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Keywords:

  • Hepatitis B vaccine;
  • immunogenicity;
  • intradermal;
  • transplant

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References

We investigated intradermal hepatitis B (HBV) vaccination in 24 renal transplant recipients who failed to develop hepatitis B surface antibody (anti-HBs) with intramuscular (i.m.) vaccination. All patients received recombinant HBV vaccine 5 μg intradermally every 2 weeks for 8 doses. Nine patients developed protective levels of anti-HBs (> 10 miu/mL) and two patients developed low levels of anti-HBs (4–6 miu/mL), giving an overall initial response rate of 45.8%. A booster of 40 μg was administered intramuscularly after 1 year. All initial responders developed an anti-HBs response (322.6 ± 92.0 miu/mL). In addition, four patients who did not respond initially to the intradermal vaccination seroconverted after the booster. Responders (62.5%) and nonresponders had comparable age, gender, immunosuppressive medications, and duration of transplant. In conclusion, renal transplant patients who fail to respond to intramuscular HBV vaccination may benefit from intradermal vaccination followed by an intramuscular booster.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References

Hepatitis B infection is associated with significant morbidity and mortality in renal transplant patients (1,2). If hepatitis B infection is contracted after renal transplantation, most unprotected patients will become chronic carriers (3). Efforts have been made to vaccinate seronegative transplant recipients with hepatitis B vaccine. However, the seroconversion rate with the conventional intramuscular (i.m.) regime is as low as 9%. Even with the enhanced regimen of four doses of double-strength intramuscular immunization, the seroconversion rate is still around 32–36% (4,5). Intradermal (i.d.) hepatitis B vaccination has been shown to be an effective and cost-saving means of inducing hepatitis B immunity (6,7). It has also been reported that intradermal vaccination is effective in seroconverting healthy individuals (8) and chronic renal failure patients (9,10) who do not respond to conventional intramuscular vaccination. In our study, we report for the first time, that intradermal hepatitis B immunization can induce immunity in transplant recipients who fail to response to high-dose intramuscular vaccine.

Materials and Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References

Twenty-four renal transplant recipients who failed to develop hepatitis B surface antibody (anti-HBs) after four or more doses of double-strength hepatitis B vaccines intramuscularly post transplant were recruited. All of them had received 4 or more doses (range 4–8 doses) of double-strength hepatitis B vaccine (40 μg of recombinant HBV vaccine) at a mean time of 44.7 ± 7.4 months post transplant, and anti-HBs checked within 4 weeks post vaccination were negative. One of the patients had also received a course of i.m. vaccination before transplant and a second course of i.m. vaccination post transplant but still failed to respond. All of the patients had no detectable hepatitis B surface antigen, hepatitis B surface antibody and hepatitis B core antibody before entry into the study.

Patients were immunized intradermally with 5 μg recombinant hepatitis B vaccine (Engerix-B, SmithKline Beecham Biologicals, Belgium) in the forearm every 2 weeks for a total of eight doses. Assays for hepatitis B surface antigen and antibody were performed by enzyme immunoassay using the Abbot Axsym autoanalyzer (AbbottGmbH, Germany). Hepatitis B core antibody was detected using the VIDAS immunoassay (VIDAS bioMeureux, France). An anti-HBs level greater than 10 miu/mL was considered protective. Initial responders were defined as those who had developed anti-HBs (= 4 min/mL) 2 weeks after the last dose of intradermal vaccine. A booster dose of hepatitis B vaccine 40 μg was given intramuscularly to the initial responders when their antibody level fell below 10 miu/mL at the 12th month.

Immunosuppressive medications included prednisolone in all patients, cyclosporin A in 22 patients, and azathioprine in 16 patients, in the form of either triple (14 patients) or double (10 patients) therapy. Immunosuppression remained unchanged during the study.

Statistical analysis was performed by the Statistical Package for Social Science (SPSS). Differences between groups were analyzed using the Mann–Whitney test. Contingency tables were analyzed using the Fisher exact test or Chi-square test as appropriate. All values were expressed as mean ± standard error and p < 0.05 was considered statistically significant.

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References

After eight doses of intradermal vaccination, nine patients (including the one who failed to respond to both courses of i.m. vaccination pre- and post-transplant) developed protective levels of anti-HBs (> 10 miu/mL), two patients developed low levels of anti-HBs (4–6 miu/mL) and 13 patients did not develop detectable antibody. For the initial responders, the anti-HBs level rose progressively with repeat doses of intradermal vaccine. Half of them developed protective levels of antibody after four doses of intradermal vaccine. The anti-HBs level peaked 2 weeks after the 8th dose of intradermal vaccination at a mean level of 75.6 ± 33.3 miu/mL (Figure 1). Ninety per cent could maintain a protective level of antibody after 6 months (mean 15.9 ± 3.0 miu/mL) and the antibody levels declined to undetectable at 1 year. However, all of them, including the two low-level responders, mounted good antibody responses (mean 322.6 ± 92.0 miu/mL) to the booster dose of intramuscular vaccine given at 12 months, indicating that they were protected from HBV infection. In order to confirm the nonresponsiveness of those 13 patients who did not have a detectable antibody level after the initial course of intradermal vaccine, an intramuscular booster dose of 40 μg HBV vaccine was given to them at a mean of 31.9 ± 0.3 months. Four of them became seropositive (anti-HBs level 13.7–1000 miu/mL) after the intramuscular booster. There was no evidence that these four late responders had any contact with HBV or HBV infection during the interim period between the i.d. course of vaccination and the i.m. booster. Thus, this combination regime of intradermal vaccine and intramuscular booster had achieved an overall response rate of 62.5% (15 persons).

image

Figure 1. Serial changes of the mean anti-HBs titers of the initial responders. Arrows: doses of intradermal hepatitis B vaccination. Bars: error bar (standard error).

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The responders and nonresponders had comparable age (39.6 ± 2.3 vs. 45.4 ± 3.1 years, p = 0.14), gender, etiology of renal failure, mode of dialysis, and type of renal transplant. The duration of dialysis before transplant of the responders tended to be shorter, with a mean duration of 8.9 ± 2.1 months as compared with 21.4 ± 9.8 months in the nonresponders, although the difference did not reach statistical significant (p = 0.13) (Table 1). The time interval from transplant to initiation of the intradermal course of vaccination for responders and nonresponders was 88.1 ± 10.5 and 63.9 ± 7.6 months, respectively (p = 0.12). The time interval from the last dose of intramuscular vaccine to the initiation of the intradermal vaccine was 18.5 ± 2.9 and 10.5 ± 2.9 months, respectively (p = 0.08). There were no significant differences in the doses of prednisolone, azathioprine and cyclosporine A between responders and nonresponders during the course of study. The serum creatinine in both groups was also comparable (Table 2).

Table 1. : Demographic and clinical data of the study population
 Responders n = 15Non-responders n = 9
Age (years)39.6 ± 2.345.4 ± 3.1
Sex (male/female)9/64/5
Underlying cause of renal failure:
 Diabetes10
 Chronic glomerulonephritis52
 Lupus nephritis01
 Alport's syndrome10
 Renal tuberculosis10
 Idiopathic76
Type of transplant
 (Cadaveric/Living donor)9/65/4
Mode of dialysis pretransplant
 (hemodialysis/peritoneal dialysis/pre-emptive)8/6/17/2/0
Duration of dialysis before transplant (months) 8.9 ± 2.121.4 ± 9.8
Table 2. : Immunosuppressive therapy and clinical data of the study population
 Responders n = 15Non-responders n = 9
  1. Triple therapy = cyclosporin A, prednisolone and azathioprine. Double I = cyclosporine A and prednisolone. Double II = azathioprine and prednisolone. i.m. = intramuscular vaccination; i.d. = intradermal vaccination.

Duration of transplant (months) 88.1 ± 10.5 63.9 ± 7.6
Time from last dose of i.m. vaccination (months) 18.5 ± 2.9 10.5 ± 2.9
Type of immunosuppressive therapy8/5/26/3/0
 (triple/double I/double II)
Prednisolone dose (mg/day)  7.3 ± 0.4/6.3 ± 0.4  8.9 ± 0.6/7.2 ± 0.5
 (i.m./i.d.)
Azathioprine dose (mg/day)   50 ± 4.7/50 ± 4.7   50 ± 0/50 ± 0
 (i.m./i.d.)
Cyclosporine A level (μg/L)158.2 ± 9.9/152.4 ± 7.4180.3 ± 6.5/168.6 ± 8.2
 (i.m./i.d.)
Serum creatinine (μmol/L)121.6 ± 6.4143.6 ± 16.4

HLA typing was available in 16 patients. One patient in each group had HLA DR7 and one patient in the responder group had HLA DR3. No patient had the extended haplotypes HLA B8-DR3-SC01 or HLA B44-DR7-FC31and no patient had HLA DR14.

Three patients from the responder group and one patient from the nonresponder group complained of mild pain and redness over the injection sites, which improved after 7–10 days with no permanent scars. No patients reported any other side-effects.

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References

Hepatitis B infection in transplant patients is associated with high morbidity and mortality. The likelihood of death from liver-related complications, including acute hepatic failure, cirrhosis or hepatocellular carcinoma, is increased 3–4-fold in hepatitis B carriers (1,2). In view of the poor immune response to hepatitis B immunization post transplant, it is generally recommended that potential transplant recipients should be immunized prior to transplant. However, there remain renal allograft recipients who have not been vaccinated for hepatitis B and those who have failed to respond to the enhanced scheme of intramuscular vaccination. They remain unprotected against hepatitis B infection.

We have presented data from 24 patients who failed to respond to four or more doses of double-strength intramuscular hepatitis B vaccine. With our 8 doses of 5 μg intradermal HBV vaccination regimen, 45.8% (11 persons) of patients responded with an antibody response, of whom 82% (9 persons) had protective levels of antibody. With a booster dose of 40 μg of intramuscular HBV vaccine 12–32 months after the initial intradermal vaccination, all initial responders responded with a high antibody response. In addition, four patients, who initially did not have detectable antibody with the intradermal course, developed protective levels of anti-HBs after the booster.

This combination of intradermal vaccine together with a double-strength intramuscular booster achieved a very good overall response rate of 62.5%.

No significant predictors of responsiveness to vaccine were identified, although the responders in our study tended to have a shorter period of pretransplant dialysis and a longer duration of transplant. No gender difference in the response rate was observed in our study, although female sex had been reported to be associated with better response previously (11). On the other hand, the underlying renal disease, the donor source, the interval from last intramuscular vaccination, the renal functions and the amount of immunosuppressive therapy did not differ between the responders and nonresponders.

Associations of certain HLA antigens with poor responsiveness to hepatitis B vaccine have been reported previously. HLA DR3, DR4, DR7, DR14 and DP2, especially those homozygotes for the extended haplotypes HLA B8-DR3-SC01, B44-DR7-FC31 (12,13) and Bw54-DR4-DQW4 (14) had been associated with poor responsiveness to hepatitis B vaccine. We could not draw any definite correlation between the HLA typing and the responsiveness of our patients, probably because of the small sample size and racial difference. Further studies with bigger patient populations are warranted for clarification of this issue.

Earlier studies have reported a lower antibody response rate in patients receiving low-dose intradermal vaccination, in particular for patients who had insulin-dependent diabetes mellitus (15). However, these studies involved a lower total dose and less frequent administration of vaccine, which probably would be insufficient for diabetic patients, who had impaired macrophage activity (15). There is emerging evidence for better immunogenicity with the intradermal route of vaccine administration in healthy subjects (6), young children (7) and renal failure patients (16), especially when equivalent doses of vaccine were used for comparison. Intradermal vaccination has also been shown to be able to induce an immune response in healthy subjects (8) and renal failure patients (9,10) who failed to response to intramuscular vaccination previously. Our observation that induction of immunity was successful in 62.5% of transplant patients who did not respond to previous intramuscular vaccination is in line with these studies.

The exact mechanism for improved immune responsiveness with the intradermal route of vaccination is not clear. There was no significant change in the dosage of immunosuppressive therapy during the two vaccination regimes. Langerhans' cells, which are important antigen-presenting cells, are present in the epidermis but not in subcutaneous tissue or muscle (17). Thus the intradermal route may be immunologically more efficient, by directly priming these antigen-presenting cells (18,19). In addition, the vaccine that was introduced via the reinforced protocol of repeated high-dose intradermal injections may be retained for an extended period in the epidermal region. Antigen persistence and appropriate antigen presentation via the Langerhans' cells may thereby stimulate the immune system more effectively and overcome the problem of reduced immune responsiveness in transplant patients. It is interesting to note that the response rate rose from 45.8% to 62.5% with an additional intramuscular booster. It had previously been reported that persons who had insufficient antibody levels after low-dose intradermal vaccination showed an antibody response comparable to that of the responders after an intramuscular booster (20). As the enzyme immunoassay for the antibody level in our laboratory is up to international standard and those four late responders had no evidence of HBV infection during the period between the intradermal course and the intramuscular booster, we believe that they had developed an immune response after the intradermal course although the antibody had not reached a detectable level initially. The intradermal vaccination probably had primed the immune system, and the immunogenicity improved further after the high-dose intramuscular booster.

The intradermal vaccination was well tolerated by our patients. The only side-effect was local skin reaction at the injection sites, which was present in four patients (16.7%). Local skin reactions leading to pigmentation and induration have been reported in 20–67% of patients receiving intradermal vaccination (6,8). We observed a higher incidence of skin reaction in the responder group, but a larger sample number is needed to draw a definite correlation between the incidence of skin reactions and the immune response to HBV vaccine.

In view of the critical importance of preventing hepatitis B virus infection in kidney transplant patients, and the lack of a therapeutic agent with proven long-term efficacy at present, any safe measure that can induce immunity in susceptible patients should be attempted. Although our study is not a randomized control study and thus could not achieve the same power as a control study, our findings suggest that intradermal hepatitis B vaccination together with a high-dose intramuscular booster may benefit renal transplant patients who have failed to respond to conventional intramuscular hepatitis B vaccination. Further multicenter clinical trials are warranted for optimization of the schedule, and to evaluate the effect of combining different routes of immunization.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Materials and Methods
  5. Results
  6. Discussion
  7. References
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