COMPOSITIONS AND METHODS FOR TREATING HEART FAILURE

Abstract

The present invention provides methods for treating chronic heart failure patients using the medication comprising neuregulin. The methods comprise first performing a companion diagnostic test of each patient before treatment; and then providing a suitable treatment to the patient according to the results of the companion diagnostic test. When the result of the test is within a favorite treatment zone, the patient is suitable for heart failure treatment by administering an effective amount of neuregulin.

Claims

1. A method of treating chronic heart failure, comprising: a) performing a companion diagnostic test of each patient before treatment; and b) providing a suitable treatment to the patient according to the results of the companion diagnostic test.

2. The method of claim 1, wherein the companion diagnostic test is the NYHA heart function classification.

3. The method of claim 1, wherein the companion diagnostic test is the test of plasma level of NT-proBNP or BNP.

4. The method of claim 1, wherein the suitable treatment comprising neuregulin treatment.

5. The method of claim 4, wherein the suitable treatment further comprising treating heart failure using one or more anti-heart failure drugs selected from a group consisting of: ACE inhibitors, ?-blockers, ARBs, diuretics, and digitalis.

6. The method of claim 2, wherein said suitable treatment is administered when the heart function is NYHA class II or III.

7. The method of claim 3, wherein said suitable treatment is administered when plasma level of NT-proBNP is ?4000 fmol/ml.

8. The method of claim 4 or 5, wherein the neuregulin is neuregulin-1.

9. The method of claim 4 or 5, wherein the neuregulin protein comprises of EGF-like domain of neuregulin-1.

10. The method of claim 4 or 5, wherein the neuregulin comprises of SEQ ID NO: 1.

11. Use of neuregulin for the preparation of medications for treating chronic heart failure patients, wherein the patients have a plasma NT-proBNP level of ?4000 fmol/ml before treatment.

12. The use of claim 11, wherein the neuregulin is neuregulin-1.

13. The use of claim 11, wherein the neuregulin comprises of EGF-like domain of neuregulin-1.

14. The use of claim 11, wherein the neuregulin comprises of SEQ ID NO: 1.

15. The use of claim 11, wherein the chronic heart failure is caused by ischemic, congenital, rheumatic, idiopathic, viral or toxic factors.

16. The use of claim 11, wherein the medication can be used together with anti-heart failure drugs.

17. The use of claim 16, wherein the anti-heart failure drugs is one or more selected from a group consisting of: ACE inhibitors, ?-blockers, ARBs, diuretics, and digitalis.

18. Use of neuregulin for the preparation of medications for treating chronic heart failure patients, wherein the patients has a class II or III heart function as classified by NYHA functional classification.

19. The use of claim 18, wherein the neuregulin is neuregulin-1.

20. The use of claim 18, wherein the neuregulin comprises of EGF-like domain of neuregulin-1.

21. The use of claim 18, wherein the neuregulin comprises of SEQ ID NO: 1.

22. The use of claim 18, wherein the chronic heart failure is caused by ischemic, congenital, rheumatic, idiopathic, viral or toxic factors.

23. The use of claim 18, wherein the medication can be used together with anti-heart failure drugs.

24. The use of claim 23, wherein the anti-heart failure drugs is one or more selected from a group consisting of: ACE inhibitors, ?-blockers, ARBs, diuretics, and digitalis.

25. A method of treating chronic heart failure comprising administering neuregulin, wherein the administration of neuregulin improves the clinical indication of the patient.

26. The method of claim 25, wherein the clinical indication is prolonged survival.

27. The method of claim 25, wherein the clinical indication is reduced re-hospitalization.

28. The method of claim 25, wherein the clinical indication is the expression level of biomarkers for diagnosis or prognosis of heart failure.

29. The method of claim 28, wherein the biomarker is NT-proBNP or BNP.

30. The method of claim 25, wherein the neuregulin is neuregulin-1.

31. The method of claim 25, wherein the neuregulin comprises of EGF-like domain of neuregulin-1.

32. The method of claim 25, wherein the neuregulin comprises of SEQ ID NO: 1.

33. The method of claim 25, wherein the neuregulin is administered to the patients for an introduction regimen.

34. The method of claim 33, wherein the introduction regimen includes an administration of neuregulin for at least consecutive 3, 5, 7 or 10 days.

35. The method of claim 33, wherein the neuregulin is administered to the patients for a maintenance regimen after the introduction regimen.

36. The method of claim 35, wherein the maintenance regimen includes an administration of neuregulin every 3, 5, 7 or 10 days.

37. A pharmaceutically effective amount of neuregulin for treating a chronic heart failure patient whose plasma level of NT-proBNP is within a favorite treatment zone prior to neuregulin treatment.

38. The pharmaceutically effective amount of neuregulin of claim 37, wherein said favorite treatment zone is ?4000 fmol/ml.

39. The pharmaceutically effective amount of neuregulin of claim 37, wherein said favorite treatment zone is between 1600 fmol/ml and 4000 fmol/ml.

40. The pharmaceutically effective amount of neuregulin of claim 37, wherein said favorite treatment zone is ?1600 fmol/ml.

41. The pharmaceutically effective amount of neuregulin of claim 37, wherein the plasma level is measured by immunoassay.

42. The pharmaceutically effective amount of neuregulin of claim 37, wherein the neuregulin is neuregulin-1.

43. The pharmaceutically effective amount of neuregulin of claim 37, wherein the neuregulin protein comprises of EGF-like domain of neuregulin-1.

44. The pharmaceutically effective amount of neuregulin of claim 37, wherein the neuregulin comprises of SEQ ID NO: 1.

45. A pharmaceutically effective amount of neuregulin for treating a chronic heart failure patient whose heart function is NYHA class II or III.

46. The pharmaceutically effective amount of neuregulin of claim 45, wherein the neuregulin is neuregulin-1.

47. The pharmaceutically effective amount of neuregulin of claim 45, wherein the neuregulin protein comprises of EGF-like domain of neuregulin-1.

48. The pharmaceutically effective amount of neuregulin of claim 45, wherein the neuregulin comprises of SEQ ID NO: 1.

49. A method of selecting a heart failure patient for treatment by neuregulin, comprises performing a companion diagnostic test before treatment and decide whether the result of the test is indicative for the treatment by neuregulin.

50. The method of claim 49, wherein the companion diagnostic test is measuring the plasma level of NT-proBNP in said patient.

51. The method of claim 50, wherein the result of the test is indicative for the treatment by neuregulin when the plasma level of NT-proBNP?4000 fmol/ml.

52. The method of claim 50, wherein the result of the test is indicative for the treatment by neuregulin when the plasma level of NT-proBNP is between 1600 fmol/ml and 4000 fmol/ml.

53. The method of claim 50, wherein the result of the test is indicative for the treatment by neuregulin when the plasma level of NT-proBNP is ?1600 fmol/ml.

54. The method of claim 50, wherein the plasma level is measured by immunoassay.

55. The method of claim 49, wherein the companion diagnostic test is evaluating heart function by NYHA heart function classification.

56. The method of claim 50, wherein the result of the test is indicative for the treatment by neuregulin when the heart function is NYHA class II or III.

57. The method of claim 49, wherein the neuregulin is neuregulin-1.

58. The method of claim 49, wherein the neuregulin protein comprises of EGF-like domain of neuregulin-1.

59. The method of claim 49, wherein the neuregulin comprises of SEQ ID NO: 1.

60. A diagnostic kit for selecting a heart failure patient for treatment by neuregulin, wherein diagnostic kit comprises immunoassay reagents to measure plasma level of NT-proBNP in a heart failure patient, wherein a plasma level of ?4000 fmol/ml is indicative of the patient being suitable for heart failure treatment by neuregulin.

61. The diagnostic kit of claim 60 wherein a plasma level of between 1600 fmol/ml and 4000 fmol/ml is indicative of the patient being suitable for heart failure treatment by neuregulin.

62. The diagnostic kit of claim 60, wherein a plasma level of ?1600 fmol/ml is indicative of the patient being suitable for heart failure treatment by neuregulin.

Description

B. EXAMPLES

Example 1: The Effect of Neucardin? Administration by Different Routes on the Survival Rate of Rats with CHF

INTRODUCTION

[0051] In this study, we used a coronary artery ligation (CAL) induced CHF model to investigate whether administration of Neucardin? by IV drip using a micro-injection pump or by subcutaneous (SC) bolus had any effects on survival rate and cardiac hemodynamics, 120 days after the initiation of administration of Neucardin? 4 weeks after CAL. Echocardiography and cardiac remodeling were also used to determine cardiac function and recovery from CAL.

2. Methods:

2.1. Test animals:

[0052] Strain, Origin: Wistar rats, Shanghai SLAC Laboratory Animal CO. LTD; Weight, 200+10 g, male;

2.2 Test Article:

2.2.1 Neucardin?

[0053] Identification: Recombinant human neuregulin-1 for injection (rhNRG-1, Neucardin?) [0054] Lot Number: 200607009 [0055] Manufacturer: Zensun (Shanghai) Sci & Tech Co., Ltd [0056] Dose form: Lyophilized powder [0057] Appearance: White or off-white cake [0058] Labeled Content of rhNRG-1: 250 ?g/vial [0059] Specific activity: 4897 U/vial [0060] Storage conditions: 2?8? C.

2.2.2 Vehicle:

[0061] Identification: Placebo for recombinant human neuregulin-1 [0062] Dose form: Lyophilized powder [0063] Appearance: White or off-white cake [0064] Composition: Human serum albumin, mannitol, phosphate, NaCl [0065] Storage conditions: 2?8? C.

2.3 Procedure:

2.3.1 to Establish the Rat CHF Model:

[0066] The LAD of the rats was ligated. Briefly, the rats were anesthetized with ketamine hydrochloride (100 mg/kg, IP) and their chest was shaved and sterilized. The rats were endotracheally intubated and mechanically ventilated with room air (respiratory rate 60 breaths/min, tidal volume 20 ml). A left thoracotomy was then performed at the 4th and 5th intercostal space and then the skin was incised along the left sternal border. The fourth rib was then cut proximal to the sternum. The pericardial sac was perforated and the heart was exposed. The LAD was ligated approximately 2 mm from its origin using a 6-0 silk suture. Subsequently, the air within the thorax was removed and the chest was closed in three layers (ribs, muscles and then skin). The rats were then allowed to resume spontaneous respiration, recover from the anesthesia and were then returned to their cages. Rats were maintained during a period of 4 weeks, then echocardiography evaluated, included in the formal study if they were shown an EF % value of 30-45%. Animals from all Groups were housed 5 per cage, fed ad libitum with standard diet and had free access to pure water. Room temperature was maintained at 21+1? C. and in a12 h light/dark cycle.

2.3.2 IV Drip Via Microinjection Pump:

[0067] The method of IV drip of vehicle or Neucardin? was through the tail vein. For this procedure, an appropriate rat restrainer was used according to the weight of the animal. The rat was placed near the restrainer and was gently placed into the apparatus. Normally the rats entered the restrainer without aid. Subsequently the tail of the rat was swabbed with a gauze dampened with alcohol to increase blood flow to the tail vein and to the intenerate skin corneum. The two lateral (on the side) tail veins were located and with the bevel of the needle facing upward with the needle almost parallel to the vein, the needle was inserted 2 mm into the tail vein 2-3 cm from the end of the tail. To confirm that the needle was successfully inserted into the tail vein, blood was extracted into the hub of the needle. The needle was fixed into the tail using medical tape. The infusion of drug or vehicle at the appropriate rate (0.2-0.4 ml/h) by microinjection pump or bolus injection was initiated.

2.3.3 SC Bolus

[0068] The SC bolus of vehicle or Neucardin? was from the back of the rat. For this procedure, an appropriate rat restrainer was used according to the weight of the animal. The back of the rat was swabbed with gauze dampened with alcohol to sterilize the skin. With the bevel of the needle facing upward with the needle almost parallel to the skin, the needle was subcutaneously inserted 3-4 cm into the back of the rat. The needle was fixed onto the back using medical tape and connected to the perfusion tube. Then, the rat was placed near the restrainer and was gently placed into the apparatus. Normally the rats entered the restrainer with no aid. After fasten the restrainer, the bolus injection was initiated.

2.3.4 Experiment Groups and Drug Infusion:

[0069] MI rats were randomized by EF % value into four Groups as follows: [0070] Group A (Negative control) for both IV and SC bolus. n=58 rats: IV drip of vehicle for 10-days by micro-injection pump at a speed of 0.2 ml/h for 8 h each day for the first 10 days, SC bolus of vehicle (same volume as Neucardin?), every 5 days until Day 120; [0071] Group B (SC bolus Neucardin?), n=58: IV drip of vehicle by micro-injection pump at a speed of 0.2 ml/h for 8 h each day in the first 10 days, SC bolus Neucardin? (10 ?g/day), every 5 days until Day 120; [0072] Group C (IV drip Neucardin?), n=57: IV drip of Neucardin? (0.625 ?g/kg/h) by micro-injection pump at a speed of 0.2 ml/h for 8 h each day for the first 10 days, SC bolus of vehicle (same volume as Neucardin?), every 5 days until Day 120. [0073] Group D (IV drip and SC bolus Neucardin?), n=57: IV drip of Neucardin? (0.625 ?g/kg/h) by micro-injection pump at a rate of 0.2 ml/h for 8 h per day for the first 10 days, SC bolus of vehicle (same volume as Neucardin?) at 1st, 6th, 11th day, and then SC bolus Neucardin? (10 ?g/kg), every 5 days from 16th day to the end.
2.3.5 Data acquisition

[0074] Survival rate; Echocardiography parameters; Hemodynamics parameters;

3. Results

3.1 Survival rate:

[0075] Table 1 illustrates the survival rates between each Group. The survival rates were 48.3%, 62.1%, 64.9% and 82.5% in the IV drip & SC bolus of vehicle Group A, SC bolus of Neucardin? Group B, IV drip of Neucardin? Group C and IV drip & SC bolus of Neucardin? Group D, respectively. All the survival rate or mean survival time of mortalities in Group B, C and D were improved or prolonged compared to Group A with Group D had best efficacy.

TABLE-US-00001 TABLE 1 Mortality, Survival rate and Mean survival time in the four Groups Survival Mean survival Start rat rat Survival time of mortalities Group Treatment number Deaths number rate (%) in days ? S.E. A Vehicle 58 30 28 48.3% 83.8 ? 5.9 B SC bolus Neucardin? 58 22 36 62.1% 91.4 ? 5.5 C IV drip Neucardin? 57 20 37 64.9% 97.5 ? 5.1 D SC bolus & IV drip Neucardin? 57 10 47 82.5% 107.5 ? 4.1

3.2 Echocardiography Parameters:

[0076] Echocardiography parameters were shown in Table 2. Four-weeks after coronary artery ligation and before administration of the test article, the CHF rats were randomized into four Groups according to their EF % values. As shown in Table 2, there were no significant differences between the four Groups before treatment (BT). 120 days after the start of administration, the EF % values were 30.7?3.1, 32.9 ?4.1, 33.5+3.4, 36.2?4.8% in the vehicle, Neucardin? via SC bolus, Neucardin? via IV drip and Neucardin? via IV drip plus SC bolus Groups, respectively. After treatment, EF % and FS % of Group B, C and D were all higher than that of Group A.

TABLE-US-00002 TABLE 2 Echocardiography parameters in the four Groups BT LVEDd LVEDs EF % FS % Group AT N (cm) (cm) (%) (%) A. Negative control BT 58 0.987 ? 0.083 0.829 ? 0.088 38.0 ? 5.5 16.2? AT 25 1.100 ? 0.089 0.961 ? 0.090 30.7 ? 3.1 12.7? B. SC bolus Neucardin? BT 58 0.992 ? 0.066 0.831 ? 0.066 38.2 ? 4.0 16.3? AT 33 1.104 ? 0.063 0.952 ? 0.070 33.1 ? 4.1 13.9? C. IV drip Neucardin? BT 57 0.985 ? 0.061 0.824 ? 0.068 38.5 ? 4.4 16.3? AT 36 1.080 ? 0.072 0.929 ? 0.073 33.4 ? 3.4 14.0? D. SC bolus & IV drip Neucardin? BT 57 0.979 ? 0.065 0.818 ? 0.066 38.7 ? 4.3 16.5? AT 44 1.052 ? 0.087 0.893 ? 0.092 36.2 ? 4.8 15.3? BT: Before treatment; AT: After treatment;

3.3 Hemodynamic Parameters:

[0077] Table 3 shows the MAP, HR, ?dp/dt, LVEDP and LVSP values as measured in the four Groups of anesthetized animals on day 121. When Neucardin? was administered by SC bolus or by IV drip alone (Group B and C), Neucardin? significantly increased dp/dt and ?dp/dt by 19.6% and 27.1%, 22.5% and 29.8% compared to Group A. When Neucardin? was administered by both IV drip and SC bolus routes (Group D), significant increases in mean arterial pressure (MAP, 112.3?5.5 mmHg), left ventricular systolic pressure (LVSP, 139.4?9.8 mmHg), ?dp/dt (7012.1+903.0 mmHg/s), ?dp/dt (?4353.2?847.6 mmHg/s) compared to vehicle were obtained. Interestingly, these values of MAP, LVSP, ?dp/dt and ?dp/dt were 10.6%, 9.2%, 38.5% and 37.8% higher than vehicle treated rats, respectively. The results showed that Group B, C and D were all better than Group A in hemodynamic parameters with Group D had best efficacy.

TABLE-US-00003 TABLE 3 Hemodynamics parameters in the four Groups SBP DBP MAP LVSP Group Treatment N (mmHg) (mmHg) (mmHg) (mmHg) A Vehicle 14 118.7 ? 11.5 94.1 ? 12.3 102.3 ? 11.7 128.5 ? 14.7 B SC bolus Neucardin? 27 123.8 ? 11.5 95.3 ? 8.9 104.9 ? 9.5 129.5 ? 13.6 C IV drip Neucardin? 25 122.5 ? 10.5 95.0 ? 7.5 104.4 ? 8.2 131.7 ? 10.0 D SC bolus & IV drip Neucardin? 35 132.6 ? 7.1 102.1 ? 5.3 112.3 ? 5.5 139.4 ? 9.8 LVEDP dp/dt (?dp/dt) Heart rate Group Treatment N (mmHg) (mmHg/s) (mmHg/s) (Beat/min) A Vehicle 14 5.8 ? 3.5 4995.6 ? 532.2 3087.5 ? 715.7 297.2 ? 16.0 B SC bolus Neucardin? 27 4.5 ? 2.8 6050.9 ? 1231.3 4013.8 ? 838.3 292.6 ? 23.0 C IV drip Neucardin? 25 4.0 ? 3.2 6199.9 ? 709.5 4098.9 ? 823.5 296.3 ? 13.5 D SC bolus & IV drip Neucardin? 35 3.9 ? 2.5 7012.1 ? 903.0 4353.2 ? 847.6 292.5 ? 19.1

4. Conclusion

[0078] A combined administration of Neucardin? by IV drip & SC bolus or administration of the peptide given by each route alone all increased the survival rate of rats with CHF induced by CAL and improved cardiac functional parameters compared to rats treated with vehicle.

Example 2: A Randomized, Double-Blinded, Multi-Center, Placebo Controlled Study to Evaluate the Efficacy and Safety of Recombinant Human Neuregulin 1 in Patients with Chronic Heart Failure Based on Standard Treatment

[0079] To evaluate the efficacy of recombinant human neuregulin-1 for injection on chronic heart failure, a phase II, double-blinded, multi-center, placebo controlled, standard treatment based study was carried out in multiple clinical centers in China. A total of 195 patients with NYHA Class II or III stable chronic heart failure were enrolled and randomized into three groups: placebo, or 0.6 ?g/kg and 1.2 ?g/kg of rhNRG-1. There were no significant variations in demographics or background therapies among groups. According to the schedule, patients were administered the drug for 10 consecutive days in the hospital first, after finishing the day 11 follow up, they were discharged from the hospital. Another two on site follow up were at day 30 and day 90. A telephone interview was conducted one year after the last patient enrolled. [0080] Investigational product: [0081] Specification: Neucardin?, 61 amino acid polypeptide comprises the EGF-like domain of Neuregulin-1 ?2 isoform, with the molecular weight of 7054 Dal (1 ?g=0.14 nmol). 250 ?g (5000 EU)/vial (lug=20 EU). [0082] Preparation: For injection. [0083] Mode of administration: Intravenously drip. [0084] Storage: in safe place, with limited access and protected from light, at 3-8? C. [0085] Placebo: [0086] Specification: Excipient for Neucardin? (250 ?g/vial without active recombinant human neuregulin-1 protein). [0087] Dosage groups:

TABLE-US-00004 Dosage 0 ?g/kg/day 0.6 ?g/kg/day 1.2 ?g/kg/day Administration Intravenous infusion Volume 50 ml Course 10 hours per day, for consecutive 10 days

Study Procedure

[0088] Criteria for participation in the trial included patients with CHF (NYHA class II or III) between the ages of 18 and 65 years old, LVEF?40%, in relatively stable clinical condition (including clinical signs, symptoms and accepted standard treatment for CHF at the target dose or maximum tolerance dose for over 1 month). Major exclusion criteria included acute myocardial infarction, hypertrophic cardiomyopathy, constrictive pericarditis, significant valve disease or congenital heart disease, severe pulmonary hypertension, systolic blood pressure ?90 mmHg or >160 mmHg, severe ventricular arrhythmia, cardiac surgery or a cerebrovascular event within the previous six months, claustrophobia or pregnant female subjects. All patients provided witnessed written consent.

[0089] Patients were randomly assigned to three groups, treated with placebo or rhNRG-1 (0.6 or 1.2 ?g/kg/day) for 10 consecutive days, after finishing the day 11 follow up, they were discharged from the hospital. Another two on site follow up were at day 30 and day 90. Blood samples of each patient were collected before treatment and at day 11, 30 and 90. Plasma NT-proBNP was tested in the core lab with NT-proBNP assays (kit from Biomedica). One year after the last patient enrolled, the telephone interview was made for collecting the information of re-hospitalizations, all telephone interviews were recorded in a special form with investigators signature.

[0090] Of the 48 patients with available re-hospitalization information in the placebo group, 12 (25.0%) were rehospitalized for worsening heart failure at least once. For the 0.6 ?g/kg group, only 4 (8.7%) of the 46 patients readmitted to the hospital (P=0.05 compared to placebo); Rehopitalization rate of the 1.2 ?g/kg group was 22.0% (11/50). The average times of re-hospitalizations was 0.458 (22/48) per patient in the placebo group, while they were reduced by 57.4% and 17.0% respectively in the 0.6 (8/41) and 1.2 ?g/kg group (19/50).

[0091] In the placebo group, the NT-proBNP were almost the same during the study while compare to the baseline. At day 11, the NT-proBNP was significantly increased in rhNRG-1 treated groups (from 1853+1512 to 2399+1841 fmol/ml in 0.6 ?g/kg group, P?0.01; from 1562?1275 to 2774?1926 fmol/ml in 1.2 ?g/kg group, P?0.01). But his increase was transient and was not caused by a worsening heart function as the cardiac function shown to be increased, the NT-proBNP decreased to the baseline level at Day 30 and Day 90 in the 1.2 ?g/kg group. Moreover, in the 0.6 ?g/kg group, the NT-proBNP was significantly reduced at day 30 (1323+1124 fmol/ml, P=0.01) and day 90 (1518+1403 fmol/ml, P=0.01) while compare to the baseline.

[0092] These results showed that rhNRG-1 treatment can reduce the re-hospitalizations and the plasma level of NT-proBNP, which may indicate rhNRG-1 can provide long-term benefits to chronic heart failure patients.

Example 3: A Randomized, Double-Blinded, Multi-Center, Placebo Controlled Survival Study of Recombinant Human Neuregulin 1 in Patients with Chronic Heart Failure Based on Standard Treatment

[0093] To evaluate the efficacy of recombinant human neuregulin-1 for injection on chronic heart failure, a phase II, double-blinded, multi-center, placebo controlled, standard treatment based study was carried out in multiple clinical centers in China. A total of 351 patients with NYHA Class III or IV stable chronic heart failure were enrolled and randomized into placebo group or rhNRG-1 group (0.6 ?g/kg). There were no significant variations in demographics or background therapies among groups. According to the schedule, patients were administered with the drug for 10 consecutive days in the hospital, after finishing the day 11 follow up, they were discharged from the hospital, and were administered with the drug once weekly from the 3rd week till the 25th week as out-patient. Blood samples of each patient were collected before treatment (baseline) and at each follow up. Plasma NT-proBNP level was tested in the core lab with NT-proBNP assays (kit from Biomedica). The survival information was collected at 52th week of the study.

Investigational Product:

[0094] Specification: Neucardin?, 61 amino acid polypeptide comprises the EGF-like domain of Neuregulin-1 ?2 isoform, with the molecular weight of 7054 Dal (lug=0.14 nmol). 250 ?g (5000 EU)/vial (lug-20 EU). [0095] Preparation: For injection. [0096] Mode of administration: Intravenously drip or infusion. [0097] Storage: in safe place, with limited access and protected from light, at 3-8? C. [0098] Placebo: [0099] Specification: Excipient for Neucardin?. 250 ?g/vial and without active recombinant human neuregulin-1 protein. [0100] Dosage and regimens:

TABLE-US-00005 Day 1-10 Week 3-25 Dose 0.6 ?g/kg/day rhNRG-1 0.8 ?g/kg/day rhNRG-1 or placebo or placebo Route Intravenous drip Intravenous infusion regimen 10 hours per day for 10 days 10 minutes infusion weekly

[0101] Criteria for participation in the trial included patients with CHF (NYHA class III or IV) between the ages of 18 and 80 years old, LVEF?40%, in relatively stable clinical condition (including clinical signs, symptoms and accepted standard treatment for CHF at the target dose or maximum tolerance dose for over 1 month). Major exclusion criteria included acute myocardial infarction, hypertrophic cardiomyopathy, constrictive pericarditis, significant valve disease or congenital heart disease, severe pulmonary hypertension, systolic blood pressure ?90 mmHg or >160 mmHg, severe ventricular arrhythmia, cardiac surgery or a cerebrovascular event within the previous six months, claustrophobia or pregnant female subjects. All patients provided witnessed written consent.

[0102] The all-cause mortality of the placebo group at 52 week is 15.91%, with 28 death in 176 patients, while the number is 9.71% in rhNRG-1 group, with 16 death in 175 patients completed the trial (Hazard ratio=0.425, 95% CI 0.222-0.813, p=0.0097). Considering the mortality caused by cardiovascular events, the number of the placebo group at 52 week is 14.77%, with 26 death in 176 patients, and 9.71% in the rhNRG-1 group. So from the results we can find around 40% decrease of the mortality of rhNRG-1 administration compared with placebo group, even the placebo group were still maintain their previous standard treatment for chronic heart failure.

[0103] We also analyzed the all-cause mortality based on the stratification of baseline NT-proBNP. When the NT-proBNP level is stratified into 3 stratums as ?1600 fmol/ml, >1600 fmol/ml and ?4000 fmol/ml, or >4000 fmol/ml, the mortality of rhNRG-1 group vs placebo group are 1.49% vs 8.49%, 8.96% vs 23.33%, and 26.67% vs 28.00%, respectively. And if the NT-proBNP level is stratified as ?4000 fmol/ml or >4000 fmol/ml, the mortality of rhNRG-1 group vs placebo group are 5.22% vs 14.89% (p=0.0092), and 26.67% vs 28.00%, respectively. These results show statistical significance that rhNRG-1 can effectively improve the survival of chronic heart failure patients.

[0104] Further, the patients were stratified with their baseline NYHA heart function class, to be class III or class IV. The all-cause mortality of class III patients in rhNRG-1 group or placebo group is 6.06% (8 death in 132 patients) and 15.49% (22 death in 142 patients), respectively, p=0.0189. While the all-cause mortality of class IV patients in rhNRG-1 group or placebo group is 20.93% (9 death in 43 patients) and 17.65% (6 death in 34 patients), respectively, p=0.778.