Mechanisms Associated With Favorable Responses to Peginterferon-Alpha Add-on Therapy Following Long-term Nucleos(t)Ide Analogue Treatment in Patients With Chronic Hepatitis B

Chronic hepatitis B virus (HBV) infection is a leading cause of liver associated morbidity
and mortality. Currently available first-line therapies for treatment of chronic hepatitis B
include pegylated interferon-alpha and the nucleos(t)ide analogues (NUCs) entecavir and
tenofovir. These were shown to effectively suppress viral replication, but their ability to
induce durable off-treatment response is limited to a small subset of patients. Combination
treatment with peginterferon and NUCs has been attempted in several randomized controlled
trials, with no apparent advantage over either agent given alone. In these studies however,
treatment with peginterferon was initiated either simultaneously or shortly after NUCs
administration. The efficacy of peginterferon following long-term viral suppression with NUCs
was only tested in one small pilot study, nevertheless showing 60% HBsAg loss rate.

The underlying mechanisms responsible for improved efficacy of peginterferon in this setting
are unknown and warrant further investigation. In this single arm study we propose to
evaluate the efficacy and mechanisms associated with response to peginterferon add-on therapy
following a minimum of 192 weeks of viral suppression induced by NUCs in a group of chronic
HBV infected patients. Sixty patients with either HBeAg positive (n=30) or negative (n=30)
chronic HBV infection will be enrolled to this study. After medical evaluation and
pretreatment liver biopsy, treatment with subcutaneous injections of pegylated interferon
alpha-2a 180 g per week will be given for a total of 24 weeks, followed by an off-treatment
evaluation period of 48 weeks. A second liver biopsy will be performed six hours following
the first peginterferon injection. Primary end-point for this study will be the change in
interferon-stimulated-genes response before and after first interferon injection in
responders versus non-responders to treatment. The responsiveness to IFN-based therapy of
treatment responders vs nonresponders will additionally be evaluated by studying intrahepatic
and peripheral blood natural killer cells. The study will also assess HBeAg and HBsAg loss
and seroconversion rates in comparison to historical controls treated with either
peginterferon or NUCs monotherapy. Finally, we will assess whether treatment responders
develop an HBV-specific T cell response similar in quantity and quality to that of patients
who spontaneously resolve HBV infection.

- INCLUSION CRITERIA:

Inclusion criteria: HBeAg positive group

1. Age >18 years and older, male or female.

2. Known serum HBsAg and HBeAg positivity at the time of screening.

3. Ongoing treatment with one or more NUCs for at least 192 weeks before study entry.
Subjects may have a brief interruption of treatment for medical reasons (e.g. breast
feeding) not to exceed 8 weeks and none within the 48 weeks before study entry.

4. HBV DNA levels <100 IU/mL, measured at least 12 months prior to, and upon enrollment
to the study.

5. ALT level less than or equal to 2 ULN based on at least two determinations taken at
least one month apart during the 24 weeks before study entry with the second being at
time of screening

6. Written informed consent

Inclusion criteria: HBeAg negative group

1. Age >18 years and older, male or female.

2. Known serum HBsAg positivity and HBeAg negativity at the time of screening.

3. Ongoing treatment with one or more NUCs for at least 192 weeks before study entry.
Subjects may have a brief interruption of treatment for medical reasons (e.g. breast
feeding) not to exceed 8 weeks and none within the 48 weeks before study entry.

4. HBV DNA levels <100 IU/mL, measured at least 12 months prior to, and upon enrollment
to the study

5. ALT level less than or equal to 2 ULN based on at least two determinations taken at
least one month apart during the 24 weeks before study entry with the second being at
time of screening

6. Written informed consent

EXCLUSION CRITERIA:

Exclusion criteria (for both eAg positive and negative patients)

1. Co-infection with HDV as defined by the presence of anti-HDV in serum and/or HDV
antigen in the liver.

2. Co-infection with HCV as defined by the presence of HCV RNA in serum.

3. Co-infection with HIV as defined by the presence of anti-HIV in serum.

4. Decompensated liver disease as defined by serum bilirubin >2.5 mg/dL (with direct
bilirubin > 0.5 mg/dL), prothrombin time of greater than 2 seconds prolonged, a serum
albumin of less than 3 g/dL, or a history of ascites, variceal bleeding or hepatic
encephalopathy.

5. Presence of other causes of liver disease, (i.e. hemochromatosis, Wilson disease,
alcoholic liver disease, severe nonalcoholic steatohepatitis defined as the presence
of marked ballooning injury on liver biopsy, alpha-1-anti-trypsin deficiency).

6. A history of organ transplantation or in the absence of organ transplantation, any
immunosuppressive therapy requiring the use of more than 5 mg of prednisone (or its
equivalent) daily.

7. Significant systemic illness other than liver diseases including congestive heart
failure, renal failure, chronic pancreatitis and diabetes mellitus with poor control,
that in the opinion of the investigator may interfere with therapy.

8. Pregnancy or inability to practice contraception in patients capable of bearing or
fathering children

9. Lactating women.

10. Hepatocellular carcinoma (HCC), or the presence of a mass on imaging studies of the
liver that is suggestive of HCC, or an alpha-fetoprotein level of greater than 500
ng/mL.

11. eGFR < 50 ml/min, serum creatinine > 1.3 mg/dl or urine protein >1 gram/24-hours

12. History of hypersensitivity to pegylated interferon-alpha

13. Platelet count <70 mm(3)/dL

14. Hgb <12 g/dL for males and <11 g/dL for females

15. Active ethanol/drug abuse/psychiatric problems such as major depression,
schizophrenia, bipolar illness, obsessive-compulsive disorder, severe anxiety, or
personality disorder that, in the investigator s opinion, might interfere with
participation in the study.

16. History of malignancy or treatment for a malignancy within the past 3 years (except
adequately treated carcinoma in situ or basal cell carcinoma of the skin).

17. Any medical condition requiring, or likely to require, chronic systemic administration
of corticosteroids or other immunosuppressive medications during the course of this
study.

18. History of immune-mediated disease, or cerebrovascular, chronic pulmonary or cardiac
disease associated with functional limitation, retinopathy, uncontrolled thyroid
disease, poorly controlled diabetes or uncontrolled seizure disorder, as determined by
a study physician.

19. Presence of conditions that, in the opinion of the investigators, would not allow the
patient to be followed in the current study for at 1 year.

20. For subjects who interrupt therapy, documentation of a viral load >1,000 IU/ml while
off therapy.