1. Hepatitis B Jay H. Hoofnagle, M.D. Division of Digestive Diseases and Nutrition National Institute of Diabetes and Digestive and Kidney Diseases National Institutes of Health FDA Advisory Panel Meeting: August 7, 2002

2. Hepatitis B HBV, small double stranded DNA virus Hepadnaviridae Infection restricted to humans and higher apes High levels in blood (10 2 to 10 10 copies/ml) Causes both acute and chronic hepatitis Parenteral, sexual and maternal-infant spread Marked geographic variation in incidence Common in Asia & Africa, uncommon in the United States and Western Europe

3. Hepatitis B Virus HBsAg HBcAg Dane ParticleTubule Sphere HBeAg

5. A A A A A A A A A A A A A A A A A A A A A A A A 2.1kb RNA 2.4kb RNA 3.5kb RNA 0.7kb RNA Pre-S1 Pre-S2 ORF-S ORF-P ORF-X ORF-C DR1 5’ +strand -strand Pre-C DR2 Hepatitis B Virus RNAs

6. Hepatitis B Viral Genome Circular, partially doubled-stranded DNA Four open reading frames –HBsAg (pre-S1, pre-S2 and S) –HBcAg (pre-core & core) –Polymerase (multifunctional) –HBxAg (transactivating factor) Replicates largely in liver Through RNA intermediate and reverse transcription

7. Infectious cycle of hepatitis B virus degradation - antigen-specific - non-specific Y Y Y Y Y Y Y Y cell death Virus - half-life: 1-2 days -production: 10 11 -10 13 /day mutation rate: 1-3 x 10 -5 /site/yr Zeuzem et al: 2000

8. Hepatitis B Virus Mutants Variations in nucleotide sequence in one of the HBV genes can result in change in the virological and, in some cases, clinical features of the infection. S gene: vaccine or HBIG escape mutants C gene: can affect disease severity or serological and clinical manifestations P gene: can effect replicative efficiency and resistance to antiviral therapy

9. Hepatitis B Core Antigen Mutants Nucleocapsid region: Pre-core and Core Pre-core: May result in inability to produce HBeAg. HBeAg-negative mutants. Most frequently, G  A at nt 1896. Core: Substitutions in core region are frequent among pts with severe disease or resistance to interferon, in areas of major B cell and T cell epitopes, thus important in T cell cytotoxicity and viral clearance

10. HBeAg-negative Variants G  A at nt 1896 creates a stop codon in the pre- core region that therefore blocks the synthesis of HBeAg. nt 1896 is in the highly structured stem-loop  encapsidation signal region of HBV RNA and base-pairs with nt 1858 If nt 1858 is a T (ayw, adr, some adw), stem loop of  is maintained by either G or A; if it is a C (adw), stem loop is disrupted by A and replication is stopped. Thus HBeAg-negative variants are more common with genotypes B, C and D than genotype A

11. Outcome of Hepatitis B Virus Infection Acute Hepatitis B Asymptomatic subclinical infection Fulminant Hepatitis Chronic Hepatitis B Inactive Carrier StateCirrhosis Liver Cancer ? 65% 35% <1% 5% 30% 50%

12. Typical Acute Hepatitis B ALT HBsAg HBeAg HBV DNA Normal Months After Exposure ALT and HBV DNA IU/L and million copies/ml Symptoms

13. Typical Chronic Hepatitis B ALT HBsAg HBeAg HBV DNA Normal Months After Exposure ALT and HBV DNA IU/L or million copies/ml

14. Chronic Hepatitis B: Transition to Inactive Carrier State ALT `` HBsAg HBeAg HBV DNA Normal Months After Exposure ALT and HBV DNA IU/L and million copies/ml Anti-HBe

15. Evolution of HBeAg Negative Mutant ALT HBsAg HBV DNA Normal ALT levels Months ALT and HBV DNA IU/L and million copies/ml Anti-HBeHBeAg

16. Chronic Hepatitis B: Three Clinical Forms: HBeAg Positive Chronic Hepatitis B HBeAg, raised ALT, HBV DNA in serum and chronic hepatitis on biopsy HBeAg Negative Chronic Hepatitis B Anti-HBe, raised ALT and HBV DNA in serum, chronic hepatitis on biopsy Inactive HBsAg Carrier State Anti-HBe, normal ALT & no HBV DNA, minimal nonspecific changes on biopsy

17. Chronic Hepatitis B: Clinical Forms: HBV DNA levels l HBeAg Positive Chronic Hepatitis B 10 7 to 10 11 copies per ml l HBeAg Negative Chronic Hepatitis B 10 4 to 10 8 copies per ml l Inactive HBsAg Carrier State < 10 1 to 10 4 copies per ml

18. 350 3.5.035 Log 10 copies/mL pg/mL 8 6 4 2 10 35,000 Dynamic Range of Detection of HBV DNA: 5 Assays HBV DNA Detection.0035

19. Genotypes of Hepatitis B Virus Aadw, adw2, ayw1US, Northern Europe, Africa Badw2, ayw1China, Indonesia, Vietnam Cadr, ayr China, Korea, Japan, Vietnam Dayw2, ayw3Mediterranian, Middle East, India Eayw4West Africa Fadw4Polynesia, US (rare) GEurope, US (rare) TypeSubtypeGeographical Distribution

20. Acute Hepatitis B Sentinel County Study: 1982-98 Currently, HBV causes 34% of viral hepatitis Decline in incidence by 76% between 1987-98 20% hospitalized, 1% fatal Gradual rise in median age (27 to 32 yrs) More common in men than women African-Americans > Hispanic whites > whites Current proportions with risk factors  Injection drug use: 14%  Men who has sex with men: 15%  Heterosexual activity: 40%  Occupational exposure: 2% Goldstein et al: 2002

21. Vaccine licensed HBsAg screening of pregnant women Routine infant immunization OSHA Rule Routine adolescent immunization Decline in MSM & HCW Decline in injecting drug users Decline in high-risk heterosexuals Year Infections per 100,000 Acute Hepatitis B Incidence in the U.S.: 1978-1998 Alter et al: CDC

22. Chronic Liver Disease: United States 1999 Hepatitis C 57% Alcohol 25% Hepatitis B Other NASH 10% Bell et al 2001 Hepatitis B accounted for only 4.4% of newly- diagnosed chronic liver disease

23. Chronic Hepatitis B Long-Term Complications Cirrhosis Hepatocellular carcinoma Glomerulonephritis Polyarteritis Nodosa

24. Chronic Hepatitis B Histology Necroinflammatory Changes (Grade) Periportal inflammation and necrosis (piecemeal necrosis, interface hepatitis) Lobular inflammation and single cell necrosis Portal inflammation Fibrosis (Stage) Portal Septa formation Bridging fibrosis Cirrhosis

25. Chronic Hepatitis B Histology Scoring Systems Histology Activity Index (Knodell) : Periportal necrosis & inflammation (0-10) Lobular necrosis & inflammation (0-4) Portal inflammation (0-4) Fibrosis None = 0 Portal fibrosis = 1 Bridging fibrosis = 3 Cirrhosis = 4

26. Chronic Hepatitis B Histology Scoring Systems Histology Activity Index (Ishak) : Periportal necrosis & inflammation (0-4) Bridging necrosis (0-6) Lobular necrosis & inflammation (0-4) Portal inflammation (0-4) Fibrosis None = 0 Portal fibrosis = 1 or 2 Bridging fibrosis = 3 or 4 Cirrhosis = 5 or 6

27. Therapy of Hepatitis B

28. Chronic Hepatitis B Goals of Therapy Improve symptoms and quality of life Decrease infectivity Prevent progression of disease Hepatic Decompensation Death from liver disease What surrogate end-points correlate with these outcomes ?

29. Therapy of Chronic Hepatitis B: Major Issues n What are appropriate end-points? n Are they the same for different forms of HBV?  Loss of HBeAg  Loss of HBsAg  Loss of HBV DNA (fall below 10 5 copies/ml)  Normalization of ALT  Improvement in histology n What amount of follow up is appropriate in assessing benefit of therapy?

30. Definition of Responses to Therapy in Chronic Hepatitis B  Type: n Virological: Loss of HBeAg and/or HBV DNA n Biochemical: Normal ALT n Histological: Improvement in histology scores n Complete: All of above & loss of HBsAg  Timing: n Initial: within first 6 mo of therapy n End-of-therapy: when therapy is stopped n Sustained: 6 or 12 mo after stopping n Maintained: present while continuing therapy

31. Virological Response in Chronic Hepatitis B Loss of HBeAg and fall of HBV DNA levels to below 10 5 copies/mL n Occurs in 25-48% of patients given a 4-5 month course of alpha interferon n Occurs in 20-32% of patients given a 12 month course of lamivudine n Occurs in 8-12% of patients on no therapy n Is this response durable and does it result in long-term improvement in disease and lack of progression to cirrhosis and HCC?

32. Virological Response in Chronic Hepatitis B Loss of HBeAg cannot be used as an endpoint in patients with HBeAg-negative disease n Generally rely upon decrease in HBV DNA to below 10 5 copies/ml n HBV DNA levels, however, can fluctuate widely, and with nucleoside therapy will rapidly return to baseline when treatment is stopped. n How durable is decrease in HBV DNA without other changes in viral status?

33. Virological Response in Chronic Hepatitis B Loss of HBsAg and development of anti-HBs n Occurs in 8% of patients given a 4-5 month course of alpha interferon n Occurs in 1-2% of patients given a 12 month course of lamivudine n Occurs in <1% of patients on no therapy n Extremely rare in treatment trials of HBeAg- negative chronic hepatitis B n This response is durable and associated with resolution of liver disease

34. Biochemical Response in Chronic Hepatitis B Fall of ALT levels into the Normal Range n Often accompanies loss of HBeAg or decrease in HBV DNA to below 10 5 copies/mL n Not durable unless the decrease in HBV DNA is durable. n Surrogate, indirect marker for decrease in necroinflammatory disease

35. Histological Response in Chronic Hepatitis B Improvements in Histology n Used in virtually all studies of antiviral therapy n Typically, improvement is called a > 2 point improvement in HAI score (0-22) compared to baseline n However, necroinflammatory scores can change rapidly and improve and worsen n Fibrosis scores represent best evidence for progression of disease and are unlikely to improve with treatment

36. Alpha Interferon n Human cytokine made by lymphocytes in response to viral infection n Acts through cell-surface receptors n Activates Jak/Stat system n Induces transcription of proteins with antiviral activity (2-5 OAS, PKR, eIF2) n Recombinant human alpha interferon n Pegylated forms now available

37. Chronic Hepatitis B Long-term Response to Interferon HBeAg HBsAg Anti-HBe Anti-HBs Alpha Interferon ALT Months After Start of Therapy Patient A HBV DNA (dot blot)

38. Chronic Hepatitis B Response to Interferon & Relapse HBeAg HBsAg Alpha Interferon ALT Months After Start of Therapy HBeAgAnti-HBe Patient B

39. HBeAg-Negative Chronic Hepatitis B Response to Interferon and Relapse HBsAg Alpha Interferon ALT HBV DNA Months After Start of Therapy Patient C Anti-HBe ++++ ---

40. Interferon for Chronic Hepatitis B: Problems Effective in only 1/3 rd of cases Expensive Side effects are common and can be severe Not appropriate for many categories of pts: –Immune suppressed –Renal failure or dialysis –Solid organ transplant –Decompensated liver disease

41. Lamivudine n Negative enantiomer of 3-thiacytidine n Both an unnatural nucleoside and chain terminator n Highly active against HBV in vitro n Dose: 100 mg, once daily by mouth n Approved for use in chronic hepatitis B as one year course of therapy n Continuous, long-term use is common but must be considered experimental

42. Lamivudine Therapy Maintained Response HBV DNA ALT 10 2 10 4 10 6 10 8 HBV DNA Levels Therapy with Lamivudine (100 mg/d) HAI Scores: Pre: 14 Yr 1: 4 Yr 4: 1 HBeAg HBsAg Patient B

43. HBeAg-Negative Chronic Hepatitis B HBsAg and Anti-HBe Lamivudine ALT Months After Start of Therapy Patient C HAI = 4 200 copies/ml HAI = 13 53.1 million copies/ml HAI = 1 <100 copies/ml Liver Biopsy HBV DNA

44. Lamivudine Therapy: Viral Resistance HBV DNA ALT 10 2 10 4 10 6 10 8 HBV DNA Levels Normal Therapy with Lamivudine (100 mg) HAI Scores: Pre: 14 Yr 1: 10 Yr 4: 17 (Cirrhosis) wt YVDD 10 Patient D

45. 9.89.9 2.5 8.4 1.3 7.1 13 9 4 99 Total HAI Scores (0 to 18) Lamivudine for Chronic Hepatitis B Histology Activity Index Scores

46. 4.2 2.9 1.3 3.9 3.3 4.3 13 4 9 Fibrosis Score (0 to 6) Lamivudine for Chronic Hepatitis B: Fibrosis Scores 99

47. Lamivudine Therapy Late Relapse HBV DNA ALT 10 2 10 4 10 6 10 8 HBV DNA Levels Therapy with Lamivudine (100 mg/d) HAI Scores: Pre: 14 Yr 1: 4 Yr 4: 1 HBeAg HBsAg Patient B

48. 85% 50%

49. Lamivudine n The major shortcoming of long-term lamivudine therapy for hepatitis B is emergence of lamivudine resistance n Occurs in 20%-30% of patients per yr, approaching 90% by 5 yrs n Loss of HBsAg, but not loss of HBeAg, appears to reliably predict long-term benefit & ability to stop lamivudine n Future studies should focus on combinations that might prevent resistance

50. Optimal Therapy of Hepatitis B? &Monotherapy or combination therapy? &For a defined period (48 wks) or continuous? &For all pts or only those with mod-severe disease? &If monotherapy, which agent? &If combination, which combination? &Standard interferon and lamivudine &Pegylated interferon and lamivudine &Lamivudine and adefovir &Lamivudine and entecavir

51. Management of Hepatitis B l Initial evaluation: Routine liver tests, HBsAg, HBeAg & anti-HBe, HBV DNA, anti-HDV, abdominal US l If ALT elevated & HBV DNA present: liver biopsy and assess for therapy l Reserve therapy for patients with significant underlying liver disease l Therapy? Lok, Heathcote & Hoofnagle: 2001

52. Therapy of Chronic Hepatitis B: Future Directions n Focus must be on combination therapy n Long term outcomes with histological verification of long-term benefit n Loss of HBsAg might be a gold standard n Appropriate directions:  Combinations of alpha interferon & lamivudine  Nucleoside combinations without cross-reactive resistance (lamivudine & adefovir or entecavir)  Novel approaches: immunological or molecular