Title: Long-Term Outcome of Hepatitis B-Positive Renal Allograft Recipients After Development

Title: Long-Term Outcome of Hepatitis B-Positive Renal Allograft Recipients after Development of Antiviral Treatment

Authors: Kyung Sun Park1, Duck Jong Han2, Jae Berm Park2, Jung Sik Park1, Su-Kil Park1

1 Division of Nephrology, Department of Internal Medicine, University of Ulsan College of Medicine, Asan Medical Center

2 Department of Surgery, University of Ulsan College of Medicine, Asan Medical Center

KSP participated in research design, performed research, data analysis and wrote article; DJH and JBP helped design research and provided urine and blood samples from kidney transplantation candidates; JSP participated in research design; SKP participated in research design, data analysis and project supervisor.

The authors declare no conflict of interest.

There is no funding source.

Word counts of Abstract and Text are 200 and 2692, respectively.

There are 2 Tables and 3 Figures.

Running Title: Long-term outcome of HBV-positive renal recipients

Correspondence: Su-Kil Park, MD, PhD.

Division of Nephrology, Department of Internal Medicine

University of Ulsan college of Medicine, Asan Medical Center

388-1 Poongnap-2 dong, Songpa-gu, Seoul, 138-736, South Korea

Phone: +82-2-3010-3263; Fax: +82-2-3010-6963; E-mail:

ABSTRACT

Background/Aims: Hepatitis B virus (HBV) infection can adversely affect the clinical outcome of kidney transplantation (KT). Short-term efficacy of lamivudine has been demonstrated for chronic hepatitis B in KT recipients (KTR).

Methods: To clarify the long-term impact of antiviral treatment for HBV-positive KTR, we retrospectively reviewed 94 HBV-positive (male 73%) and 282 age-sex matched HBV-negative patients who underwent KT from February 1997 to November 2009, after lamivudine had come into wide use.

Results: Mean follow-up was 75.7 months. Fifty-six patients received antiviral agent for prophylaxis, and other 18 for HBV reactivation. During follow-up, 15 died, with 5 deaths being HBV related. Although the patient survival rate was lower for HBV-positive than HBV-negative KTRs (89% vs. 94% at 5 years, 78% vs. 88% at 10 years, p=0.031), graft survival was comparable (86% vs. 92% at 5 years, 73% vs. 81% at 10 years, p=0.113). Of the 26 hepatitis B e antigen (HBeAg)-positive patients, 14 experienced HBV reactivations, but all survived with stable liver chemistry, except for one who died of hepatocellular carcinoma. Among 57 HBeAg-negative patients, 12 died, whereas the remaining 45 survived without hepatic dysfunction.

Conclusion: Long-term outcomes of HBV-positive KTRs may be favorable after antiviral agents have been introduced.

Keywords: Hepatitis B, Kidney transplantation, Lamivudine

Short summary

We clarified the long-term outcomes of hepatitis B-positive renal allograft recipients who underwent transplantation after antiviral agents had come into wide use in a relatively large cohort with long duration after kidney transplantation. Treatment of hepatitis B-positive renal allograft recipients with antiviral agents still resulted in lower patient survival rates in HBsAg-positive than in HBsAg-negative KT recipients. However, graft survival was not adversely affected by HBV infection.

INTRODUCTION

Hepatitis B virus (HBV) infection has been recognized as a major concern for kidney transplantation (KT). Immunosuppressive therapy after KT enhances viral replication and may lead to liver-related complications, including chronic active hepatitis, cirrhosis and hepatocellular carcinoma (HCC), thus increasing the morbidity and mortality of renal allograft recipients (KTRs) [5, 7, 8, 15, 16, 18, 20]. Liver biopsy has been recommended for HBV-positive patients to identify high risk patients for hepatic decompensation after KT [19]. If the biopsy shows severe hepatitis or cirrhosis, KT is not recommended [11]. Although KT is usually acceptable in patients with mild hepatitis, HBV infection is associated with a significant risk of hepatic failure after KT in these patients.

Lamivudine, the (-) enantiomer of 2’-3’ dideoxy-3’-thiacytidine, is a potent inhibitor of HBV replication [14] and is a safe and effective therapy for chronic hepatitis B (CHB) in KTRs [9, 13, 17, 21]. Preemptive or prophylactic treatment with lamivudine before hepatic dysfunction has shown favorable outcomes in KTRs, including reduction in viral load, improvements in liver chemistry and prevention of permanent histological deterioration [2, 6]. Other agents used to treat CHB after KT include adefovir, a nucleotide analogue of adenosine monophosphate, and entecavir, a carbocyclic analogue of 2’-deoxyguanosine [22, 23]. However, these uncontrolled studies assessed only short-term outcomes after transplantation. The long-term impact of these antiviral agents in HBV-positive KTRs has not been clarified.

We therefore retrospectively investigated the long-term outcomes of HBV-positive KTRs who underwent KT after antiviral agents had come into wide use, focusing on changes in liver chemistry and serological viral markers, such as HBV DNA and hepatitis B e antigen (HBeAg). We also evaluated patient and graft survival compared with HBV-negative KTRs.

PATIENTS AND METHODS

This study was approved by the institutional review board of Asan Medical Center, which has obtained certification from FERCAP (Forum for Ethical Review Committees in Asia and the Western Pacific).

Populations

Since 1997, lamivudine has been used at our center to treat HBV-positive KTRs. The case records of the 1863 patients who underwent KT between February 1997 and November 2009 at Asan Medical Center were reviewed; among these, 102 (5.5%) tested positive for hepatitis B surface antigen (HBsAg) before transplantation. After exclusion of 8 patients who underwent kidney-liver co-transplantation, 94 subjects (69 males, 73%) were included. To compare patient and graft survival, we selected as a control group 282 age- and sex-matched HBV-negative patients who underwent KT after 1997.

Data collection

Data were obtained from the electronic medical recording system of Asan Medical Center. Data collected included baseline demographic characteristics, biochemical liver profile, serological viral markers such as HBsAg, HBeAg, antibody against HBeAg (anti-HBe), HBV DNA and antibody against hepatitis C virus (anti-HCV), induction and maintenance immunosuppression, acute rejection episodes, date and cause of death and date of graft loss. HBsAg, HBeAg and anti-HBe were measured by immunoradiometric assays. HBV DNA titer in serum was measured by hybridization or real-time polymerase chain reaction (PCR). A serum concentration of HBV DNA of > 0.5 pg/mL determined by hybridization or > 50 copies/mL HBV DNA determined by real-time PCR was considered a positive result. Baseline liver histology was obtained for most of the patients immediately before transplantation.

Definitions

Prophylaxis in this study means that the serum alanine aminotransferase (ALT) concentrations is normal when the antiviral drug is prescribed at the time of transplantation, which includes post treatment cases before transplantation. HBV reactivation was defined as an increase of serum ALT two times greater than the upper limit of normal with new appearance of HBV DNA or more than 2-log elevation of HBV DNA titer without other cause of hepatic dysfunction such as alcohol, medication or toxin. Biochemical response (BCR) to antiviral treatment was defined as decreased serum ALT within normal limits (< 40 IU/L). HBeAg seroconversion was defined as the clearance of HBeAg and the appearance anti-HBe. HBeAg reversion was defined as the reappearance of lost HBeAg. Acute rejection was considered when clinically suspected or biopsy-proven. Patient death with functioning kidney was included in graft loss.

Antiviral treatment

Lamivudine (100 mg daily) was used as the initial therapy before adefovir or entecavir was available. Patients who had developed HBV reactivation with suspicion of lamivudine resistance were maintained on lamivudine before the availability of adefovir or entecavir. After these agents were available, these patients were added on adefovir (10 mg daily) or switched to entecavir (1 mg daily), after consultation with a hepatologist. Dosages were adjusted according to allograft function.

Antiviral prophylaxis was at the discretion of each clinician because there were no clear guidelines for prophylaxis of HBV-positive KTRs. Moreover a patient’s finances were crucial because prophylactic antiviral agents for HBV-positive KTRs are not covered by health insurance in Korea.

Lamivudine resistance

The most common mutation that causes lamivudine resistance involves the substitution of methionine in the tyrosine-methionine-aspartate-aspartate (YMDD) motif of the HBV DNA polymerase by valine or isoleucine rtM204V/I, the so-called ‘YMDD mutation’[14]. Direct sequencing of HBV DNA and PCR–restriction fragment length polymorphism (RFLP) analysis were used to detect lamivudine resistance.

Immunosuppression

Patients were administered 500 mg methylprednisolone intravenously during surgery and were then tapered. Maintenance immunosuppression consisted of low-dose prednisolone and cyclosporine or tacrolimus and azathioprine or mycophenolate mofetil. Acute rejection was treated with intravenous methylprednisolone.

Statistical analysis

All statistical analyses were performed with SPSS software (Statistical Package for the Social Science, version 18.0). Quantitative and qualitative variables were compared using Student’s t-test and Chi-square test. Patient and graft survival were analyzed using the Kaplan-Meier method and compared using the log rank test. A p-value < 0.05 was considered statistically significant.

RESULTS

Baseline Characteristics

The mean age of the 94 HBV-positive patients was 40.5 years, and the mean follow-up was 75.7 months ranging from 1 to 162 months (Table 1). None of these patients was positive for anti-HCV. At transplantation, 26 patients were positive and 57 were negative for HBeAg, whereas no information was available for the other 11. In addition, 32 were positive and 51 were negative for HBV DNA, with no information available for the other 11. Mean serum ALT was 19.4 IU/L. Liver biopsies were performed in 71 patients, with histology showing mild inflammation in 67, moderate to severe hepatitis in 2 and cirrhosis in 2. Of the 94 patients, 52 received antiviral prophylaxis, 48 with lamivudine and 4 with entecavir, just before transplantation and 4 were receiving antiviral treatment, 3 with lamivudine and 1 with entecavir, for HBV reactivation that had occurred before transplantation. Both patients with cirrhosis were prescribed antiviral prophylaxis, 1 with lamivudine and 1 with entecavir. Seventy patients received renal grafts from living donors and 24 from deceased donors. Twenty one donors (7 living, 14 deceased) were positive for HBsAg.

Patient and Graft survival

During follow-up, 15 patients died, 5 of HBV-related causes. The 5- and 10-year survival rates of these patients were 89% and 78%, respectively (Fig. 1A). Graft losses developed in 19 patients, with 5- and 10-year graft survival rates of 86% and 73%, respectively (Fig. 1B). The log rank test showed that patient survival rate was significantly lower in HBV-positive patients than in HBV-negative controls (p = 0.031) but that graft survival rates were comparable (p = 0.113).

Subgroup analysis of patient and graft survival according to donor HBV positivity, recipient HBeAg and HBV DNA positivity, and antiviral prophylaxis was performed. The 10-year survival rates of patients who received allograft from HBV-positive and HBV-negative donors were 55% and 83%, respectively (p = 0.985), and the 10-year graft survival rates were 39% and 80% (p = 0.404), respectively. Moreover, there was no significant difference in patient and graft survival rates between subgroups that did and did not receive antiviral prophylaxis. Interestingly, HBeAg-positive and HBV DNA-positive subjects had better outcomes than HBeAg-negative and HBV DNA-negative patients, respectively. This may be resulted from significantly younger age of HBeAg-positive patients (42 vs. 36, p = 0.007). In multivariate analysis, HBeAg and HBV DNA were not significant factor for patient and graft survival. The 5- and 10-year survival rates of each subgroup are summarized in Table 2.

Outcomes of hepatitis B virus infection

Of the 26 HBeAg-positive patients, 14 were receiving antiviral prophylaxis at transplantation (Fig. 2). HBV reactivation occurred in 8 at a mean 46.0 months (range, 7 to 140 months) after transplantation. Four of these had the YMDD mutation and were switched to adefovir. Of these, 3 achieved BCR, at 1, 5 and 10 months after HBV reactivation, respectively, and the fourth showed no further progression of hepatic dysfunction. The remaining 4 patients who were maintained on first-line agents achieved BCRs 3, 6, 6 and 6 months after HBV reactivation, respectively. Eighty months after KT, 1 of these 4 patients acquired HBeAg clearance without seroconversion. Although YMDD mutation was also detected in two other patients who did not experience HBV reactivation, they were maintained on lamivudine. Their liver chemistry remained stable for 70 and 139 months, respectively.

Twelve HBeAg-positive patients did not receive antiviral prophylaxis. Of these, 6 showed HBV reactivation at a mean 61.3 months (range, 9 to 107 months) after KT. After lamivudine treatment, all showed BCR, at a mean 10.7 months, (range 4 to 23 months) after reactivation. Two patients experienced a second episode of HBV reactivation due to the YMDD mutation. Both were switched to adefovir and achieved a second BCR. One patient was diagnosed with HCC 96 months after KT and died 17 months later. During a mean follow-up of 99 months (range, 1 to 162 months) after KT, 25 HBeAg-positive patients survived without hepatic failure or HCC.

Of 57 HBeAg-negative patients, 35 were started on antiviral prophylaxis (Fig. 3). HBV reactivation occurred in 14 of these 35 patients at a mean 39.1 months (range 3 to 102 months) after KT. The YMDD mutation was detected in 11 of these patients, who were switched to adefovir or entecavir. BCRs were observed in 13 patients at a mean 4.9 months (range 1 to 16 months) after reactivation. Three patients died of hepatic failure at 20, 90 and 105 months after KT, respectively. Two patients experienced second episodes of HBV reactivation, at 20 and 100 months after KT, respectively. After management with adefovir or entecavir, their hepatic function remained stable. HCC developed in 2 patients at 4 and 31 months after KT. One died of cytomegalovirus pneumonia, whereas the second remains alive. Another 4 patients died of pulmonary thromboembolism, postoperative complications and pneumonia, at 2, 8, 8 and 68 months after KT, respectively. On this progress, 4 patients experienced HBeAg reversion and 1 patient showed HBsAg clearance.

Of 22 HBeAg-negative patients who did not receive antiviral prophylaxis, 14 experienced HBV reactivation at a mean 10.3 months (range 3 to 46 months) after KT. After treatment with lamivudine or entecavir, all showed BCR, at a mean 6.2 months (range 2 to 28 months) after reactivation. Four experienced a second episode of HBV reactivation, at 14, 40, 41 and 63 months, respectively, after KT. Of these, 1 died 75 months after KT and the other 3 patients have survived without hepatic dysfunction. An additional 3 subjects also died, at 7, 45 and 132 months after KT. Three patients showed HBeAg reversion, at 6, 12 and 32 months, respectively, after KT.

Of 11 patients for whom data about HBeAg were not available, 7 received lamivudine prophylaxis. Of these, 1 patient experienced HBV reactivation 24 months after KT due to a YMDD mutation. This patient showed a BCR after treatment with adefovir for 10 months. Two patients died at 2 and 43 months after KT.