1. Case Three: Serendipitous testing in hepatitis
Educational Workshops 2015
Case Three: Serendipitous testing in hepatitis
We are grateful to Emmanuel Nsutebu, Consultant
Infectious Diseases Physician at Royal Liverpool
University Hospital for composing this case.

2. May 2010 55 year old man Born in the Phillipines -moved to UK 2005
New diagnosis of light chain multiple myeloma after presenting with back pain
Previously fit and well
Works as plant operator; married with 6 children

3. Treatment planned…
8 cycles of chemotherapy with cyclophosphamide, thalidomide and dexamethasone
Followed by high dose mephalan autologous stem cell transplant

4. Who would you screen for hepatitis B?
1. All patients before immunosuppression?
2. Selectively, based on risk factors?

5. Prevalence of Hepatitis B carriers
Figure Worldwide prevalence of hepatitis B carriers and primary hepatocellular carcinoma. (Courtesy Centers for Disease Control and Prevention, Atlanta.)
From Murray et. al., Medical Microbiology 5th edition, 2005, Chapter 66, published by Mosby Philadelphia,,

6. Interpreting hepatitis B test results
Marker
Abbreviation
Interpretation
Hepatitis B surface antigen
HBsAg
Current infection
Hepatitis B core antibody
Anti- HBc
Infection (past or current)
Hepatitis B surface antibody
Anti -HBs
Immunity (vaccine or past infection)
Hepatitis B e antigen
HBeAg
Marker of high viral replication
Hepatitis B DNA
Not detected
Positive <2000 iu/ml
Positive >2000 iu/ml
No virus detectable in blood
Infection – low HBV level
Infection – high HBV level

7. Chronic Hepatitis B infection

8. Natural History of Chronic HBV Infection
Immunotolerance
Immune Clearance
Immune Control
(Nonreplicative)
HBV DNA
HBeAg HBeAg- HBeAb+
HBsAg HBsAg- HBsAb+
ALT
5-30 Yrs
Mos-Yrs
Mos-Yrs
Infection
Yim HJ, et al. Hepatology. 2006;43:S173-S181.

9. Natural History of Chronic HBV Infection
Immunotolerance
Immune Clearance
Immune Control
(Nonreplicative)
HBV DNA
Most transplant Patients
Normal ALT
Low/undetectable HBV DNA
HBsAg+ and HBeAg-
or HBsAg-, anti-HBc+
HBeAg HBeAg- HBeAb+
HBsAg HBsAg- HBsAb+
ALT
5-30 Yrs
Mos-Yrs
Mos-Yrs
Infection
Yim HJ, et al. Hepatology. 2006;43:S173-S181.

10. What tests would you use to screen for hepatitis B?
1. Hepatitis B surface antigen?
2. Hepatitis B core antibody?
3. Hepatitis B DNA?

11. Results

12. What form of prophylaxis can be considered if he had been exposed to hepatitis B?
Lamivudine
Tenofovir
Entecavir

13. Pre-emptive therapy vs watch and wait
Randomized, controlled trials (RCTs) have demonstrated the efficacy of prophylactic therapy compared to early therapy when reactivation occurs

14. Use of Preemptive Lamivudine Reduces Risk of HBV-Related Hepatitis
HBsAg-positive patients with lymphoma treated with high-dose chemotherapy randomized to “preemptive” vs “on-demand” lamivudine
100
Preemptive LAM 75
Survival Free From Hepatitis Due to HBV Reactivation 50
P = .002 by log-rank test
On-demand LAM
(if HBV DNA increased) 25 10 20 30 40
Pts at Risk, n Preemptive LAM
On-demand LAM Wk
15 15
12 13
10 10
9 4
6 2
Lau GK, et al. Gastroenterology. 2003;125:

15. Value of Preemptive Antivirals
HBsAg-positive patients with NHL treated with CHOP randomized to “preemptive” vs “on-demand” lamivudine
100 80
On-demand group: start LAM if ALT > 1.5 x ULN
Preemptive group: start LAM on Day 1 of CHOP 60
HBsAg Patients (%) 48 40 36 20 20 8 8
HBV Reactivation and Hepatitis Flare
HBV Reactivation and ALT >10 x ULN
HBV Reactivation and Jaundice
Death
(After ChemoTx)
Preemptive antivirals decrease HBV reactivation
Hsu C, et al. Hepatology. 2008;47:

16. Choice of Antiviral Therapy and Monitoring
Choice of therapy affected by HBV DNA level
HBV DNA < 2000 IU/mL: any therapy can be used (including lamivudine)
HBV DNA > 2000 IU/mL: entecavir or tenofovir
Choice of therapy affected by duration of therapy
> 12 months: entecavir or tenofovir because they have a high resistance barrier
HBV DNA and ALT should be monitored every 3 months
EASL. J Hepatol. 2009;50: Lok AS, et al. Hepatology. 2009;50:

17. What about core antibody positive, surface antibody positive patients?
Presence of HBsAb appears to be protective, especially when titres are >100
However the risk in patients who have “occult” hepatitis B (HBsAb, HBcAb and DNA +ve) may be higher
Provide antiviral therapy for patients receiving rituximab or bone marrow/stem cell transplant even if surface antibody positive

18. April 2011 Stem cell transplant takes place
Post- transplant the patient develops a progressive transaminitis (raised ALT) over the course of the following year.
Ultrasound is normal, no new drugs, no alcohol
He received 2 pools of platelets at the time of his stem cell transplant
No new sexual partners

19. Graph of LFTs vs time
Stem Cell Transplant
Date
ALT

20. What could be responsible for his LFTs? And what tests should be done?

21. Results April 2011 CMV PCR negative
Hepatitis C Antibody negative and PCR negative
Hepatitis B surface antigen negative
Hepatitis A IgM negative and IgG positive
Hepatitis E antibody negative
Liver ultrasound scan and dopler normal
…. What now?

22. The GP calls January 2012…
“I sent a liver screen and the patient is positive for hepatitis B!”
The sample went to a different lab and was tested with a different assay
Rechecked in initial lab:
HBsAg negative on Siemens Centaur XP (the assay used initially)
HBsAg POSITIVE on Abbot Architect (the assay used in the different lab)
Anti-HBc now positive on Siemens Centaur XP – was negative before transplant

23. What has happened? And what to do now?
Differential diagnosis: 1. 2. 3.
Plan: ?

24. Results Wife is hepatitis B surface antigen negative
Donor platelets HBV DNA negative
Further HBV testing on patient’s serum:
HBV PCR positive viral load 106
HBV e antigen positive, e antibody negative

25. Results II
Surface antigen positive on 3 other commercial assays, but negative on Siemens Centaur XP
Mutations in sAg gene: L110I, I126T, F134S and G145R, associated with non-detection on commercial assays
Retrospective testing carried out…

26. Results of retrospective testing using various hepatitis B tests

27. Do You Ever Really Get Rid of HBV?
cccDNA
Immune control—not clearance
“Resolved HBV” a misnomer—still HBV DNA in liver
Werle-Lapostolle B, et al. Gastroenterology. 2004;126:

28. Do You Ever Really Get Rid of HBV?
cccDNA
Immune control—not clearance
“Resolved HBV” a misnomer—still HBV DNA in liver
Werle-Lapostolle B, et al. Gastroenterology. 2004;126:

29. Do You Ever Really Get Rid of HBV?
T cell
cccDNA
T cell
T cell
Immune control—not clearance
“Resolved HBV” a misnomer—still HBV DNA in liver
Werle-Lapostolle B, et al. Gastroenterology. 2004;126:

30. Along Comes Immune Suppression
T cell
HIV
Steroids
Chemotx
cccDNA
T cell
T cell
Immune control can be lost
Immune-mediated liver damage with immune reconstitution
Werle-Lapostolle B, et al. Gastroenterology. 2004;126:

31. Along Comes Immune Suppression
T cell
HIV
Steroids
Chemotx
cccDNA
T cell
T cell
Immune control can be lost
Immune-mediated liver damage with immune reconstitution
Werle-Lapostolle B, et al. Gastroenterology. 2004;126:

32. Hoofnagle JH. Hepatology. 2009;49(5 suppl):S156-S165.
HBV Reactivation
HBeAg+ HBeAg HBeAb+
HBeAg+
Immunotolerance
Immune Clearance
HBV DNA
ALT
5-30 Yrs
Mos-Yrs
Mos-Yrs
Infection
Hoofnagle JH. Hepatology. 2009;49(5 suppl):S156-S165.

33. Hoofnagle JH. Hepatology. 2009;49(5 suppl):S156-S165.
HBV Reactivation
HBeAg+ HBeAg HBeAb+
HBeAg+
Immunotolerance
Immune Suppression
Immune Clearance
HBV DNA
ALT
5-30 Yrs
Mos-Yrs
Mos-Yrs
Infection
Hoofnagle JH. Hepatology. 2009;49(5 suppl):S156-S165.

34. HBV and immunosuppression
All immunosuppressive therapy can cause flares
Risk of reactivation ranges from 20-50% with immunosuppression
Clinically, reactivation can range from asymptomatic illness to liver failure
Flares are associated with 10% mortality

35. Risk Factors for HBV Reactivation
Agent
Use of rituximab and bone marrow/stem cell transplantation are highest risk factors
No direct comparisons between agents
Anti-TNF: more reports with infliximab and adalimumab than etanercept
In general, more immunosuppressive = higher risk
Duration
Increased risk with prolonged therapy
Reports after single doses
HBV profile
HBsAg positive >> anti-HBc alone
HBV DNA positive at baseline

36. Agents Reported to Cause HBV Reactivation
Anti-TNF (infliximab, adalimumab, etanercept)
Other (rituximab, cyclosporine)
Antimetabolite (methotrexate)
Immunomodulatory Therapy
Purine Analogues (azathioprine/6mp)
Steroids (prednisone, budesonide)
Roche B, et al. Liver Int. 2011;31(suppl 1):

37. Rituximab: A Particular Problem
Monoclonal antibody against CD20 (B-cell marker)
Reduces B-cell numbers and antibody levels
Increasingly used as part of CHOP-R, EPOCH-R
Increased risk of HBV reactivation, including HBsAg-negative patients
Reverse seroconversion: reappearance of HBsAg in previously HBsAg-negative patient due to loss of immune control
Yeo W, et al. Hepatology. 2006;43: Papamichalis P, et al. Clin Res Hepatol Gastroenterol. 2012;36:84-93.

38. What if hepatitis B core antibody +ve and DNA -ve?
Monitor LFTs
Very low risk if hepatitis B surface antibody positive and titre >100
If profound immunosuppression such as transplant, rituximab or chemotherapy
Use Lamivudine, Tenofovir or Entecavir

39. Summary of Management *Caveats:
Screen all patients
HBsAg, anti-HBc
HBsAg+
HBsAg-,
anti-HBc+
Positive
HBV DNA
HBV DNA
Positive
Negative
Best strategy uncertain
Monitoring for flare probably
acceptable in some circumstances
Prophylaxis probably sensible in others
e.g. rituximab
Option: HBsAg q mo, HBV DNA q 3 mos
Until 6-12 mos posttherapy
< 2000 IU/mL &
<12 months RX
≥ 2000 IU/mL or
>12 months Rx
LAM x 6-12 mos
posttherapy
ETV/TDF until HBV endpoints
Watch for withdrawal flares
*Caveats:
If concern about monitoring  err on side of treatment
High risk: anti-HBs negative older men  consider up-front treatment

40. What is the diagnosis? And should any treatment be given?
Diagnosis: reactivation of hepatitis B with escape mutant
Anti-HBc was negative prior to transplant due to an assay which did not detect anti-HBc. When it was tested retrospectively with another assay it was weakly positive (see slide 28)
He probably also had low anti-HBc levels at the time due to immunosupression secondary to myeloma.
Treatment should be offered with Tenofovir or Entecavir

41. Outcome… Started tenofovir treatment 300mg OD
After 6 months – ALT normal and HBV viral load undetectable

42. Learning points
All patients should be tested for hepatitis B prior to starting immunosuppression as per EASL and NICE guidelines
Reactivation of hepatitis B is associated with high mortality but is entirely preventable
Lamivudine, Tenofovir and Entecavir are highly effective preventing reactivation in immunosuppressed patients
Hepatitis B surface antigen mutants can cause significant diagnostic difficulty!