzo[a]quinolizine-3-carboxylic acid (1.0 g).1H NMR (400 MHz, OMSO-d₆): δ 8.76 (s, 1H), 7.52 (s, 1H), 7.45 (s, 1H), 7.09 (s, 1H), 4.43 (dd, 1H), 4.08 (m, 2H), 3.88 (s, 3H), 3.48 (t, 2H), 3.13-3.17 (m, 2H), 2.01 (m, 2H), 1.61 - 1.66 (m, 1H), 0.88 (d, 3H), 0.71 (d, 3H). MS obsd. (ESI⁺) [(M+H)⁺] : 402. [a]_D ²⁰ = +71.19° (0.059%, CH₃CN). Example 2: Preparation of Form B of compound (I)

10 mg of compound (I) was weighed and transferred to a 1-mL vial. 100 mL mixture of IPA and water (3: 1, v/v) was added to the previous vial to form a clear solution. The clear solution was evaporated slowly to induce precipitation at room temperature. The resulting solid was isolated and analysed for XRPD analysis, DSC analysis and TGA analysis. The XRPD pattern of Form B of compound (I) is shown in FIG. 1. Major peaks and their related intensities in the XRPD pattern are shown in Table 1 below.

Experimental conditions:

The XRPD spectrum of the solid was obtained by PANalytical EMPYREAN X-ray powder diffractometer (Cu-Κα radiation) at 40KV and 40mA. The sample was recorded between 4 and 40° in 2Θ with a step size of 0.026° and a scanning rate of 3.35 min.

The DSC curve was acquired by TA Q2000. The sample was heated from 30°C to 300°C at a rate of 10°C /min.

The TGA analysis was operated on TA Q5000. The sample was heated from 30°C to 300°C at a rate of 10°C /min. Table 1. X-Ray Powder Diffraction peaks of Form B of compound (I)

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%]

9.46 8286.04 9.34483 60.18

11.84 5226.40 7.47407 37.96

11.99 4058.04 7.38083 29.47

13.78 214.46 6.42747 1.56

14.79 3529.35 5.99082 25.63

15.62 13139.84 5.67342 95.44

17.51 9365.81 5.06561 68.02

18.71 706.16 4.74313 5.13

19.56 581.52 4.53909 4.22

20.19 6787.06 4.39904 49.29

20.43 2450.43 4.34813 17.80

21.70 2993.64 4.09481 21.74

21.85 5934.43 4.06718 43.10

23.07 12391.47 3.85501 90.00

24.13 3548.27 3.68764 25.77

25.23 440.61 3.52977 3.20

25.42 719.56 3.50364 5.23

25.66 573.52 3.47187 4.17

26.43 13768.28 3.37211 100.00

26.90 1840.29 3.31408 13.37

27.80 166.91 3.20883 1.21

28.78 585.62 3.10166 4.25

29.19 1137.48 3.05964 8.26

29.35 1588.78 3.04270 11.54

29.68 826.05 3.01029 6.00

30.27 1658.12 2.95267 12.04 31.16 585.02 2.87082 4.25

31.85 799.26 2.80951 5.81

33.22 524.65 2.69672 3.81

33.99 325.49 2.63796 2.36

34.41 395.94 2.60619 2.88

35.68 235.22 2.51624 1.71

36.26 422.19 2.47731 3.07

37.05 108.69 2.42627 0.79

38.38 425.09 2.34557 3.09

DSC and TGA results are shown in FIG. 2 and FIG. 3 respectively, indicating the dehydration temperature of Form B of compound (I) is around 56°C.

FIG. 4 shows the X-ray structure of Form B of compound (I), indicating that Form B is a monohydrate of compound (I). The single crystal X-ray intensity data were collected at 100K using a Gemini R Ultra diffractometer (Cu-Κα radiation, λ= 1.54184 A). The crystal data and structure refinement is shown in Table 2.

Table 2. Crystal data and structure refinement of Form B of compound (I).

Empirical formula C22 H29 N O7

Formula weight 419.46

Temperature 100(2) K

Wavelength 1.54184 A

Crystal system, space group Monoclinic, P2(l)

a = 9.30001(13) A

Alpha= 90 deg.

b = 11.72942(18) A

Unit cell dimensions

Beta = 95.127(1 l)deg.

c = 9.39100(11) A

Gamma= 90 deg. Volume 1020.31(2) A³

Z, Calculated density 2, 1.365 mg/mm³

Absorption coefficient 0.842 mm^"1

F(000) 448

Crystal size 0.60 x 0.50 x 0.40 mm³

Theta range for data collection 4.73 to 67.07 deg.

-10<=h<=10

Limiting indices -13<=k<=13

-11<=1<=11

Reflections collected/unique 19188 / 3342 [R(int) = 0.0276]

Completeness to theta = 67.07 97.9 %

Refinement method Full-matrix least-squares on F

Data/ restraints/ parameters 3342 / 1 / 287

Goodness-of-fit on F2 1.031

Rl = 0.0242

Final R indices [I>2sigma(I)]

wR2 = 0.0655

Absolute structure parameter 0.04(11)

Largest diff. peak and hole 0.194 and -0.179 e.A^"3

Example 3: Preparation of Form A of compound (I)

10 mg of form B was weighed into a 1-mL vial. The sample was stored at 80°C for 3 days. The resulting solid was collected for XRPD analysis, DSC analysis and TGA analysis. The XRPD pattern of Form A of compound (I) is shown in FIG. 5. Major peaks and their related intensities in the XRPD pattern are shown in Table 3 below.

The XRPD spectrum of the solid was obtained by PANalytical EMPYREAN X-ray powder diffractometer (Cu-Κα radiation) at 40KV and 40mA. The sample was recorded between 4 and 40° in 2Θ with a step size of 0.026° and a scanning rate of 3.35 min. The DSC curve was acquired by TA Q2000. The sample was heated from 30°C to 200°C at a rate of 10°C /min.

The TGA analysis was operated on TA Q5000. The sample was heated from 30°C to 350°C at a rate of 10°C /min. Table 3. X-Ray Powder Diffraction peaks of Form A of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [ ]

6.40 1170.89 13.81608 8.79

7.54 12801.41 11.72615 96.11

9.35 2485.17 9.45939 18.66

10.17 13319.06 8.69731 100.00

10.83 2778.14 8.16850 20.86

11.20 247.39 7.89824 1.86

11.88 124.88 7.45144 0.94

13.14 1020.12 6.73854 7.66

13.63 1325.78 6.49565 9.95

14.59 2516.08 6.07279 18.89

15.62 4092.38 5.67364 30.73

16.10 509.47 5.50611 3.83

16.64 750.78 5.32719 5.64

18.05 8501.67 4.91414 63.83

18.80 686.39 4.71915 5.15

19.36 784.95 4.58466 5.89

20.12 1135.96 4.41391 8.53

20.46 4384.69 4.33990 32.92

21.64 3107.06 4.10691 23.33

22.61 2342.23 3.93330 17.59

22.75 2408.78 3.90800 18.09 23.62 7646.92 3.76624 57.41

25.61 236.62 3.47867 1.78

26.47 1885.93 3.36729 14.16

27.51 399.05 3.24282 3.00

28.77 822.72 3.10310 6.18

29.56 129.24 3.02197 0.97

30.22 600.21 2.95707 4.51

32.27 329.55 2.77375 2.47

33.17 85.11 2.70116 0.64

34.92 116.64 2.56971 0.88

36.07 80.90 2.49041 0.61

37.18 50.07 2.41852 0.38

37.90 96.91 2.37396 0.73

38.81 69.50 2.32028 0.52

DSC and TGA results are shown in FIG. 6 and FIG. 7 respectively, indicating the melting point of Form A of compound (I) is around 172°C.

Example 4: Preparation of Form C of compound (I)

10 mg of form A was weighed into a 1-mL vial. O. lmL of ethanol was added to the previous vial. The sample was shaken for 24 hours. The resulting solid was collected for XRPD analysis and DSC analysis. The XRPD pattern of Form C of compound (I) is shown in FIG. 8. Major peaks and their related intensities in the XRPD pattern are shown in Table 4 below.

The DSC curve was acquired by TA Q2000. The sample was heated from 30°C to 200°C at a rate of 10°C /min. Table 4. X-Ray Powder Diffraction peaks of Form C of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%]

5.51 1046.43 16.03364 14.72

6.97 859.57 12.67883 12.10

7.25 764.70 12.20028 10.76

7.76 7015.00 11.38928 98.71

8.31 1954.95 10.64496 27.51

8.48 6806.16 10.43070 95.77

8.92 1702.20 9.91008 23.95

9.24 1413.63 9.57344 19.89

9.45 1340.68 9.35793 18.86

10.34 7106.76 8.55874 100.00

10.69 3268.43 8.27565 45.99

11.99 296.19 7.37854 4.17

12.46 1180.33 7.10524 16.61

13.42 1238.32 6.59947 17.42

13.85 1679.09 6.39344 23.63

14.05 3045.95 6.30277 42.86

14.57 3744.96 6.08055 52.70

15.21 2253.67 5.82608 31.71

16.29 1584.74 5.44235 22.30

16.49 1835.31 5.37480 25.82

16.73 4154.20 5.29779 58.45

17.43 5871.81 5.08812 82.62

18.07 3640.12 4.90793 51.22

18.53 1002.75 4.78827 14.11

19.01 183.53 4.66905 2.58

19.39 3742.09 4.57720 52.66 19.73 965.04 4.49949 13.58

20.10 2650.59 4.41762 37.30

20.80 1681.85 4.27009 23.67

21.12 1317.95 4.20645 18.55

21.31 1873.41 4.17056 26.36

21.57 3224.05 4.12017 45.37

21.97 3139.24 4.04571 44.17

22.13 1436.34 4.01761 20.21

22.33 525.84 3.98085 7.40

22.81 1381.06 3.89818 19.43

23.31 230.72 3.81658 3.25

23.59 275.24 3.77174 3.87

23.83 635.10 3.73443 8.94

24.19 706.14 3.67997 9.94

24.58 1123.66 3.62169 15.81

25.12 791.35 3.54533 11.14

25.32 2102.80 3.51742 29.59

25.84 2538.41 3.44833 35.72

26.16 261.83 3.40700 3.68

26.42 4335.38 3.37382 61.00

26.65 1669.26 3.34481 23.49

26.88 256.52 3.31641 3.61

27.14 782.36 3.28547 11.01

27.43 1976.06 3.25201 27.81

DSC result is shown in FIG. 9, indicating the melting point of Form C of compound (I) is around 160°C.

Example 5: Preparation of Form E of compound (I) 10 mg of compound (I) was weighed into a variable temperature chamber. The sample was heated to 180°C, and cooled at 10°C/min to 25 °C. The resulting solid was collected for XRPD analysis and DSC analysis. The XRPD pattern of Form E of compound (I) is shown in FIG. 10. Major peaks and their related intensities in the XRPD pattern are shown in Table 5 below. The XRPD spectrum of the solid was obtained by PANalytical EMPYREAN X-ray powder diffractometer (Cu-Κα radiation) at 40KV and 40mA. The sample was recorded between 4 and 40° in 2Θ with a step size of 0.026° and a scanning rate of 3.35 min

Table 5. X-Ray Powder Diffraction peaks of Form E of compound (I)

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [ ]

5.18 515.69 17.06870 5.68

6.96 250.40 12.70183 2.76

8.58 2287.21 10.30605 25.21

8.82 8624.91 10.02551 95.05

9.20 1105.65 9.61434 12.18

9.45 549.12 9.36376 6.05

10.41 1903.14 8.49955 20.97

11.06 1204.30 8.00349 13.27

11.48 147.20 7.71049 1.62

12.02 265.80 7.36559 2.93

12.41 367.50 7.13435 4.05

12.70 1003.62 6.96995 11.06

13.19 334.82 6.71482 3.69

14.01 1001.75 6.31978 11.04

14.56 685.83 6.08299 7.56

15.07 1558.28 5.87754 17.17

15.65 116.63 5.66250 1.29

16.43 1886.09 5.39477 20.79 17.00 3031.09 5.21475 33.40

17.29 3403.80 5.12893 37.51

17.71 9074.20 5.00685 100.00

18.38 638.29 4.82780 7.03

19.23 962.83 4.61553 10.61

20.48 367.71 4.33671 4.05

21.35 654.99 4.16111 7.22

21.92 932.43 4.05556 10.28

22.27 1107.59 3.99152 12.21

22.70 1417.02 3.91746 15.62

23.58 485.83 3.77273 5.35

24.09 936.93 3.69495 10.33

25.09 351.05 3.54914 3.87

25.62 415.65 3.47702 4.58

26.01 467.82 3.42559 5.16

26.74 4703.68 3.33342 51.84

27.98 115.08 3.18884 1.27

28.76 139.77 3.10397 1.54

29.50 128.15 3.02779 1.41

30.32 542.29 2.94779 5.98

33.59 46.69 2.66792 0.51

34.58 104.34 2.59378 1.15

35.16 90.84 2.55279 1.00

35.97 202.56 2.49671 2.23

38.83 54.06 2.31926 0.60

DSC result is shown in FIG. 11, indicating the melting point of Form E of compound (I) is around 79.4°C.

Example 6: Preparation of Form K of compound (I) 10 mg of compound (I) was weighed into a variable temperature chamber. The sample was heated to 180°C, and then cooled at 10°C/min to 100 °C. The resulting solid was analyzed by XRPD. Cooling of Form K at 10°C/min to 25 °C afforded Form E. The XRPD pattern of Form K of compound (I) is shown in FIG. 16. Major peaks and their related intensities in the XRPD pattern are shown in Table 6 below.

Table 6. X-Ray Powder Diffraction peaks of Form K of compound (I)

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [ ]

5.19 291.19 17.02686 9.38

8.48 1388.64 10.42315 44.71

9.09 3105.96 9.72904 100.00

9.87 217.97 8.96602 7.02

10.40 1092.87 8.50477 35.19

10.99 172.19 8.05282 5.54

12.98 388.62 6.82098 12.51

13.64 89.51 6.49146 2.88

14.20 123.30 6.23830 3.97

14.78 235.19 5.99336 7.57

15.51 572.37 5.71266 18.43

16.71 1505.92 5.30511 48.48

17.23 195.92 5.14732 6.31

17.61 245.64 5.03671 7.91

18.23 2032.90 4.86707 65.45

19.73 155.53 4.50028 5.01

20.13 122.58 4.41130 3.95

21.40 684.78 4.15306 22.05 21.97 770.99 4.04643 24.82

22.37 408.18 3.97489 13.14

22.59 690.96 3.93665 22.25

23.08 245.32 3.85345 7.90

24.11 87.22 3.69094 2.81

24.50 423.71 3.63404 13.64

26.23 367.30 3.39778 11.83

27.56 150.68 3.23715 4.85

28.81 24.93 3.09890 0.80

30.87 58.57 2.89675 1.89

37.08 24.13 2.42464 0.78

Example 7: Preparation of Form F of compound (I)

30 mg of compound (I) Form B was weighed into a variable temperature chamber. The sample was placed in RH 10% at 20°C for four hours. The formed solid was analyzed by XRPD. 10 mg of Form F was weighed into a 1-mL vial. Form F changed to Form A after being stored at 80°C for 3 days. The XRPD pattern of Form F of compound (I) is shown in FIG. 12. Major peaks and their related intensities in the XRPD pattern are shown in Table 7 below.

The XRPD spectrum of the solid was obtained by PANalytical EMPYREAN X-ray powder diffractometer (Cu-Κα radiation) at 40KV and 40mA. The sample was recorded between 4 and 40° in 2Θ with a step size of 0.013° and a scanning rate of 5.08 min.

Table 7. X-Ray Powder Diffraction peaks of From F of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%]

6.82 30.76 12.96406 0.46

7.56 265.07 11.69206 4.00

9.51 6634.90 9.30241 100.00

11.83 1319.24 7.48155 19.88

12.02 1376.36 7.36267 20.74 13.78 67.47 6.42494 1.02

14.58 473.99 6.07389 7.14

14.84 660.43 5.96939 9.95

15.61 3439.30 5.67625 51.84

17.48 1828.14 5.07265 27.55

18.69 106.50 4.74711 1.61

19.55 123.94 4.54117 1.87

20.10 1070.76 4.41747 16.14

20.47 575.44 4.33788 8.67

21.91 2286.29 4.05736 34.46

23.13 3454.71 3.84588 52.07

24.19 626.26 3.67878 9.44

25.51 206.60 3.49119 3.11

26.44 2984.09 3.37140 44.98

26.94 355.51 3.30950 5.36

28.79 136.22 3.10077 2.05

Example 8: Preparation of From G of compound (I)

30 mg of compound (I) Form B was weighed into a variable temperature chamber. The sample was placed in RH 10% at 25°C for four hours. The resulting solid was analyzed by XRPD. 10 mg of Form G was weighed into a 1-mL vial. Form G changed to Form A after being stored at 80°C for 3 days. The XRPD pattern of Form G of compound (I) is shown in FIG. 13. Major peaks and their related intensities in the XRPD pattern are shown in Table 8 below.

Table 8. X-Ray Powder Diffraction peaks of From G of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%] 6.84 46.83 12.92205 1.33

7.62 298.00 11.60590 8.45

9.56 3526.25 9.24771 100.00

11.76 1048.36 7.52470 29.73

12.09 1251.22 7.31823 35.48

13.75 81.70 6.43919 2.32

14.40 320.61 6.14990 9.09

14.91 452.78 5.94227 12.84

15.57 2688.19 5.69315 76.23

17.38 1637.08 5.10227 46.43

18.06 53.52 4.91194 1.52

18.66 141.89 4.75532 4.02

19.57 190.28 4.53658 5.40

19.97 811.05 4.44607 23.00

20.47 260.38 4.33906 7.38

21.89 1277.62 4.06009 36.23

23.11 2529.19 3.84612 71.72

23.18 2728.39 3.84364 77.37

24.34 419.17 3.65434 11.89

25.72 180.64 3.46085 5.12

26.47 2158.61 3.36495 61.22

27.04 121.72 3.29471 3.45

28.72 121.11 3.10536 3.43

29.27 262.86 3.04841 7.45

30.16 161.12 2.96060 4.57

31.91 77.22 2.80267 2.19

34.46 63.38 2.60030 1.80

35.86 30.31 2.50200 0.86 36.69 31.66 2.44763 0.90

38.47 53.34 2.33810 1.51

Example 9: Preparation of From H of compound (I)

30 mg of compound (I) Form B was weighed into a variable temperature chamber. The sample was placed in RH 10% at 30°C for four hours. The resulting solid was analyzed by XRPD. 10 mg of Form H was weighed into a 1-mL vial. Form F changed to Form A after being stored at 80°C for 3 days. The XRPD pattern of From H of compound (I) is shown in FIG. 14. Major peaks and their related intensities in the XRPD pattern are shown in Table 9 below.

Table 9. X-Ray Powder Diffraction peaks of From H of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%]

7.09 28.27 12.46403 0.76

7.53 278.95 11.73850 7.49

9.48 3234.58 9.33204 86.82

9.74 3725.81 9.07966 100.00

11.66 1169.11 7.58886 31.38

12.03 2146.03 7.35911 57.60

13.66 92.70 6.48301 2.49

14.23 491.62 6.22526 13.19

15.01 515.26 5.90235 13.83

15.42 3118.60 5.74536 83.70

16.97 854.59 5.22394 22.94

17.30 1542.56 5.12601 41.40

18.53 108.58 4.78786 2.91

19.79 972.73 4.48631 26.11 20.85 201.54 4.26084 5.41

21.92 1701.27 4.05201 45.66

22.00 1823.02 4.04055 48.93

22.55 262.93 3.94335 7.06

23.09 3434.68 3.85135 92.19

23.60 527.16 3.76978 14.15

24.30 641.93 3.66295 17.23

25.20 102.10 3.53352 2.74

25.78 232.54 3.45544 6.24

26.32 1992.79 3.38668 53.49

27.60 182.80 3.23155 4.91

28.55 118.90 3.12695 3.19

29.26 192.41 3.05247 5.16

29.93 162.48 2.98585 4.36

32.25 129.74 2.77554 3.48

33.37 22.76 2.68496 0.61

34.64 71.27 2.58992 1.91

35.68 55.48 2.51662 1.49

38.35 54.72 2.34734 1.47

Example 10: Preparation of Form J of compound (I)

30 mg of compound (I) Form B was weighed into a variable temperature chamber. The sample was placed in RH 10% at 35°C for four hours. The resulting solid was analyzed by XRPD. 10 mg of form J was weighed into a 1-mL vial. Form J changed to Form A after being stored at 80°C for 3 days. The XRPD pattern of From J of compound (I) is shown in FIG. 15. Major peaks and their related intensities in the XRPD pattern are shown in Table 10 below. The XRPD spectra of the solid were obtained by PANalytical EMPYREAN X-ray powder diffractometer (Cu-Κα radiation) at 40KV and 40mA. The sample was recorded between 4 and 40° in 2Θ with a step size of 0.013° and a scanning rate of 5.08 min.

Table 10. X-Ray Powder Diffraction peaks of Form J of compound (I).

Pos. [°2Th.] Height [cts] d- spacing [A] Rel. Int. [%]

6.43 154.40 13.74499 6.14

7.61 2512.82 11.62425 100.00

9.63 1047.33 9.18596 41.68

9.81 968.27 9.01482 38.53

10.21 1432.04 8.66457 56.99

10.82 260.18 8.17452 10.35

11.67 278.74 7.58124 11.09

12.12 482.30 7.30243 19.19

13.13 113.40 6.74477 4.51

13.67 151.68 6.47830 6.04

14.23 104.21 6.22389 4.15

14.59 233.92 6.06952 9.31

15.45 840.82 5.73358 33.46

15.63 608.47 5.66796 24.21

16.69 128.18 5.31279 5.10

16.99 199.83 5.21827 7.95

17.36 280.51 5.10876 11.16

18.05 443.46 4.91417 17.65

18.82 45.78 4.71476 1.82

19.36 101.38 4.58494 4.03

19.76 238.67 4.49411 9.50

20.50 502.60 4.33241 20.00

21.63 354.72 4.10857 14.12 22.01 587.04 4.03810 23.36

22.60 487.30 3.93430 19.39

23.17 1046.43 3.83941 41.64

23.61 1156.94 3.76773 46.04

24.37 183.79 3.65218 7.31

25.63 81.54 3.47543 3.25

26.33 625.25 3.38493 24.88

27.53 78.12 3.23973 3.11

28.77 143.75 3.10292 5.72

30.27 95.69 2.95232 3.81

34.75 30.22 2.58189 1.20

Example 11: Solid state stability study of Form A, Form B and Form C

6 mg of Form A, Form B or Form C was weighed into a 20 mL glass vial and stored at the conditions and the time indicated in Table 11 respectively. After incubation, chemical stability of the samples was tested by HPLC, and physical stability was tested by XRPD.

Table 11. Solid state stability of Form A, Form B and Form C

Degradation

Testing ; Condition Time XRPD

(%)

80°C 24 hours 0.04 Unchanged

1 month 0.12 Form B

40°C RH75 2 months 0.12 Form B

3 months 0.13 Form B

Form A 1 month 0.12 Partially changed to Form B

25°C RH60 2 months 0.13 Partially changed to Form B

3 months 0.13 Almost Form B

1 month 0.07 Partially changed to Form B

2-8°C

2 months 0.09 Partially changed to Form B 3 months 0.09 Partially changed to Form B

80°C 24 hours 0.09 Changed to Form A

1 month -0.02 Unchanged

40°C RH75 2 months 0.01 Unchanged

3 months 0.01 Unchanged

1 month 0.07 Unchanged

Form B

25°C RH60 2 months 0.05 Unchanged

3 months 0.06 Unchanged

1 month 0.03 Unchanged

2-8°C 2 months 0 Unchanged

3 months 0.01 Unchanged

80°C 24 hours 0.09 Unchanged

1 month 0.08 Partially changed to Form B

40°C RH75 2 months 0.08 Partially changed to Form B

3 months 0.09 Partially changed to Form B

1 month 0.08 Unchanged

Form C

25°C RH60 2 months 0.08 Unchanged

3 months 0.08 Unchanged

1 month 0.08 Unchanged

2-8°C 2 months 0.08 Unchanged

3 months 0.08 Unchanged

Example 12: Photo-stability of Form A, Form B and From C

6 mg of Form A, Form B or Form C was weighed into a 20 mL glass vial and stored at the conditions indicated in Table 12 respectively. After incubation, chemical stability of the samples was tested by HPLC.

Table 12. Photo-stability of Form A, Form B and Form C Conditions Time point Purity TRS Increase

Initial Form A — 99.61 —

Light (Form A solid state) 1.2 million Lux- hours 99.56 0.05

Initial Form B — 99.46% —

Light (Form B solid state) 1.2 million Lux- hours 98.24% 1.41%

Initial Form C — 99.65 —

Light (Form C solid state) 1.2 million Lux- hours 99.36 0.29

Example 13: HPLC method for chemical purity test

HPLC condition is disclosed here in Table 13.

Table 13. HPLC method for chemical purity test

Instrument Aglent 1260 HPLC system

Column Waters XSELECT CSH18(4.6 x 150mm x 3.5μπι)

Column Temperature 30°C

Mobile Phase A 0.5% FA in water

Mobile Phase B 0.5% FA in ACN

Time A% B%

0.00 75 25

20.00 50 50

Gradient Program 23.00 10 90

28.00 10 90

28.01 75 25

33.00 25 25

Diluent ACN: water=l: l

Detector UV 250nm

Flow rate 0.7mL/min

Injection volume 5 μL·

Nominal concentration 0.3 mg/mL Example 14: The capacity to inhibit HBsAg materials and methods HBV cell line

HepG2.2.15 cells (Acs et al. Proc Natl Acad Sci U S A, 84, (1987), 4641-4), a

constitutively HBV-expressing cell line were cultured in DMEM+Glutamax-I medium

(Invitrogen, Carlsbad, CA, USA), supplemented with 10% fetal bovine serum (Invitrogen) and G418 (Invitrogen) at a final concentration of 200 mg/L and maintained in 5% C0₂ at 37°C.

HBsAg Assay

HepG2.2.15 cells were seeded in duplicate into white, 96-well plates at 1.5 x 10⁴ cells/well. The cells were treated with a three-fold serial dilution series of the compounds in DMSO. The final DMSO concentration in all wells was 1% and DMSO was used as no drug control.

The HBsAg chemiluminescence immunoassay (CLIA) kit (Autobio Diagnostics Co., Zhengzhou, China, Catalog number: CL0310-2) was used to measure the levels of secreted HBV antigens semi-quantitatively. For the detection 50 μίΛνεΙΙ culture supernatant was used and HBsAg was quantified using HBsAg chemiluminescence immunoassay (CLIA) kit (Autobio Diagnostics Co., Zhengzhou, China, Catalog number: CL0310-2), 50 μΐ_^ of the supernatant was transferred to the CLIA assay plate and 50 μΐ_^ of enzyme conjugate reagent was added into each well. The plates were sealed and gently agitated for 1 hour at room temperature. The

supernatant-enzyme-mixture was discarded and wells were washed 6 times with 300 μΐ_^ of PBS. The residual liquid was removed by plating the CLIA plate right side down on absorbent tissue paper. 25 μΐ_^ of substrates A and B were added to each well. Luminance was measured using a luminometer (Mithras LB 940 Multimode Microplate Reader) after 10 minutes incubation. Dose- response curves were generated and the IC₅₀ value was extrapolated by using the E-WorkBook Suite (ID Business Solutions Ltd., Guildford, UK). The IC₅₀ was defined as the compound concentration (or conditioned media log dilution) at which HBsAg secretion was reduced by 50% compared to the no drug control.

Compound (I) was tested for their capacity to inhibit HBsAg as described herein and found to have lCso of 0.002 μΜ.

Claims

1. A crystalline form of compound (I):

2. A crystalline form according to claim 1, wherein the form is Form A, From B, Form C, Form E, Form F, Form G, Form H, Form J, From K or a combination thereof.

3. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form A that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.54°+0.10°, 10.17°+0.10°, 15.62°+0.10°, 18.05°+0.10°, 20.46°+0.10° and 23.62°+0.10°.

4. A crystalline form according to claim 3, wherein the crystalline form is Form A that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.54°+0.10°, 9.35°+0.10°, 10.17°+0.10°, 10.83°+0.10°, 13.63°+0.10°, 14.59°+0.10°, 15.62°+0.10°, 18.05°+0.10°, 20.46°+0.10°, 21.64°+0.10°, 22.61°+0.10°,

22.75°+0.10°, 23.62°+0.10° and 26.47°+0.10°. 5. A crystalline form according to claim 3 or 4, wherein the crystalline form is Form A that exhibits an X-ray powder diffraction (XRPD) pattern shown in FIG.

5.

6. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.46 °+0.10°, 11.84 °+0.10°, 15.62 °+0.10°, 17.51 °+0.10°, 20.19 °+0.10°, 21.85 °+0.10°, 23.07 °+0.10° and 26.43°+0.10°.

7. A crystalline form according to claim 6, wherein the crystalline form is Form B that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.46 °+0.10°, 11.84 °+0.10°, 11.99 °+0.10°, 14.79 °+0.10°, 15.62 °+0.10°, 17.51 °+0.10°, 20.19 °+0.10°, 20.43 °+0.10°, 21.70 °+0.10°, 21.85 °+0.10°, 23.07 °+0.10°, 24.13 °+0.10°, 26.43 °+0.10°, 26.90 °+0.10°, 29.35 °+0.10° and 30.27°+0.10°.

8. A crystalline form according to claim 6 or 7, wherein the crystalline form B that exhibits a X-ray powder diffraction (XRPD) pattern shown in FIG. 1.

9. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form C that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.76°+0.10°, 8.48°+0.10°, 10.34°+0.10°, 16.73°+0.10°, 17.43°+0.10° and 26.42°+0.10°. 10. A crystalline form according to claim 9, wherein the crystalline form is Form C that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.76°+0.10°, 8.48°+0.10°, 10.34°+0.10°, 10.69°+0.10°, 14.05°+0.10°,

14.57°+0.10°, 15.21°+0.10°, 16.73°+0.10°, 17.43°+0.10°, 18.07°+0.10°, 19.39°+0.10°,

20.1°+0.10°, 21.57°+0.10°, 21.97°+0.10°, 25.84°+0.10° and 26.42°+0.

10°.

11. A crystalline form according to claim 9 or 10, wherein the crystalline form is Form C that exhibits an X-ray powder diffraction (XRPD) pattern shown in FIG. 7.

12. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form E that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.58°+0.10°, 8.82°+0.10°, 17.00°+0.10°, 17.29°+0.10°, 17.71°+0.10° and 26.74°+0.10°.

13. A crystalline form according to claim 12, wherein the crystalline form is Form E that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.58°+0.10°, 8.82°+0.10°, 9.20°+0.10°, 10.41°+0.10°, 11.06°+0.10°,

12.70°+0.10°, 14.01°+0.10°, 15.07°+0.10°, 16.43°+0.10°, 17.00°+0.10°, 17.29°+0.10°,

17.71°+0.10°, 19.23°+0.10°, 21.92°+0.10°, 22.27°+0.10°, 22.70°+0.10°, 24.09°+0.10° and 26.74°+0.10°.

14. A crystalline form according to claim 12 or 13, wherein the crystalline form is Form E that exhibits an X-ray powder diffraction (XRPD) pattern shown in FIG. 10.

15. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form K that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 8.48 °+0.10°, 9.09 °+0.10°, 10.40 °+0.10°, 16.71 °+0.10°, 18.23 °+0.10° and 21.97 °+0.10°.

16. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form F that exhibits an X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.51°+0.10°, 15.61°+0.10°, 17.48°+0.10° , 21.91°+0.10°, 23.13°+0.10° and 26.44°+0.10°.

17. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form G that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.56°+0.10°, 15.57°+0.10°, 17.38°+0.10°, 23.11°+0.10°, 23.18°+0.10° and 26.47°+0.10°.

18. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form H that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 9.48°+0.10°, 9.74°+0.10°, 12.03°+0.10°, 15.42°+0.10°, 23.09°+0.10° and 26.32°+0.10°.

19. A crystalline form according to claim 1 or 2, wherein the crystalline form is Form J that exhibits a X-ray powder diffraction (XRPD) pattern with characteristic peaks expressed in degrees 2-theta at 7.61°+0.10°, 9.63°+0.10°, 9.81°+0.10°, 10.21°+0.10°, 23.17°+0.10° and 23.61°+0.10°.

20. A pharmaceutical composition comprising the crystalline form of anyone of the claims 1 to 19 and a pharmaceutically acceptable carrier, excipient, diluent, adjuvant, vehicle or a combination thereof.

21. The use of the amorphous or crystalline form of any one of claims 1 to 19 or the pharmaceutical composition of claim 20 for the manufacture of a medicament for the treatment or prophylaxis of a viral disease in a patient.

22. The use according to claim 21, wherein the viral disease is hepatitis B infection or a disease caused by hepatitis B infection.

23. A method for the treatment or prophylaxis of hepatitis B infection or a disease caused by hepatitis B infection, which method comprises administering an therapeutically effective amount of the amorphous or crystalline form as defined in any one of claims 1 to 19 or the

pharmaceutical composition of claim 20.

24. The invention as hereinbefore described.