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Inactivated hepatitis A vaccine: active and passive immunoprophylaxis in chimpanzees
Inactivated hepatitis A vaccine: active and passive immunoprophylaxis in chimpanzees Robert H. Purcell *+, Erik D'Hondt¢, Richard Bradbury~, Suzanne U. Emerson*, Sugantha Govindarajan' and Leonard Binn' Studies q/" active and passive inmmnoprot)h.t'Mxis were carried out it, chimpanzees to determine whether a candidate hepatitis A virus (HA V) vaccine could stimulate antibody m HA V (anti-HA 17) that was qualitatively shTfilar to anti-HA V sth~lulated by natural iJ?/Eelion. Normal immune globulin ( I,g ) was prepared.fi'om plasma obtained./kom human vohmteers be/are and after vaccination with the HA V vaccine, and these preparations or commereia/h' prepared I,~ were administered to chimpanzees. Protective q~ca
hmmmoprophylaxis: hepatitis A ~irus: animal model: immunoglobulin
INTRODUCTION Hepatitis A continues to be an important medical problem throughout the world. In developed countries such as the USA. it accounts for -- 20-25% of clinical acute hepatitis. In such countries it is a particular problem for high-risk groups, including staff of day-care centres, indigenous populations, travellers to developing countries and military personnel who must travel to high-risk regions. In developing countries virtually everyone is infected at an early age. There is, therefore, a contintuing need for hepatitis A vaccines in both developed and developing countries. tLaboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, ~National Institutes of Health, Building 7, Room 202, Bethesda, Maryland 20892, USA. =SmithKline Beecham Biologicals, Rixensart, Belgium. §Bioqual Inc., Rockville, Maryland, USA. 'Rancho Los Amigos Hospital, Downey, California, USA. 'Walter Reed Army Institute of Research, Washington, DC, USA. *To whom correspondence should be addressed
S148
Vaccine, Vol. 10, Suppl. 1, 1992
The protective efficacy of hepatitis A virus (HAV) vaccines in field trials poses special problems because of the low clinical attack rate among infants and children (who are the populations at risk) in developing countries and the relatively low overall infection rate in the general population of developed countries. As a supplement to such studies, it is appropriate to attempt to demonstrate the efficacy of candidate HAV vaccines in surrogates of man, namely, susceptible chimpanzees and marmosets. Such animal model systems also provide a means for quantifying the protective response to vaccines and for separating the humoral from the cellular immune responses of the vaccinees. The mechanism of immune protection against HAV is not well understood, but it has been repeatedly documented that antibodies to HAV (anti-HAV) are alone capable of protecting against illness: numerous studies have demonstrated that normal immune globulin (lg), when administered at the recommended dose of 0.02 0.04 ml/kg, is capable of preventing clinical hepatitis following natural challenge. The seruna of individuals 0264-410X/92/100S148-04 ~ 1992 Butterworth-Heinemann Ltd
Protective efficacy of hepatitis A vaccine: R. H. Purcell et al.
who have received such Ig has been shown to contain a relatively low level of anti-HAV t. Thus a small amount of antibody to HAV, in the absence of cell-mediated immunity, is capable of affording protection. It would be interesting to demonstrate in a similar manner that antibodies induced by the vaccine and passively administered to chimpanzees could afford protection. The purpose of this study was to investigate the degree of equivalence between the protective efficacy of the antibodies induced by the vaccine and antibodies stimulated by natural infection. Such equivalence of quality should be demonstrated at a relatively low antibody titre, preferably comparable to the antibody titres detected in an early blood sample of recipients of commercial normal Ig.
MATERIALS AND METHODS
Chimpanzees Twelve chimpanzees were used in this study. They were all born in captivity and were raised, maintained and entered into research protocols under conditions that met or exceeded all US regulations and requirements. All protocols were approved by appropriate US regulatory bodies. The chimpanzees were bled weekly and tested for levels (in units) of serum alanine aminotransferase (ALT), isocitrate dehydrogenase (ICD) and ~,-glutamyltransferase (GGT). Stool samples were collected daily from chimpanzees and stored at minus 20°C. Percutaneous needle biopsies of the liver were obtained weekly throughout the study. These were divided into pieces and fixed in buffered formalin or snap-frozen. Formalin-fixed tissue was embedded, sectioned, stained with hematoxylin and eosin and examined for histopathological changes. Frozen liver tissue was sectioned and stained by the indirect immunofluorescence (IMF) method with mouse monoclonal antiHAV (K32F2N) and fluroescein-labelled goat antimouse globulin2.3.
Serologic tests Serial chimpanzee sera were tested for anti-HAV with HAVAB tests (Abbott Laboratories, North Chicago, IL, USA). Sera were also tested for anti-HAV and for the IgM class of anti-HAV with non-commercial enzymelinked immunosorbent assay (ELISA) methodology (SmithKline Beecham Biologicals, Rixensart, Belgium). Selected sera were tested for neutralizing antibody to HAV with a radioimmunofocus inhibition assay4.
lnocula The following five inocula were used in this study: (a) Inactivated hepatitis A vaccine prepared by SmithKline Beecham Biologicals. This vaccine was from a pre-licensing lot and consisted of HAV, strain HM ! 755, grown in MRC-5 human diploid fibroblasts. The dose of vaccine used in this study contained 360 arbitrary units as measured by ELISA (one-half of the proposed standard dose). The vaccine was adjuvanted with alum and administered intramuscularly in a volume of 0.5 ml. The dose was repeated one month later. (b,c) Immune globulins prepared, for intravenous administration, from the plasma of volunteers before and after vaccination with hepatitis A vaccine, respectively. (d) Commercial
immune globulin for intramuscular administration (commercial normal Ig). The three lg preparations were administered 48 h before challenge. (e) Virulent hepatitis A virus, strain HM-175, used as the challenge virus for determination of protective efficacy. The virus was contained in a suspension of faeces obtained during the acute phase of hepatitis A in an Australian patienP. The challenge dose of virus was ~ 1000 chimpanzee infectious doses, administered intravenously 24 weeks after the administration of the first dose of vaccine and 48 h after administration of the Ig preparations.
Detection of HAV RNA by hybridization Suspensions of stool (10%) in buffer (10mM Tris, 150 mM NaCI, pH 7.2) were prepared and RNA was extracted from them by the proteinase K/phenol method and tested for the presence of HAV RNA by slot-blot hybridization with a riboprobe consisting of 32p-labelled negative-sense RNA of HAV, strain HM-175 v.
RESULTS The chimpanzees, the inocula they received and the results obtained with the various assays are summarized in Table 1 and described below.
Antibody response Both vaccinated chimpanzees developed anti-HAV including IgM anti-HAV, three weeks after vaccination. Both total and IgM anti-HAV reached high levels 2-3 weeks after the second vaccination. IgM anti-HAV diminished in titre 1 and 5 weeks later, respectively, but total anti-HAV remained near peak levels (2512 and 15 849 mIU/ml, respectively) for the remainder of the study. Barely detectable levels of anti-HAV were present prior to the challenge in the chimpanzees that were passively infused with the various Ig preparations (Table 1). However, beginning approximately 2-5 weeks following challenge with virulent HAV, all eight of the chimpanzees passively immunized with Ig and the two uninoculated control chimpanzees developed high titres of anti-HAV, indicating that all were infected by the challenge virus. In general, the anti-HAV response (total, IgM and neutralizing anti-HAV) was slightly delayed and the titres were slightly lower in animals that received anti-HAV, regardless of the source, when compared with control animals. Vaccinated chimpanzees did not develop a significant rise in the titre of total or IgM anti-HAV following challenge with virulent HAV, suggesting that there was little or no replication of the virus, in contrast to the results obtained with all of the other chimpanzees following challenge.
RNA hybridization The chimpanzees that were vaccinated with hepatitis A vaccine or that received commercial Ig did not shed detectable HAV in their stools following challenge with virulent HAV. In contrast, the chimpanzees receiving the pre-vaccination Ig or low-dose post-vaccination Ig shed virus between weeks 2 and 6 postchallenge. One of two animals receiving the higher dose of postvaccination Ig and one of the two positive control animals shed detec-
Vaccine, Vol. 10, Suppl. 1, 1992 $149
Protective efficacy of hepatitis A vaccine." R. H. Purcefl et al. Table1
Active or passive immunization with hepatitis A vaccine
Chimpanzee No. 1380 1332 1383 1441 1374 1422 1382 1406 1396 1420 1413 1442
Immunoprophylaxis Vaccine Vaccine Ig, pre-vaccination Ig, pre-vaccination Ig, post-vaccination, low v o l u m e Ig, post-vaccination, low volume Ig, post-vaccination, high volume Ig, post-vaccination, high volume Ig, commercial Ig, commercial None None
Neutralizing anti-HAV (reciprocal titre) at time of challenge
Virological hybrid
IMF
> > < <
10 000 10 000 10 10
+ +
-
10
+
<10
+
40 40 40 40 <10 < 10
Evidence of infection following challenge Increase in anti-HAV (fold) a
Hepatitis
Total
Peak ALT
IgM
Histology
2 2 _>2628 _>468
4 4 _>7800 _>4000
47 ( ) 40 ( ) 346 ( + ) 113 ( + )
_> 120
_> 1600
51 ( - )
±
_>131
_>1600
80(±)
+
-
+
_>209
_>3200
29 ( - )
-
+ + -
± -
_>322 ->440 _>323 _>2612 _>2702
_>3200 _> 1000 _>2000 _>8000 _>8000
66 ( ) 9 ( ) 48 ( ) 835(+) 103 ( + )
+ -
+
NT + +
alncrease in amount of anti-HAV (measured as mlU for total or reciprocal titre for IgM) between time of challenge and peak level following challenge expressed as fold. NT, Not tested (chimpanzee could not be biopsied for medical reasons). See text for definitions
table virus between weeks 1 and 3 post-challenge. The sensitivity of this assay was ~ 105.s infectious doses of HAV as determined with a standard pool of cell-culture derived virus.
Immunofluorescence Liver biopsies from only two of the chimpanzees were positive for HAV antigen by immunofluorescence. A single biopsy (week 4) from one of the two chimpanzees that received pre-vaccination Ig and one biopsy (week 6) from one chimpanzee that received the higher volume of postvaccination Ig were positive.
Protection against hepatitis As seen in Table l, only the chimpanzees that received experimental Ig prepared from the plasma of vaccinees before vaccination and the chimpanzees that received nothing (positive controls) developed consistent liver enzyme elevations and histopathological changes indicative of hepatitis. In contrast, chimpanzees that were passively immunized with Ig containing anti-HAV were partially or completely protected against hepatitis A following challenge. Thus, all chimpanzees that had detectable anti-HAV prior to challenge, regardless of its source (vaccination or Ig from vaccinated or convalescent humans) were protected against hepatitis A, and all chimpanzees that lacked detectable anti-HAV developed hepatitis A. The lowest virus-neutralizing titre of passively acquired anti-HAV that was protective in chimpanzees was 1:10.
CONCLUSIONS Inactivated hepatitis A vaccine protected chimpanzees against intravenous challenge with virulent HAV as demonstrated by virological, biochemical, histological and serological criteria. Anti-HAV stimulated by vaccination with inactivated HAV vaccine is capable of preventing hepatitis A when administered passively. In this
S150
V a c c i n e , Vol. 10, S u p p l . 1, 1992
respect, it mimics commercial Ig containing anti-HAV stimulated by natural infection with HAV. Although both preparations prevented hepatitis A, neither preparation prevented infection with HAV when exposure was by the intravenous route. In contrast, Ig prepared from human vaccinees before vaccination failed to protect against either illness or infection, thus validating the specificy of the passive protection with the Ig preparations. Thus, anti-HAV stimulated by vaccination of volunteers with inactivated HAV vaccine was capable of passive immunoprophylaxis against hepatitis A when administered to chimpanzees. The reason why passively acquired anti-HAV protected only against hepatitis but not infection, whereas vaccine apparently protected against both, is not known but may simply reflect the higher titres of anti-HAV stimulated by the vaccine. Alternatively, cell-mediated immune respones may have been involved, but the excellent protective effect of traces of passively-acquired antiHAV suggests that cell-mediated responses are not essential and may not be important. It is surprising but gratifying that such a small amount of anti-HAV is protective. It suggests that the titre of anti-HAV present in commercial normal Ig is still adequate to prevent hepatitis A and is likely to be sufficient in the future, even if average levels continue to fall. It is also gratifying to know that anti-HAV stimulated by vaccination is protective when passively administered. This indicates that vaccination with hepatitis A vaccine can be used as a means of stimulating a high titre of anti-HAV in plasmapheresis donors for the purpose of preparing hepatitis A Ig. Active immunoprophylaxis with hepatitis A vaccine is preferable to passive immunoprophylaxis in almost all instances, and such vaccines should largely replace the administration of Ig for the prevention of hepatitis A in the future. The rapid appearance of anti-HAV following only one dose of vaccine suggests that protection is afforded by vaccination within days of administration of the vaccine, making vaccination an attractive alternative to Ig, even in situations where exposure may occur prior to administration of booster doses of vaccine. Further
Protective efficacy o f hepatitis A vaccine: R. H. Purcell et al.
more, the extremely high level of anti-HAV stimulated by vaccine, compared to Ig, makes it likely that vaccination in the community will also prevent not only hepatitis but infection with HAV. Such prevention of infection will minimize spread of HAV more effectively than passive immunoprophylaxis. Finally, the high level of antiHAV stimulated by HAV vaccines promises long-term immunity, a distinct advantage over Ig, which must be administered repeatedly to sustain protection. Future studies will determine how often booster doses of vaccine must be administered and whether subsequent exposure of vaccinees to HAV will stimulate a protective anamnestic anti-HAV response in vaccinees who have lost detectable anti-HAV over time. ACKNOWLEDGEMENT © US Government.
REFERENCES 1 Stapleton, J., Jansen, R. and Lemon, S. Neutralizing antibody to hepatitis A virus in immune serum globulin and in the sera of human recipients of immune serum globulin. Gastroenterology 1985, 89, 637-642 MacGregor, A., Kornitschuk, M., Hurrell, J.G.R., Lehmann, N.I., Coulepis, A.G., Locarnini, S.A. etaL Monoclonal antibodies against hepatitis A virus. J. Clin. Microbiol. 1983, 18, 1237-1243 Anderson, D.A., Coulepis, A.G., Chenoweth, M.P. and Gust, I.D. Indirect immunofluorescense assay for the detection of hepatitis A virusspecific serum immunoglobulins. J. Clin. Microbiol. 1986, 24, 163-165 Lemon, S.M. and Binn, L.N. Serum neutralizing antibody response to hepatitis A virus. J. Infect. Dis. 1983, 148, 1033-1039 Andre, F.E., Hepburn A. and D'Hondt, E. Inactivated candidate vaccines for hepatitis A. Prog. Med. ViroL 1990, 37, 72-95 Gust, I.D., Lehmann, N.I., Crowe, S., McCrorie, M., Locarnini, S.A. and Lucas, C.R. The origin of the HM175 strain of hepatitis A virus. J. Infect. Dis. 1985, 151,365-367 Emerson, S.U., McRill, C., Rosenblum, B., Feinstone, S. and Purcell, R.H. Mutations responsible for adaptation of hepatitis A virus to efficient growth in cell culture. J. Virol. 1991, 65, 4882-4886
Vaccine, Vol. 10, Suppl. 1, 1992
$151
hmmmoprophylaxis: hepatitis A ~irus: animal model: immunoglobulin
INTRODUCTION Hepatitis A continues to be an important medical problem throughout the world. In developed countries such as the USA. it accounts for -- 20-25% of clinical acute hepatitis. In such countries it is a particular problem for high-risk groups, including staff of day-care centres, indigenous populations, travellers to developing countries and military personnel who must travel to high-risk regions. In developing countries virtually everyone is infected at an early age. There is, therefore, a contintuing need for hepatitis A vaccines in both developed and developing countries. tLaboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, ~National Institutes of Health, Building 7, Room 202, Bethesda, Maryland 20892, USA. =SmithKline Beecham Biologicals, Rixensart, Belgium. §Bioqual Inc., Rockville, Maryland, USA. 'Rancho Los Amigos Hospital, Downey, California, USA. 'Walter Reed Army Institute of Research, Washington, DC, USA. *To whom correspondence should be addressed
S148
Vaccine, Vol. 10, Suppl. 1, 1992
The protective efficacy of hepatitis A virus (HAV) vaccines in field trials poses special problems because of the low clinical attack rate among infants and children (who are the populations at risk) in developing countries and the relatively low overall infection rate in the general population of developed countries. As a supplement to such studies, it is appropriate to attempt to demonstrate the efficacy of candidate HAV vaccines in surrogates of man, namely, susceptible chimpanzees and marmosets. Such animal model systems also provide a means for quantifying the protective response to vaccines and for separating the humoral from the cellular immune responses of the vaccinees. The mechanism of immune protection against HAV is not well understood, but it has been repeatedly documented that antibodies to HAV (anti-HAV) are alone capable of protecting against illness: numerous studies have demonstrated that normal immune globulin (lg), when administered at the recommended dose of 0.02 0.04 ml/kg, is capable of preventing clinical hepatitis following natural challenge. The seruna of individuals 0264-410X/92/100S148-04 ~ 1992 Butterworth-Heinemann Ltd
Protective efficacy of hepatitis A vaccine: R. H. Purcell et al.
who have received such Ig has been shown to contain a relatively low level of anti-HAV t. Thus a small amount of antibody to HAV, in the absence of cell-mediated immunity, is capable of affording protection. It would be interesting to demonstrate in a similar manner that antibodies induced by the vaccine and passively administered to chimpanzees could afford protection. The purpose of this study was to investigate the degree of equivalence between the protective efficacy of the antibodies induced by the vaccine and antibodies stimulated by natural infection. Such equivalence of quality should be demonstrated at a relatively low antibody titre, preferably comparable to the antibody titres detected in an early blood sample of recipients of commercial normal Ig.
MATERIALS AND METHODS
Chimpanzees Twelve chimpanzees were used in this study. They were all born in captivity and were raised, maintained and entered into research protocols under conditions that met or exceeded all US regulations and requirements. All protocols were approved by appropriate US regulatory bodies. The chimpanzees were bled weekly and tested for levels (in units) of serum alanine aminotransferase (ALT), isocitrate dehydrogenase (ICD) and ~,-glutamyltransferase (GGT). Stool samples were collected daily from chimpanzees and stored at minus 20°C. Percutaneous needle biopsies of the liver were obtained weekly throughout the study. These were divided into pieces and fixed in buffered formalin or snap-frozen. Formalin-fixed tissue was embedded, sectioned, stained with hematoxylin and eosin and examined for histopathological changes. Frozen liver tissue was sectioned and stained by the indirect immunofluorescence (IMF) method with mouse monoclonal antiHAV (K32F2N) and fluroescein-labelled goat antimouse globulin2.3.
Serologic tests Serial chimpanzee sera were tested for anti-HAV with HAVAB tests (Abbott Laboratories, North Chicago, IL, USA). Sera were also tested for anti-HAV and for the IgM class of anti-HAV with non-commercial enzymelinked immunosorbent assay (ELISA) methodology (SmithKline Beecham Biologicals, Rixensart, Belgium). Selected sera were tested for neutralizing antibody to HAV with a radioimmunofocus inhibition assay4.
lnocula The following five inocula were used in this study: (a) Inactivated hepatitis A vaccine prepared by SmithKline Beecham Biologicals. This vaccine was from a pre-licensing lot and consisted of HAV, strain HM ! 755, grown in MRC-5 human diploid fibroblasts. The dose of vaccine used in this study contained 360 arbitrary units as measured by ELISA (one-half of the proposed standard dose). The vaccine was adjuvanted with alum and administered intramuscularly in a volume of 0.5 ml. The dose was repeated one month later. (b,c) Immune globulins prepared, for intravenous administration, from the plasma of volunteers before and after vaccination with hepatitis A vaccine, respectively. (d) Commercial
immune globulin for intramuscular administration (commercial normal Ig). The three lg preparations were administered 48 h before challenge. (e) Virulent hepatitis A virus, strain HM-175, used as the challenge virus for determination of protective efficacy. The virus was contained in a suspension of faeces obtained during the acute phase of hepatitis A in an Australian patienP. The challenge dose of virus was ~ 1000 chimpanzee infectious doses, administered intravenously 24 weeks after the administration of the first dose of vaccine and 48 h after administration of the Ig preparations.
Detection of HAV RNA by hybridization Suspensions of stool (10%) in buffer (10mM Tris, 150 mM NaCI, pH 7.2) were prepared and RNA was extracted from them by the proteinase K/phenol method and tested for the presence of HAV RNA by slot-blot hybridization with a riboprobe consisting of 32p-labelled negative-sense RNA of HAV, strain HM-175 v.
RESULTS The chimpanzees, the inocula they received and the results obtained with the various assays are summarized in Table 1 and described below.
Antibody response Both vaccinated chimpanzees developed anti-HAV including IgM anti-HAV, three weeks after vaccination. Both total and IgM anti-HAV reached high levels 2-3 weeks after the second vaccination. IgM anti-HAV diminished in titre 1 and 5 weeks later, respectively, but total anti-HAV remained near peak levels (2512 and 15 849 mIU/ml, respectively) for the remainder of the study. Barely detectable levels of anti-HAV were present prior to the challenge in the chimpanzees that were passively infused with the various Ig preparations (Table 1). However, beginning approximately 2-5 weeks following challenge with virulent HAV, all eight of the chimpanzees passively immunized with Ig and the two uninoculated control chimpanzees developed high titres of anti-HAV, indicating that all were infected by the challenge virus. In general, the anti-HAV response (total, IgM and neutralizing anti-HAV) was slightly delayed and the titres were slightly lower in animals that received anti-HAV, regardless of the source, when compared with control animals. Vaccinated chimpanzees did not develop a significant rise in the titre of total or IgM anti-HAV following challenge with virulent HAV, suggesting that there was little or no replication of the virus, in contrast to the results obtained with all of the other chimpanzees following challenge.
RNA hybridization The chimpanzees that were vaccinated with hepatitis A vaccine or that received commercial Ig did not shed detectable HAV in their stools following challenge with virulent HAV. In contrast, the chimpanzees receiving the pre-vaccination Ig or low-dose post-vaccination Ig shed virus between weeks 2 and 6 postchallenge. One of two animals receiving the higher dose of postvaccination Ig and one of the two positive control animals shed detec-
Vaccine, Vol. 10, Suppl. 1, 1992 $149
Protective efficacy of hepatitis A vaccine." R. H. Purcefl et al. Table1
Active or passive immunization with hepatitis A vaccine
Chimpanzee No. 1380 1332 1383 1441 1374 1422 1382 1406 1396 1420 1413 1442
Immunoprophylaxis Vaccine Vaccine Ig, pre-vaccination Ig, pre-vaccination Ig, post-vaccination, low v o l u m e Ig, post-vaccination, low volume Ig, post-vaccination, high volume Ig, post-vaccination, high volume Ig, commercial Ig, commercial None None
Neutralizing anti-HAV (reciprocal titre) at time of challenge
Virological hybrid
IMF
> > < <
10 000 10 000 10 10
+ +
-
10
+
<10
+
40 40 40 40 <10 < 10
Evidence of infection following challenge Increase in anti-HAV (fold) a
Hepatitis
Total
Peak ALT
IgM
Histology
2 2 _>2628 _>468
4 4 _>7800 _>4000
47 ( ) 40 ( ) 346 ( + ) 113 ( + )
_> 120
_> 1600
51 ( - )
±
_>131
_>1600
80(±)
+
-
+
_>209
_>3200
29 ( - )
-
+ + -
± -
_>322 ->440 _>323 _>2612 _>2702
_>3200 _> 1000 _>2000 _>8000 _>8000
66 ( ) 9 ( ) 48 ( ) 835(+) 103 ( + )
+ -
+
NT + +
alncrease in amount of anti-HAV (measured as mlU for total or reciprocal titre for IgM) between time of challenge and peak level following challenge expressed as fold. NT, Not tested (chimpanzee could not be biopsied for medical reasons). See text for definitions
table virus between weeks 1 and 3 post-challenge. The sensitivity of this assay was ~ 105.s infectious doses of HAV as determined with a standard pool of cell-culture derived virus.
Immunofluorescence Liver biopsies from only two of the chimpanzees were positive for HAV antigen by immunofluorescence. A single biopsy (week 4) from one of the two chimpanzees that received pre-vaccination Ig and one biopsy (week 6) from one chimpanzee that received the higher volume of postvaccination Ig were positive.
Protection against hepatitis As seen in Table l, only the chimpanzees that received experimental Ig prepared from the plasma of vaccinees before vaccination and the chimpanzees that received nothing (positive controls) developed consistent liver enzyme elevations and histopathological changes indicative of hepatitis. In contrast, chimpanzees that were passively immunized with Ig containing anti-HAV were partially or completely protected against hepatitis A following challenge. Thus, all chimpanzees that had detectable anti-HAV prior to challenge, regardless of its source (vaccination or Ig from vaccinated or convalescent humans) were protected against hepatitis A, and all chimpanzees that lacked detectable anti-HAV developed hepatitis A. The lowest virus-neutralizing titre of passively acquired anti-HAV that was protective in chimpanzees was 1:10.
CONCLUSIONS Inactivated hepatitis A vaccine protected chimpanzees against intravenous challenge with virulent HAV as demonstrated by virological, biochemical, histological and serological criteria. Anti-HAV stimulated by vaccination with inactivated HAV vaccine is capable of preventing hepatitis A when administered passively. In this
S150
V a c c i n e , Vol. 10, S u p p l . 1, 1992
respect, it mimics commercial Ig containing anti-HAV stimulated by natural infection with HAV. Although both preparations prevented hepatitis A, neither preparation prevented infection with HAV when exposure was by the intravenous route. In contrast, Ig prepared from human vaccinees before vaccination failed to protect against either illness or infection, thus validating the specificy of the passive protection with the Ig preparations. Thus, anti-HAV stimulated by vaccination of volunteers with inactivated HAV vaccine was capable of passive immunoprophylaxis against hepatitis A when administered to chimpanzees. The reason why passively acquired anti-HAV protected only against hepatitis but not infection, whereas vaccine apparently protected against both, is not known but may simply reflect the higher titres of anti-HAV stimulated by the vaccine. Alternatively, cell-mediated immune respones may have been involved, but the excellent protective effect of traces of passively-acquired antiHAV suggests that cell-mediated responses are not essential and may not be important. It is surprising but gratifying that such a small amount of anti-HAV is protective. It suggests that the titre of anti-HAV present in commercial normal Ig is still adequate to prevent hepatitis A and is likely to be sufficient in the future, even if average levels continue to fall. It is also gratifying to know that anti-HAV stimulated by vaccination is protective when passively administered. This indicates that vaccination with hepatitis A vaccine can be used as a means of stimulating a high titre of anti-HAV in plasmapheresis donors for the purpose of preparing hepatitis A Ig. Active immunoprophylaxis with hepatitis A vaccine is preferable to passive immunoprophylaxis in almost all instances, and such vaccines should largely replace the administration of Ig for the prevention of hepatitis A in the future. The rapid appearance of anti-HAV following only one dose of vaccine suggests that protection is afforded by vaccination within days of administration of the vaccine, making vaccination an attractive alternative to Ig, even in situations where exposure may occur prior to administration of booster doses of vaccine. Further
Protective efficacy o f hepatitis A vaccine: R. H. Purcell et al.
more, the extremely high level of anti-HAV stimulated by vaccine, compared to Ig, makes it likely that vaccination in the community will also prevent not only hepatitis but infection with HAV. Such prevention of infection will minimize spread of HAV more effectively than passive immunoprophylaxis. Finally, the high level of antiHAV stimulated by HAV vaccines promises long-term immunity, a distinct advantage over Ig, which must be administered repeatedly to sustain protection. Future studies will determine how often booster doses of vaccine must be administered and whether subsequent exposure of vaccinees to HAV will stimulate a protective anamnestic anti-HAV response in vaccinees who have lost detectable anti-HAV over time. ACKNOWLEDGEMENT © US Government.
REFERENCES 1 Stapleton, J., Jansen, R. and Lemon, S. Neutralizing antibody to hepatitis A virus in immune serum globulin and in the sera of human recipients of immune serum globulin. Gastroenterology 1985, 89, 637-642 MacGregor, A., Kornitschuk, M., Hurrell, J.G.R., Lehmann, N.I., Coulepis, A.G., Locarnini, S.A. etaL Monoclonal antibodies against hepatitis A virus. J. Clin. Microbiol. 1983, 18, 1237-1243 Anderson, D.A., Coulepis, A.G., Chenoweth, M.P. and Gust, I.D. Indirect immunofluorescense assay for the detection of hepatitis A virusspecific serum immunoglobulins. J. Clin. Microbiol. 1986, 24, 163-165 Lemon, S.M. and Binn, L.N. Serum neutralizing antibody response to hepatitis A virus. J. Infect. Dis. 1983, 148, 1033-1039 Andre, F.E., Hepburn A. and D'Hondt, E. Inactivated candidate vaccines for hepatitis A. Prog. Med. ViroL 1990, 37, 72-95 Gust, I.D., Lehmann, N.I., Crowe, S., McCrorie, M., Locarnini, S.A. and Lucas, C.R. The origin of the HM175 strain of hepatitis A virus. J. Infect. Dis. 1985, 151,365-367 Emerson, S.U., McRill, C., Rosenblum, B., Feinstone, S. and Purcell, R.H. Mutations responsible for adaptation of hepatitis A virus to efficient growth in cell culture. J. Virol. 1991, 65, 4882-4886
Vaccine, Vol. 10, Suppl. 1, 1992
$151