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Hepatitis B associated liver failure following bone marrow transplantation
Journal of Hepatology 1991; 21: 512-511 Printed in Denmark All rights reserved Munksgaard Copenhagen
Coplrigh!0 for
European Association the Study of the Liver 1997
Journal of Hepatology ISSN 01684278
Case Report
Hepatitis B associated liver failure following bone marrow transplantation M. Caselitz, H. Link’, R. Hein2, H. Maschek3, IS. Baker, H. Poliwoda’
and M. I? Manns
Dept. of Gastroenterology and Hepatology, ‘Dept. of Haematology and Oncology, ‘Dept. of Immunology and 3Dept. of Pathology, Medical School of Hannover. Germany
Background: Several cases have been reported showing clearance of HBsAg in chronic hepatitis B carriers due to adoptive transfer of immunity by an hepatitis B immunised hone marrow. Case Report: We report on a 27-year-old man with chronic myelocytic leukemia and asymptomatic chronic hepatitis B who received allogeneic bone marrow transplantation (BMT). The donor was his HLA identical brother with natural immunity against hepatitis B. Before BMT the donor had received an additional dose of recombinant hepatitis B vaccine. Twenty days after BMT alanine aminotransferase levels increased and graft versus host disease of the skin was observed. Elevation of liver enzymes was initially attributed to graft versus host disease of the liver and the patient received high doses of steroids in addition to standard immunosuppression. Alanine aminotransferase levels increased up to a maximum on day 52 while the HBV DNA level peaked on day
38 after BMT. A liver biopsy showed reactivation of hepatitis B and treatment with steroids was tapered down. Although alanine aminotransferase and HBV DNA levels decreased, liver function deteriorated. The patient died 130 days after BMT due to liver failure. Conclusion: This report indicates that disturbance of the balance between HBV replication and immune control after BMT may result in fatal reactivation of hepatitis B. Careful monitoring, including HBV DNA level and early liver biopsy, of patients with chronic hepatitis B undergoing BMT as well as determination of the HBV immune status of the BMT donor is suggested and necessary.
C
hand, fatal hepatocellular injury after BMT in association with hepatitis B has also been reported (5). Reactivation of chronic hepatitis B in patients with immunosuppression secondary to cancer chemotherapy has also been observed in the past (6). These studies demonstrate a sensitive balance between the viral infection and the transplanted immune system in these patients. We report the fatal outcome of hepatitis B reactivation in a patient with chronic hepatitis B following bone marrow transplantation from a hepatitis B immune donor.
hepatitis B can cause serious problems in patients undergoing bone marrow transplantation (BMT). The risk for these patients is a matter of controversy in the literature. Several cases have been reported showing clearance of HBsAg in chronic hepa: titis B carriers due to adoptive transfer by anti-HBs antigen positive bone marrow (l-4). On the other HRONIC
Parts of this work were presented as a poster at the annual meeting of the EASL, Copenhagen, Denmark August 1995.
Key words: Adoptive immune transfer; Bone marrow transplantation; Hepatitis B; Immunosuppression; Liver failure.
Received 26 August 1996; revised 25 April; accepted 25 April 1997
Correspondence:
M. I? Manns, Department of Gastroenterology and Hepatology, Zentrum Innere Medizin und Medizinische Hochschule Hannover, Dermatologie, 30625 Hannover, Germany. Tel: 0049-51 l-5323305. Fax: 0049-511-5324896. 572
Methods Commercial enzyme linked immunosorbent assays (ELISA) from Abbott Diagnostica (Wiesbaden, Germany) and Abbott Laboratories (North Chicago, IL, USA) were used for the detection of HBs Ag (Auszyme
Fatal hepatitis B following BMT
Monoklonal), HBeAg (HBe-EIA), anti-HBs (AUSABEIA), anti-HBe (Anti HBe-EIA), anti-HDV (Delta EIA), anti-HBc (Core-EIA) anti-HCV and anti HBcIgM. The tests were performed according to the recommendations of the manufacturer. Serum levels of hepatitis B virus (HBV) DNA were measured quantitatively by a molecular hybridisation assay using 125 I-labelled HBV-DNA (Abbott hepatitis B viral DNA). HBV-DNA precore/core sequence was amplified by polymerase chain reaction (PCR) with synthetic oligonucleotide primers spanning the precore/core region (7).
Case Report A 27-year-old male, born in Turkey but living in Germany since childhood, was diagnosed as having chronic myelocytic leukemia (CML) in chronic phase in February 1993. From February to September 1993 chemotherapy (busulfan 2-8 mg/day, orally) was administered. The patient was also an asymptomatic HBV carrier exhibiting the following hepatitis B serology: HBsAg positive; HBeAg negative; anti-HBc positive, antiHBcIgM negative, anti-HBe positive, HBV-DNA below 1 pg/ml and HBV-DNA-PCR positive. Hepatitis D superinfection was excluded by anti-HDV negativity The alanine aminotransferase (ALT) levels were slightly increased to 35 U/l (normal range ~22 U/l). A chronic portal hepatitis with slight inflammatory activity and weak HBsAg immunhistochemical staining was found in a liver biopsy. Immunhistochemical staining was negative for HBcAg before bone marrow transplantation (Fig. 1). Other than HBV infection there
TABLE
1
Review of the literature:
successful
Author
Year
No. of patients
Clearance
Chen et al. (1) Locasciulli et al. (8) Reed et al. (4) Lok et al. (3) Ban et al. (2)
1990 1990 1991 1992 1993
2
2 and 7 months 351 days 13 days and 3 years 9 months 14 days
TABLE
was no previous history of jaundice, liver disease or blood transfusion. Tests for antibodies against herpes simplex virus (HSV), cytomegalovirus (CMV) and hepatitis C (HCV) were negative. The PCR for HCV RNA was negative. The test for hepatitis A showed a positive result for IgG and a negative result for IgM. Since his brother was MHC class I identical (A 24,31; B 49,50; Cw 6,7) and a disparity in any MHC class II antigens was excluded by a non-reactive mixed lymphocyte culture (MLC), an allogeneic bone marrow transplantation was performed. The donor had naturally acquired immunity against hepatitis B (HBsAg negative; anti-HBs positive; anti-HBc positive). His anti-HBs level was 548 IU/l, 42 days before BMT. The donor received an additional single dose (1 ml) of recombinant hepatitis B vaccine (Engerix-B; SmithKline Beecham, Munich, Germany) intramuscularly according to Ilan et al. (2). The patient’s laboratory tests before BMT showed a peripheral white blood cell count of 25 000 leucocytes/l, platelets 106/1 and haemoglobin 15 g/dl. Bone marrow aspiration demonstrated increased megakaryopoiesis and granulopoiesis, but no signs of acceleration of the chronic myeloid leukemia. Before BMT the patient was conditioned with busulfan (16 mg/kg) and cyclophosphamide (120 mg/kg). The patient then received a bone marrow transplantation of (4.4~ lO*)/kg bodyweight of nucleated bone marrow cells. A standard immunosuppression with cyclosporin A and methotrexate was started to prevent graft versus host disease (GvHD). Neutrophil leucocytes recovered >500/1 on day 17 after BMT
clearance
1 2 1 1
of HBsAG
after bone marrow of HBsAg
transplantation after BMT
(BMT) Remarks Hepatitis Hepatitis
B serology B serology
of one recipient incomplete of the donor incomplete
Additional vaccination acquired immunity
of the donor
with naturally
2
Review of the literature:
fatal hepatitis
B after bone marrow
transplantation
(BMT)
Author
Year
No. of patients
Onset of the fulminant hepatitis after BMT
Remarks
Pariente et al. (5) Webster et al. (9)
1988 1989
1 1
90 days 56 days
Donor HBsAg negative; Autologous BMT
further
hepatitis
B serology
unclear.
573
M. Caselitz et al.
Fig. 1. Liver biopsy before BMT. Chronic portal hepatitis with slight fibrosis (HE, X250).
mgld
Fig. 2. ALT level, HB V DNA level and steroid therapy after BMT.
BMT
250
800
600 Prednisolone
-10
0
10
20
30
40
50
60
70
80
mg/d
90 100110120130"
days
stimulated by granulocyte-colony stimulating factor (G-CSF). Ten days after BMT a mild graft versus host disease of the skin (grade 1) was observed. The patient received high doses of steroids (maximum 160 mg/day prednisolone), which were gradually tapered down. Meanwhile the ALT levels started to rise and additional graft versus host disease of the liver was suspected. The patient received a second therapy with high doses of steroids starting 30 days after BMT. The ALT level did not change and a third treatment with 574
steroids (maximum 240 mg/day prednisolone) was started 40 days after BMT. Another strong elevation of liver enzymes was observed and a liver biopsy was performed on day 44. Hepatitis B reinfection was shown by histopathology (Fig. 1 vs. Fig. 3). The active lobular hepatitis with necrosis was accompanied by the immunohistochemical proof of HBc Ag in more than 60% of hepatic cells. HBs Ag was positive in more than 80% of the cells by immunohistochemical methods. Retrospectively HBV DNA level increased up to 192
Fatal hepatitis B following BMT
Fig. 3. Liver biopsy after BMT. Acute lobular hepatitis with swelling of hepatocytes, single cell necrosis and lymphocytic injiltration (HE, X400).
Fig. 4. ALT level and therapy after BMT.
800 600 400
n ~
=10
0
10 20 30 40 50 60 70 80 90 10011u1zu1x.J
BMT
pg/ml at day 38 while the peak ALT level of 1100 U/l was seen at day 52 after BMT (Fig. 2). The hepatitis B serology during the post BMT period showed a positive result for HBsAg, anti-HBc and anti-HBe. There were no positive findings for HBeAg and anti-HBs was found to be fluctuating between borderline and weak positivity after 38 days, when HBV DNA peaked at 192 pg/ml. The HBV DNA level decreased thereafter below detection limit by hybridisation. However, PCR for HBV DNA remained positive until the patient’s death. Sequencing of the precore region did not reveal any obvious stop codon mutation, but did show a serine to
days
phenylalanine mutation at position 11. This is within the signal sequence necessary for proper cleavage for the first amino acids of the immature HBeAg. Due to hepatitis reinfection, treatment with steroids was reduced to low dose immunosuppression 48 days after BMT. Prostaglandin E2 (720 mg/day iv.) was used 50 days after BMT (Fig. 4). Although ALT levels decreased to 600 U/l and HBV DNA levels decreased to 37 pg/ml, liver function deteriorated. Therapy with prostaglandin E2 was discontinued after 10 days. Acyclovir was given in doses ranging from 3x400 to 3~ 1025 mgiday on days l-8 and days 145 1 for prophylaxis and treatment of generalised herpes zoster (Fig. 4). 575
M. Caselitz et al.
A continuing decrease of HBV DNA, ALT, albumin and cholinesterase levels as well as prothrombin time, factor V and VII were observed. Ascites developed after 76 days, and hepatic encephalopathy 110 days after BMT. The patient developed sepsis and pulmonary aspergillosis and died 130 days after BMT before an allogenie donor liver graft became available for treatment of liver failure. Permission for a post-mortem examination was denied by the patient’s relatives.
Discussion The patient described above with asymptomatic chronic hepatitis B developed fulminant hepatitis B after allogeneic bone marrow transplantation (BMT) from a hepatitis B immune donor (anti-HBs positive, anti-HBc positive). Several cases have been reported in recent years in which adoptive transfer of immunity led to clearance of HBsAg (Table 1). Ilan et al. (2) described a patient with chronic hepatitis B who cleared HBV infection within 4 weeks after an allogeneic BMT from a HBsAg negative, anti-HBs positive and anti-HBc positive donor. This donor had also received an additional active vaccination 45 days before BMT. Only a mild elevation of ALT was observed in the recipient. These data suggest that the additional active vaccination of the donor was not likely to be the reason for the liver failure in our patient. Reed et al. (4) retrospectively identified three HBsAg positive patients who received bone marrow of donors with naturally acquired immunity against hepatitis B. One patient became anti-HBs positive and HBsAg negative 13 days after BMT. Two patients remained HBsAg positive, but became negative for HBeAg and positive for anti-HBe about 75 days after transplantation. In a fourth patient during the long-term followup, a clearance of HBsAg was observed 3 years, and confirmed 7 years, post BMT. Unfortunately, the donor’s hepatitis serology is unknown in this patient. No evidence of fulminant hepatitis was found in these HBsAg positive patients. Chen et al. (1) also studied HBV markers in two HBsAg positive patients. Both cleared HBsAg and developed a persistent level of anti-HBs 7 and 2 months after BMT. Locasciulli et al. (8) found clearance of HBsAg in one of four patients with chronic hepatitis B before BMT. The donor’s hepatitis serology is unknown. Another successful clearance of HBsAg after BMT was reported by Lok et al. (3). None of the reported patients developed severe hepatic injury. In contrast, fulminant hepatitis after allogeneic bone marrow transplantation is reported by Pariente et al. (Table 2) (5). The authors observed a reactivation of 576
chronic hepatitis B 88 days after a successful BMT. Unfortunately, it remains unclear whether the donor had immunity against hepatitis B. An increase of HBsAg concentration, conversion from anti-HBe to HBeAg, and the appearance of anti-HBc-IgM and HBV DNA in the serum were observed. These results suggest an acute exacerbation of HBV replication. In contrast to these findings we found increased levels of HBV DNA in the serum of the presented patient, but did not observe a change from anti-HBe to HBeAg and no appearance of anti-HBc-IgM in our patient. The literature also contains some other reports of fatal hepatitis after BMT. Webster et al. (9) reported a case of fatal hepatitis B after autologous BMT. Their Turkish patient, suffering from acute lymphocytic leukemia, was positive for anti-HBs and anti-HBc and negative for HBsAg before BMT. HBs antigen was first detected 5 weeks after BMT and the patient died 10 weeks later due to the fulminant course of activated hepatitis B. The fatal hepatitis B in this patient seemed to be the result of immunosuppression associated with leukemia and the autologous BMT, since the patient had natural immunity to hepatitis B, most probably acquired many years ago in childhood, before BMT (Table 2). In contrast to our case, this patient underwent autologous BMT and therefore it was not necessary to differentiate between reactivated hepatitis B and GvHD before treatment. Two cases of a fulminant hepatitis C (HCV) post allogenic BMT have been reported by Kanamori et al. (10). The fatal reactivation of HCV developed within 5 months after BMT and was coincident with the withdrawal of the immunosuppression with cyclosporin A and methotrexate. In contrast to our case, these patients did not receive steroids before elevation of liver enzymes. As described below, the reactivation of the hepatitis B in our patient was related to the therapy with corticosteroids. However, these cases have in common that the imbalance between immunosuppression and reconstitution of the transplanted allogenic immune system may have led to activation of the immune system and the induction of hepatocellular injury in hepatitis C and B. Similar courses of ALT and HBV DNA polymerase levels (Fig. 2) have also been reported in chronic active hepatitis B during immunosuppressive therapy (11,12), which increases HBV DNA replication. The decrease of HBV DNA level and the rising ALT levels following the withdrawal of steroids may be explained by reactivation of a functional immune system. These findings agree with an observed increased number of peripheral blood lymphocytes. These data may confirm that high doses of cortico-
Fatal hepatitis B following BMT
steroids and their withdrawal play an important role in the acute exacerbation of hepatitis B after BMT. Further investigations are required to understand the role of corticosteroids and to develop alternative therapeutic strategies for patients with chronic hepatitis B and GvH disease following BMT. Additionally, exacerbation of the clinical course due to adoptive transfer of immunity might have played a role in this patient, because anti-HBs occurred 38 days after BMT and remained fluctuating until death. On the other hand, the patient received immunoglobulins (gammaglobulin; 40 g/week) because the recipient was cytomegalovirus seropositive and immunoglobulins have been proven to reduce the incidence and severity of acute GvH disease, thereby reducing the death rate of the recipients (13). Anti-HBc IgM was not detectable, either. However, there is no proof of adoptive immune transfer in this patient. In spite of high viral replication, no HBeAg was found. On the nucleotide level, no mutation of the precore region could be identified which produced a stop codon or disabled the initiation of translation. We did observe a mutation within the signal sequence tracing the immature HBeAg to the endoplasmatic reticulum, where the first 19 amino acids will be cleaved. Unfortunately, we do not have available any material before BMT for sequence analysis. It remains unclear whether this mutant especially alters the clinical course. The possibility of a co-infection of the liver with hepatitis B and herpes simplex virus after BMT was excluded by a negative result for in situ hybridisation of herpes simplex virus in the liver biopsy. Because of the reported beneficial effect of oral and intravenous prostaglandin E2 (PG E2) in patients with fulminant and subfulminant viral hepatitis we started to treat the patient with intravenous PG E2 (14). Unfortunately, liver function did not improve in our patient and treatment was discontinued because of severe side effects after 10 days.
Conclusions High dose immunosuppression and later reestablishment of the donor’s immune systems in BMT recipients can influence the course of chronic hepatitis B. The clearance of HBsAg in patients suffering from chronic hepatitis B by BMT is well documented (see above). This case may demonstrate the potential hazards with HBsAg positive bone marrow recipients and the difficult differential diagnosis between GvHD and HBV reinfection of the liver. The choice of treatment may be decisive for the clinical outcome of such patients. Care-
ful monitoring, including HBV DNA level and early liver biopsy, of patients with chronic hepatitis B undergoing BMT as well as determination of the HBV immune status of the BMT donor is suggested and necessary.
References 1. Chen PM, Fan S, Liu CJ, Hsieh RK, Liu JH, Chuang MW, et al. Changing of hepatitis B virus markers in patients with bone marrow transplantation. Transplantation 1990; 49: 708-13. 2. Ban Y, Nagler A, Adler R, Tur-Kaspa R, Slavin S, Shouval D. Ablation of persistent hepatitis B by bone marrow transplantation from a hepatitis B-immune donor. Gastroenterology 1993; 104: 1818-21. 3. Lok AH. Liang RH, Chung HT. Recovery of chronic hepatitis B (letter). Ann Intern Med 1992; 116: 957-8. 4. Reed EC, Myerson D, Corey L, Meyers JD. Allogeneic marrow transplantation in patients positive for hepatitis B surface antigen. Blood 1991; 77: 195-200. 5. Pariente EA, Goudeau A, Dubois F, Degott C, Gluckman E, Devergie A, et al. Fulminant hepatitis due to reactivation of chronic hepatitis B virus infection after allogeneic bone marrow transplantation. Dig Dis Sci 1988; 33: 1185-91. 6. Hoofnagle JH, Dusheiko GM, Schafer DF, Jones A, Micetich K, Young RC, et al. Reactivation of chronic hepatitis B virus infection by cancer chemotherapy. Ann Intern Med 1982; 96: 447-9. 7. Tillmann HL, Trautwein C, Waker D, Mitichitaka K, Kubicka S, Bilker KHW, et al. Clinical relevance of mutations in the precore Genome of the hepatitis B virus. Gut 1995; 37: 568-73. 8. Locasciulli A, Bacigalupo A, Van-Lint MT, Chemello L, Pontisso P Occhini D, et al. Hepatitis B virus (HBV) infection and liver disease after allogeneic bone marrow transplantation: a report of 30 cases. Bone Marrow Transplant 1990; 6: 25-9. 9. Webster A, Brenner MK, Prentice HG, Griffiths PD. Fatal hepatitis B reactivation after autologous bone marrow transplantion. Bone Marrow Transplant 1989; 4(2): 207-8. 10. Kanamori H, Fukawa H, Maruta A, Harano H, Kodama F, Matsuzaki M, et al. Case report: fulminant hepatitis C viral infection after allogenic bone marrow transplantation. Am J Med Sci 1992; 303(2): 109-l 1. 11. Rakela J, Redecker AG, Weliky B. Effect of short term prednisone therapy on aminotransferase levels and hepatitis B virus markers in chronic type B hepatitis. Gastroenterology 1983; 84: 956-60. 12. Scullard GH, Smith CI, Merigan TC, Robinson WS, Gregory PB. Effects of immunosuppressive therapy on viral markers in chronic active hepatitis. Gastroenterology 1981; 81: 987-91. 13. Sullivan KM, Kopecky KJ, Jocom J, Fisher L, Buckner CD, Meyers JD, et al. Immunomodulatory and antimicrobial efficacy of intravenous immunoglobulin in bone marrow transplantation. N Engl J Med 1990; 323: 705-12. 14. Sinclair SB, Greig PD, Blendis LM, Abecassis EA, Roberts EA, Philipps MJ, et al. Biochemical and clinical response of fulminant viral hepatitis to administration of prostaglandin E: a preliminary report. J Clin Invest 1989; 84: 1063-9.
577
Coplrigh!0 for
European Association the Study of the Liver 1997
Journal of Hepatology ISSN 01684278
Case Report
Hepatitis B associated liver failure following bone marrow transplantation M. Caselitz, H. Link’, R. Hein2, H. Maschek3, IS. Baker, H. Poliwoda’
and M. I? Manns
Dept. of Gastroenterology and Hepatology, ‘Dept. of Haematology and Oncology, ‘Dept. of Immunology and 3Dept. of Pathology, Medical School of Hannover. Germany
Background: Several cases have been reported showing clearance of HBsAg in chronic hepatitis B carriers due to adoptive transfer of immunity by an hepatitis B immunised hone marrow. Case Report: We report on a 27-year-old man with chronic myelocytic leukemia and asymptomatic chronic hepatitis B who received allogeneic bone marrow transplantation (BMT). The donor was his HLA identical brother with natural immunity against hepatitis B. Before BMT the donor had received an additional dose of recombinant hepatitis B vaccine. Twenty days after BMT alanine aminotransferase levels increased and graft versus host disease of the skin was observed. Elevation of liver enzymes was initially attributed to graft versus host disease of the liver and the patient received high doses of steroids in addition to standard immunosuppression. Alanine aminotransferase levels increased up to a maximum on day 52 while the HBV DNA level peaked on day
38 after BMT. A liver biopsy showed reactivation of hepatitis B and treatment with steroids was tapered down. Although alanine aminotransferase and HBV DNA levels decreased, liver function deteriorated. The patient died 130 days after BMT due to liver failure. Conclusion: This report indicates that disturbance of the balance between HBV replication and immune control after BMT may result in fatal reactivation of hepatitis B. Careful monitoring, including HBV DNA level and early liver biopsy, of patients with chronic hepatitis B undergoing BMT as well as determination of the HBV immune status of the BMT donor is suggested and necessary.
C
hand, fatal hepatocellular injury after BMT in association with hepatitis B has also been reported (5). Reactivation of chronic hepatitis B in patients with immunosuppression secondary to cancer chemotherapy has also been observed in the past (6). These studies demonstrate a sensitive balance between the viral infection and the transplanted immune system in these patients. We report the fatal outcome of hepatitis B reactivation in a patient with chronic hepatitis B following bone marrow transplantation from a hepatitis B immune donor.
hepatitis B can cause serious problems in patients undergoing bone marrow transplantation (BMT). The risk for these patients is a matter of controversy in the literature. Several cases have been reported showing clearance of HBsAg in chronic hepa: titis B carriers due to adoptive transfer by anti-HBs antigen positive bone marrow (l-4). On the other HRONIC
Parts of this work were presented as a poster at the annual meeting of the EASL, Copenhagen, Denmark August 1995.
Key words: Adoptive immune transfer; Bone marrow transplantation; Hepatitis B; Immunosuppression; Liver failure.
Received 26 August 1996; revised 25 April; accepted 25 April 1997
Correspondence:
M. I? Manns, Department of Gastroenterology and Hepatology, Zentrum Innere Medizin und Medizinische Hochschule Hannover, Dermatologie, 30625 Hannover, Germany. Tel: 0049-51 l-5323305. Fax: 0049-511-5324896. 572
Methods Commercial enzyme linked immunosorbent assays (ELISA) from Abbott Diagnostica (Wiesbaden, Germany) and Abbott Laboratories (North Chicago, IL, USA) were used for the detection of HBs Ag (Auszyme
Fatal hepatitis B following BMT
Monoklonal), HBeAg (HBe-EIA), anti-HBs (AUSABEIA), anti-HBe (Anti HBe-EIA), anti-HDV (Delta EIA), anti-HBc (Core-EIA) anti-HCV and anti HBcIgM. The tests were performed according to the recommendations of the manufacturer. Serum levels of hepatitis B virus (HBV) DNA were measured quantitatively by a molecular hybridisation assay using 125 I-labelled HBV-DNA (Abbott hepatitis B viral DNA). HBV-DNA precore/core sequence was amplified by polymerase chain reaction (PCR) with synthetic oligonucleotide primers spanning the precore/core region (7).
Case Report A 27-year-old male, born in Turkey but living in Germany since childhood, was diagnosed as having chronic myelocytic leukemia (CML) in chronic phase in February 1993. From February to September 1993 chemotherapy (busulfan 2-8 mg/day, orally) was administered. The patient was also an asymptomatic HBV carrier exhibiting the following hepatitis B serology: HBsAg positive; HBeAg negative; anti-HBc positive, antiHBcIgM negative, anti-HBe positive, HBV-DNA below 1 pg/ml and HBV-DNA-PCR positive. Hepatitis D superinfection was excluded by anti-HDV negativity The alanine aminotransferase (ALT) levels were slightly increased to 35 U/l (normal range ~22 U/l). A chronic portal hepatitis with slight inflammatory activity and weak HBsAg immunhistochemical staining was found in a liver biopsy. Immunhistochemical staining was negative for HBcAg before bone marrow transplantation (Fig. 1). Other than HBV infection there
TABLE
1
Review of the literature:
successful
Author
Year
No. of patients
Clearance
Chen et al. (1) Locasciulli et al. (8) Reed et al. (4) Lok et al. (3) Ban et al. (2)
1990 1990 1991 1992 1993
2
2 and 7 months 351 days 13 days and 3 years 9 months 14 days
TABLE
was no previous history of jaundice, liver disease or blood transfusion. Tests for antibodies against herpes simplex virus (HSV), cytomegalovirus (CMV) and hepatitis C (HCV) were negative. The PCR for HCV RNA was negative. The test for hepatitis A showed a positive result for IgG and a negative result for IgM. Since his brother was MHC class I identical (A 24,31; B 49,50; Cw 6,7) and a disparity in any MHC class II antigens was excluded by a non-reactive mixed lymphocyte culture (MLC), an allogeneic bone marrow transplantation was performed. The donor had naturally acquired immunity against hepatitis B (HBsAg negative; anti-HBs positive; anti-HBc positive). His anti-HBs level was 548 IU/l, 42 days before BMT. The donor received an additional single dose (1 ml) of recombinant hepatitis B vaccine (Engerix-B; SmithKline Beecham, Munich, Germany) intramuscularly according to Ilan et al. (2). The patient’s laboratory tests before BMT showed a peripheral white blood cell count of 25 000 leucocytes/l, platelets 106/1 and haemoglobin 15 g/dl. Bone marrow aspiration demonstrated increased megakaryopoiesis and granulopoiesis, but no signs of acceleration of the chronic myeloid leukemia. Before BMT the patient was conditioned with busulfan (16 mg/kg) and cyclophosphamide (120 mg/kg). The patient then received a bone marrow transplantation of (4.4~ lO*)/kg bodyweight of nucleated bone marrow cells. A standard immunosuppression with cyclosporin A and methotrexate was started to prevent graft versus host disease (GvHD). Neutrophil leucocytes recovered >500/1 on day 17 after BMT
clearance
1 2 1 1
of HBsAG
after bone marrow of HBsAg
transplantation after BMT
(BMT) Remarks Hepatitis Hepatitis
B serology B serology
of one recipient incomplete of the donor incomplete
Additional vaccination acquired immunity
of the donor
with naturally
2
Review of the literature:
fatal hepatitis
B after bone marrow
transplantation
(BMT)
Author
Year
No. of patients
Onset of the fulminant hepatitis after BMT
Remarks
Pariente et al. (5) Webster et al. (9)
1988 1989
1 1
90 days 56 days
Donor HBsAg negative; Autologous BMT
further
hepatitis
B serology
unclear.
573
M. Caselitz et al.
Fig. 1. Liver biopsy before BMT. Chronic portal hepatitis with slight fibrosis (HE, X250).
mgld
Fig. 2. ALT level, HB V DNA level and steroid therapy after BMT.
BMT
250
800
600 Prednisolone
-10
0
10
20
30
40
50
60
70
80
mg/d
90 100110120130"
days
stimulated by granulocyte-colony stimulating factor (G-CSF). Ten days after BMT a mild graft versus host disease of the skin (grade 1) was observed. The patient received high doses of steroids (maximum 160 mg/day prednisolone), which were gradually tapered down. Meanwhile the ALT levels started to rise and additional graft versus host disease of the liver was suspected. The patient received a second therapy with high doses of steroids starting 30 days after BMT. The ALT level did not change and a third treatment with 574
steroids (maximum 240 mg/day prednisolone) was started 40 days after BMT. Another strong elevation of liver enzymes was observed and a liver biopsy was performed on day 44. Hepatitis B reinfection was shown by histopathology (Fig. 1 vs. Fig. 3). The active lobular hepatitis with necrosis was accompanied by the immunohistochemical proof of HBc Ag in more than 60% of hepatic cells. HBs Ag was positive in more than 80% of the cells by immunohistochemical methods. Retrospectively HBV DNA level increased up to 192
Fatal hepatitis B following BMT
Fig. 3. Liver biopsy after BMT. Acute lobular hepatitis with swelling of hepatocytes, single cell necrosis and lymphocytic injiltration (HE, X400).
Fig. 4. ALT level and therapy after BMT.
800 600 400
n ~
=10
0
10 20 30 40 50 60 70 80 90 10011u1zu1x.J
BMT
pg/ml at day 38 while the peak ALT level of 1100 U/l was seen at day 52 after BMT (Fig. 2). The hepatitis B serology during the post BMT period showed a positive result for HBsAg, anti-HBc and anti-HBe. There were no positive findings for HBeAg and anti-HBs was found to be fluctuating between borderline and weak positivity after 38 days, when HBV DNA peaked at 192 pg/ml. The HBV DNA level decreased thereafter below detection limit by hybridisation. However, PCR for HBV DNA remained positive until the patient’s death. Sequencing of the precore region did not reveal any obvious stop codon mutation, but did show a serine to
days
phenylalanine mutation at position 11. This is within the signal sequence necessary for proper cleavage for the first amino acids of the immature HBeAg. Due to hepatitis reinfection, treatment with steroids was reduced to low dose immunosuppression 48 days after BMT. Prostaglandin E2 (720 mg/day iv.) was used 50 days after BMT (Fig. 4). Although ALT levels decreased to 600 U/l and HBV DNA levels decreased to 37 pg/ml, liver function deteriorated. Therapy with prostaglandin E2 was discontinued after 10 days. Acyclovir was given in doses ranging from 3x400 to 3~ 1025 mgiday on days l-8 and days 145 1 for prophylaxis and treatment of generalised herpes zoster (Fig. 4). 575
M. Caselitz et al.
A continuing decrease of HBV DNA, ALT, albumin and cholinesterase levels as well as prothrombin time, factor V and VII were observed. Ascites developed after 76 days, and hepatic encephalopathy 110 days after BMT. The patient developed sepsis and pulmonary aspergillosis and died 130 days after BMT before an allogenie donor liver graft became available for treatment of liver failure. Permission for a post-mortem examination was denied by the patient’s relatives.
Discussion The patient described above with asymptomatic chronic hepatitis B developed fulminant hepatitis B after allogeneic bone marrow transplantation (BMT) from a hepatitis B immune donor (anti-HBs positive, anti-HBc positive). Several cases have been reported in recent years in which adoptive transfer of immunity led to clearance of HBsAg (Table 1). Ilan et al. (2) described a patient with chronic hepatitis B who cleared HBV infection within 4 weeks after an allogeneic BMT from a HBsAg negative, anti-HBs positive and anti-HBc positive donor. This donor had also received an additional active vaccination 45 days before BMT. Only a mild elevation of ALT was observed in the recipient. These data suggest that the additional active vaccination of the donor was not likely to be the reason for the liver failure in our patient. Reed et al. (4) retrospectively identified three HBsAg positive patients who received bone marrow of donors with naturally acquired immunity against hepatitis B. One patient became anti-HBs positive and HBsAg negative 13 days after BMT. Two patients remained HBsAg positive, but became negative for HBeAg and positive for anti-HBe about 75 days after transplantation. In a fourth patient during the long-term followup, a clearance of HBsAg was observed 3 years, and confirmed 7 years, post BMT. Unfortunately, the donor’s hepatitis serology is unknown in this patient. No evidence of fulminant hepatitis was found in these HBsAg positive patients. Chen et al. (1) also studied HBV markers in two HBsAg positive patients. Both cleared HBsAg and developed a persistent level of anti-HBs 7 and 2 months after BMT. Locasciulli et al. (8) found clearance of HBsAg in one of four patients with chronic hepatitis B before BMT. The donor’s hepatitis serology is unknown. Another successful clearance of HBsAg after BMT was reported by Lok et al. (3). None of the reported patients developed severe hepatic injury. In contrast, fulminant hepatitis after allogeneic bone marrow transplantation is reported by Pariente et al. (Table 2) (5). The authors observed a reactivation of 576
chronic hepatitis B 88 days after a successful BMT. Unfortunately, it remains unclear whether the donor had immunity against hepatitis B. An increase of HBsAg concentration, conversion from anti-HBe to HBeAg, and the appearance of anti-HBc-IgM and HBV DNA in the serum were observed. These results suggest an acute exacerbation of HBV replication. In contrast to these findings we found increased levels of HBV DNA in the serum of the presented patient, but did not observe a change from anti-HBe to HBeAg and no appearance of anti-HBc-IgM in our patient. The literature also contains some other reports of fatal hepatitis after BMT. Webster et al. (9) reported a case of fatal hepatitis B after autologous BMT. Their Turkish patient, suffering from acute lymphocytic leukemia, was positive for anti-HBs and anti-HBc and negative for HBsAg before BMT. HBs antigen was first detected 5 weeks after BMT and the patient died 10 weeks later due to the fulminant course of activated hepatitis B. The fatal hepatitis B in this patient seemed to be the result of immunosuppression associated with leukemia and the autologous BMT, since the patient had natural immunity to hepatitis B, most probably acquired many years ago in childhood, before BMT (Table 2). In contrast to our case, this patient underwent autologous BMT and therefore it was not necessary to differentiate between reactivated hepatitis B and GvHD before treatment. Two cases of a fulminant hepatitis C (HCV) post allogenic BMT have been reported by Kanamori et al. (10). The fatal reactivation of HCV developed within 5 months after BMT and was coincident with the withdrawal of the immunosuppression with cyclosporin A and methotrexate. In contrast to our case, these patients did not receive steroids before elevation of liver enzymes. As described below, the reactivation of the hepatitis B in our patient was related to the therapy with corticosteroids. However, these cases have in common that the imbalance between immunosuppression and reconstitution of the transplanted allogenic immune system may have led to activation of the immune system and the induction of hepatocellular injury in hepatitis C and B. Similar courses of ALT and HBV DNA polymerase levels (Fig. 2) have also been reported in chronic active hepatitis B during immunosuppressive therapy (11,12), which increases HBV DNA replication. The decrease of HBV DNA level and the rising ALT levels following the withdrawal of steroids may be explained by reactivation of a functional immune system. These findings agree with an observed increased number of peripheral blood lymphocytes. These data may confirm that high doses of cortico-
Fatal hepatitis B following BMT
steroids and their withdrawal play an important role in the acute exacerbation of hepatitis B after BMT. Further investigations are required to understand the role of corticosteroids and to develop alternative therapeutic strategies for patients with chronic hepatitis B and GvH disease following BMT. Additionally, exacerbation of the clinical course due to adoptive transfer of immunity might have played a role in this patient, because anti-HBs occurred 38 days after BMT and remained fluctuating until death. On the other hand, the patient received immunoglobulins (gammaglobulin; 40 g/week) because the recipient was cytomegalovirus seropositive and immunoglobulins have been proven to reduce the incidence and severity of acute GvH disease, thereby reducing the death rate of the recipients (13). Anti-HBc IgM was not detectable, either. However, there is no proof of adoptive immune transfer in this patient. In spite of high viral replication, no HBeAg was found. On the nucleotide level, no mutation of the precore region could be identified which produced a stop codon or disabled the initiation of translation. We did observe a mutation within the signal sequence tracing the immature HBeAg to the endoplasmatic reticulum, where the first 19 amino acids will be cleaved. Unfortunately, we do not have available any material before BMT for sequence analysis. It remains unclear whether this mutant especially alters the clinical course. The possibility of a co-infection of the liver with hepatitis B and herpes simplex virus after BMT was excluded by a negative result for in situ hybridisation of herpes simplex virus in the liver biopsy. Because of the reported beneficial effect of oral and intravenous prostaglandin E2 (PG E2) in patients with fulminant and subfulminant viral hepatitis we started to treat the patient with intravenous PG E2 (14). Unfortunately, liver function did not improve in our patient and treatment was discontinued because of severe side effects after 10 days.
Conclusions High dose immunosuppression and later reestablishment of the donor’s immune systems in BMT recipients can influence the course of chronic hepatitis B. The clearance of HBsAg in patients suffering from chronic hepatitis B by BMT is well documented (see above). This case may demonstrate the potential hazards with HBsAg positive bone marrow recipients and the difficult differential diagnosis between GvHD and HBV reinfection of the liver. The choice of treatment may be decisive for the clinical outcome of such patients. Care-
ful monitoring, including HBV DNA level and early liver biopsy, of patients with chronic hepatitis B undergoing BMT as well as determination of the HBV immune status of the BMT donor is suggested and necessary.
References 1. Chen PM, Fan S, Liu CJ, Hsieh RK, Liu JH, Chuang MW, et al. Changing of hepatitis B virus markers in patients with bone marrow transplantation. Transplantation 1990; 49: 708-13. 2. Ban Y, Nagler A, Adler R, Tur-Kaspa R, Slavin S, Shouval D. Ablation of persistent hepatitis B by bone marrow transplantation from a hepatitis B-immune donor. Gastroenterology 1993; 104: 1818-21. 3. Lok AH. Liang RH, Chung HT. Recovery of chronic hepatitis B (letter). Ann Intern Med 1992; 116: 957-8. 4. Reed EC, Myerson D, Corey L, Meyers JD. Allogeneic marrow transplantation in patients positive for hepatitis B surface antigen. Blood 1991; 77: 195-200. 5. Pariente EA, Goudeau A, Dubois F, Degott C, Gluckman E, Devergie A, et al. Fulminant hepatitis due to reactivation of chronic hepatitis B virus infection after allogeneic bone marrow transplantation. Dig Dis Sci 1988; 33: 1185-91. 6. Hoofnagle JH, Dusheiko GM, Schafer DF, Jones A, Micetich K, Young RC, et al. Reactivation of chronic hepatitis B virus infection by cancer chemotherapy. Ann Intern Med 1982; 96: 447-9. 7. Tillmann HL, Trautwein C, Waker D, Mitichitaka K, Kubicka S, Bilker KHW, et al. Clinical relevance of mutations in the precore Genome of the hepatitis B virus. Gut 1995; 37: 568-73. 8. Locasciulli A, Bacigalupo A, Van-Lint MT, Chemello L, Pontisso P Occhini D, et al. Hepatitis B virus (HBV) infection and liver disease after allogeneic bone marrow transplantation: a report of 30 cases. Bone Marrow Transplant 1990; 6: 25-9. 9. Webster A, Brenner MK, Prentice HG, Griffiths PD. Fatal hepatitis B reactivation after autologous bone marrow transplantion. Bone Marrow Transplant 1989; 4(2): 207-8. 10. Kanamori H, Fukawa H, Maruta A, Harano H, Kodama F, Matsuzaki M, et al. Case report: fulminant hepatitis C viral infection after allogenic bone marrow transplantation. Am J Med Sci 1992; 303(2): 109-l 1. 11. Rakela J, Redecker AG, Weliky B. Effect of short term prednisone therapy on aminotransferase levels and hepatitis B virus markers in chronic type B hepatitis. Gastroenterology 1983; 84: 956-60. 12. Scullard GH, Smith CI, Merigan TC, Robinson WS, Gregory PB. Effects of immunosuppressive therapy on viral markers in chronic active hepatitis. Gastroenterology 1981; 81: 987-91. 13. Sullivan KM, Kopecky KJ, Jocom J, Fisher L, Buckner CD, Meyers JD, et al. Immunomodulatory and antimicrobial efficacy of intravenous immunoglobulin in bone marrow transplantation. N Engl J Med 1990; 323: 705-12. 14. Sinclair SB, Greig PD, Blendis LM, Abecassis EA, Roberts EA, Philipps MJ, et al. Biochemical and clinical response of fulminant viral hepatitis to administration of prostaglandin E: a preliminary report. J Clin Invest 1989; 84: 1063-9.
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