Overcoming Original (Antigenic) Sin

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Overcoming Original (Antigenic) Sin

Postby malernee » Wed Aug 02, 2006 11:17 am

antigenic sin
Our immune system protects us against death by infection. A major component of the immune system is generation of antibodies, protein molecules that bind specific antigens. To recognize invading pathogens, the immune system performs a search of the amino acid sequence space of possible antibodies. To find useful antibodies in the effectively infinite protein sequence space, the immune system has evolved a hierarchical strategy. Once the immune system has been faced with an antigen, a state of memory is established, which allows the immune system to respond more rapidly and effectively upon subsequent encounters with the same antigen. Although our immune system is highly effective, some limitations have been reported. The phenomenon known as ``original antigenic sin'' is the tendency for antibodies produced in response to exposure to influenza virus antigens to suppress the creation of new, different antibodies in response to exposure to different versions of the flu. The phenomenon of original antigenic sin has been observed in the flu, dengue fever, human immunodeficiency virus (HIV), and other viruses.

Using random energy models, we investigate the dynamics of original antigenic sin in the immune system. The phenomenon of original antigenic sin is explained as stemming from localization of the immune system response in antibody sequence space. This localization is a result of the roughness in sequence space of the evolved antibody affinity constant for antigen and is observed for diseases with high year-to-year mutation rates, such as influenza. These results suggest several implications for vaccination strategies against viruses and cancers.

http://64.233.161.104/search?q=cache:HL ... =clnk&cd=1
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1: Parasite Immunol. 1993 Apr;15(4):187-93. Related Articles, Links


'Original antigenic sin', T cell memory, and malaria sporozoite immunity: an hypothesis for immune evasion.

Good MF, Zevering Y, Currier J, Bilsborough J.

Molecular Immunology Laboratory, Queensland Institute of Medical Research, Brisbane, Australia.

Prior to any exposure to malaria, most adults have T cells specific for malaria parasites and various malaria proteins. The protein for which this has been shown more than any other is the circumsporozoite protein (CSP) of Plasmodium falciparum. These T cells can be present in high frequency and appear to have arisen through exposure to other (non-malaria) organisms. Although T cells are thought to provide protection against sporozoites, these T cells specific for cross-reactive organisms are clearly unable to protect against malaria, and may be preferentially expanded following exposure to malaria sporozoites. Thus, cross-reactive organisms have the potential to skew the repertoire of sporozoite-induced T cells and affect the induction of protective immunity. This is analogous to the concept of 'original antigenic sin' whereby prior exposure to one strain of influenza virus was shown to be able to divert the antibody response to a second challenging strain to focus on the shared (cross-reactive) epitopes.

PMID: 7685075 [PubMed - indexed for MEDLINE]
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Another reason to require revaccination programs show efficacy is the risk of antigenic sin.


Nat Med. 2003 Jul;9(7):820-2.
Original antigenic sin and apoptosis in the pathogenesis of dengue hemorrhagic fever.

Mongkolsapaya J, Dejnirattisai W, Xu XN, Vasanawathana S, Tangthawornchaikul N, Chairunsri A, Sawasdivorn S, Duangchinda T, Dong T, Rowland-Jones S, Yenchitsomanus PT, McMichael A, Malasit P, Screaton G.

MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, OX3 9DS, UK.

Dengue virus presents a growing threat to public health in the developing world. Four major serotypes of dengue virus have been characterized, and epidemiological evidence shows that dengue hemorrhagic fever (DHF), the more serious manifestation of the disease, occurs more frequently upon reinfection with a second serotype. We have studied dengue virus-specific T-cell responses in Thai children. During acute infection, few dengue-responsive CD8+ T cells were recovered; most of those present showed an activated phenotype and were undergoing programmed cell death. Many dengue-specific T cells were of low affinity for the infecting virus and showed higher affinity for other, probably previously encountered strains. Profound T-cell activation and death may contribute to the systemic disturbances leading to DHF, and original antigenic sin in the T-cell responses may suppress or delay viral elimination, leading to higher viral loads and increased immunopathology.

PMID: 12808447 [PubMed - indexed for MEDLINE]
and apoptosis in the pathogenesis of dengue hemorrhagic fever.

Mongkolsapaya J, Dejnirattisai W, Xu XN, Vasanawathana S, Tangthawornchaikul N, Chairunsri A, Sawasdivorn S, Duangchinda T, Dong T, Rowland-Jones S, Yenchitsomanus PT, McMichael A, Malasit P, Screaton G.

MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, Oxford, OX3 9DS, UK.

Dengue virus presents a growing threat to public health in the developing world. Four major serotypes of dengue virus have been characterized, and epidemiological evidence shows that dengue hemorrhagic fever (DHF), the more serious manifestation of the disease, occurs more frequently upon reinfection with a second serotype. We have studied dengue virus-specific T-cell responses in Thai children. During acute infection, few dengue-responsive CD8+ T cells were recovered; most of those present showed an activated phenotype and were undergoing programmed cell death. Many dengue-specific T cells were of low affinity for the infecting virus and showed higher affinity for other, probably previously encountered strains. Profound T-cell activation and death may contribute to the systemic disturbances leading to DHF, and original antigenic sin in the T-cell responses may suppress or delay viral elimination, leading to higher viral loads and increased immunopathology.

PMID: 12808447 [PubMed - indexed for MEDLINE]
art malernee dvm
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Immunology: The original sin of killer T cells

ANDREW J. MCMICHAEL



The phrase 'original antigenic sin' was first used to describe the antibody response to influenza virus. After an initial infection, reinfection (or vaccination) with a new strain of the virus boosted the concentration of antibodies specific for the earlier infecting strain1,2. Although these antibodies cross-reacted with the new virus, they had higher affinity for the original infecting strain. This was immediately seen to have big implications for vaccine design -- a vaccine based on a new strain of influenza virus might be unable to prime antibodies to the intended virus in people who had already been infected with a related viral strain.

Until now, original antigenic sin has been regarded as largely an antibody phenomenon. But, on page 482 of this issue, Klenerman and Zinkernagel3 describe original sin in the response of cytotoxic T lymphocytes (CTLs) to lymphocytic choriomeningitis virus (LCMV). Their work was stimulated by studies of people with the human immunodeficiency virus (HIV), who sometimes mount a CTL response to an immunodominant strain of the virus with weak or no response to the other immunogenic variants present4,5.

The authors attacked the problem in vivo. They infected mice with strains of LCMV that were either normal (wild type) or mutated at the immunodominant epitopes recognized by the CTL. For each mutated strain studied, they found that the CTL response was asymmetrical. Mice infected with the wild-type virus showed only a weak cross-reactive specificity when challenged with the mutant strain, reacting mainly to the wild-type virus. The low reactivity of CTLs against the mutant strain was also associated with delayed clearance of this strain by the immune system. In contrast, the CTL responses from mice infected with the mutant virus cross-reacted equally with the mutant and wild-type strains. Thus, the order in which the mice are exposed to different variants of the virus could have a significant effect on the outcome of the How does this happen? A possible explanation6 for antibody original antigenic sin emerged when helper T cells were discovered, and it was realized that T cells react with peptide fragments of viral proteins that are often conserved between different strains. So, helper T cells, primed by the original virus and then stimulated by the new infection, might activate memory B cells that are specific for the original virus (although it is not clear why antibodies specific for the new variant are not stimulated too). In the same way, the phenomenon observed by Klenerman and Zinkernagel3 might reflect the very strong CTL memory response to LCMV infection. Studies7-9 of acute LCMV infection in mice showed that massive CTL responses occurred, whereby 25-50% of all CD8-positive T cells were virus specific. Even after recovery, 10% of peripheral CD8-positive T cells were specific for the immunodominant LCMV epitopes. A weakly cross-reacting new virus might, therefore, reactivate these plentiful CTL memory cells more readily than the much less abundant naive CTL precursors (Fig. 1). Although this mechanism depends on very high levels of memory CTLs, similar numbers are seen in persistent HIV infection10, where original antigenic sin may also occur4,5.

infection.http://www.37c.com.cn/topic/004/spotlig ... uno/07.htm
***
Just as in the initial description of original antigenic sin, the new work3 has implications for vaccine design. Many groups are now trying to design CTL-inducing vaccines for the control of variable viruses such as HIV and hepatitis C virus. Original antigenic sin means that a monovalent vaccine (such as a peptide) intended to stimulate CTLs may not work if the virus varies at that epitope. Not only would the vaccine-induced CTL response fail to control infection with any variant virus, but the CTL response to the equivalent epitope in that virus might also be impaired. The vaccine might even make the infection worse. The probable solution is to choose several well-conserved epitopes for the vaccine -- but HIV is so variable, these may be in short supply.



Andrew J. McMichael is at the Institute of Molecular Medicine, John Radcliffe Hospital, Oxford OX3 9DU, UK.
e-mail: amcmichael@hammer.imm.ox.ac.uk




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Klenerman, P., Meier, U. C., Phillips, R. E. & McMichael, A. J. Eur. J. Immunol. 25, 1927-1931 (1995). Links
Askinas, B. A., McMichael, A. J. & Webster, R. G. in Basic and Applied Influenza Research (ed. Beare, A. S.) 157-188 (CRC Press, Boca Raton, FL, 1982). Links
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Flu Shot
10.30.03
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Should you bother getting a flu shot this fall? The last three flu seasons have been pretty mild, and the vaccine can’t protect you entirely. But as this ScienCentral News video reports, one scientist found out why skipping a flu shot one year can increase your flu risk the next.

Nothing to Sneeze At

Every winter, some people, mainly those who are elderly or suffering from chronic illnesses, die of influenza. So every fall, health clinics, doctors' offices, and pharmacies offer flu vaccinations. When one scientist got his recently, he also got some surprising news.

Bioengineer Michael W. Deem, professor of bioengineering, physics, and astronomy at Rice University in Houston, Texas, went to get his flu shot at a local pharmacy. The nurse who vaccinated Deem mentioned that he ran some risk if he skipped his flu shot the following year: “If I was getting the flu shot that year, and not the year after, I might be more likely to get the flu the year after."

The nurse's warning startled Deem: shouldn't the effects of a flu vaccination last for years, he wondered, the way vaccinations for polio or smallpox do? He decided to look into what other researchers had found out about the immune system and how it works. The nurse, he discovered, had been right: the risk she mentioned is due to a biological phenomenon known as “original antigenic sin”, first discovered in people and farm animals in 1953.

The term is a nod to both science and theology. An antigen is biologists’ term for an invading organism, such as the influenza virus. Original sin is a theological concept that accounts for human flaws. Original antigenic sin describes an apparent failing in the human immune system: it may recognize a certain strain of a disease, such as this year’s strain of flu virus, but then tries to combat an entirely different strain by "remembering" how it fought the first strain it encountered. As a result, if you skip a flu shot one year, Deem says, you may be more likely to get the flu during that year, compared to your chances of illness if you had never gotten a flu shot in previous years. "It’s as if the immune system didn't have any memory whatsoever, and simply started to learn about this year's flu from scratch," Deem says.


Three antibody fragments bind to the protein surface of influenza virus.
image: Protein Data Bank, Rutgers University
When the human body is confronted with antigens, such as flu virus or a vaccine, it defends itself with antibodies—proteins produced in white blood cells—that track down and fend off invading antigens. A portion of each antibody containing just the right sequence latches onto the part of an antigen with the exact matching sequence to form this bond. Then the antibody either kills the antigen or signals other immune cells for help.

Next time you are exposed to that particular antigen, you have antibodies that usually protect you. But viruses that mutate as quickly as influenza does, may have several different areas where antibodies could latch on—areas that may change every year, foiling existing antibodies.

"The flu changes from year to year because of random mutations," says Deem. "When someone gets the flu, lymph nodes under the arms and under the cheeks swell because they're producing different antibodies. Those antibodies become better and better at recognizing the flu. Those antibodies are still around the next year, and sometimes those antibodies don't respond as well to a new flu as new antibodies would. So the immune system’s memory actually inhibits the system from learning something new about the flu next year."

At Rice, Deem and his research team developed a computational model that demonstrates how original antigenic sin operates. "Our interest was in understanding this phenomenon of original antigenic sin in a quantitative way," Deem explains.

At present, to help combat new flu strains, every flu vaccine contains three strains, and one strain is changed annually. Deem hopes that with his model, “perhaps we could point towards better design efforts for changing the flu vaccine from year to year."

His research also could contribute to the development of vaccines for other viral diseases to which original antigenic sin applies as well—HIV, chlamydia, hepatitis, and dengue fever. Deem points out that their common feature is either multiple strains, as in dengue fever, or very rapid mutation, as in HIV and the flu.

Meanwhile, he recommends a flu shot every fall: "The flu shot protects you quite a bit against the flu this year, and your increased susceptibility next year from original antigenic sin is much smaller than the increased protection this year. And if you get the flu shot next year, you’ll be protected against next year's flu.” Anyone who is particularly susceptible should not skip an annual flu shot.

Deem's research appeared in the August 8, 2003 issue of the journal Physical Review Letters and was presented on September 8, 2003 at the national meeting of the American Chemical Society. It was funded by the National Science Foundation (NSF), and the Camille and Henry Dreyfus Foundation, Inc.
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Overcoming Original (Antigenic) Sin
David E. Anderson, Maria P. Carlos, Lynn Nguyen, and Jose´ V. Torres
School of Medicine, Medical Microbiology and Immunology, University of California–Davis, Davis, California 95616
Original antigenic sin describes a phenomenon in
which the antibody response elicited in an individual
after a secondary viral infection reacts more strongly
to the viral variant that originally infected the individual.
As T helper cells play critical roles in promoting
antibody responses, a similar phenomenon may
hold true for T helper cell responses. This concept is
particularly relevant to the development of vaccines
against viruses such as human immunodeficiency virus
and hepatitis C virus, in which myriad viral variants
are present throughout the human population.
We have compared the effects of priming the immune
system with a single peptide epitope or with a cocktail
of related peptides based on the epitope. Our data
demonstrate that immunization with multiple peptide
variants expands a more broadly reactive and durable
T helper cell response than does immunization with a
single peptide. This vaccine strategy may circumvent
original antigenic sin. © 2001 Academic Press
Key Words: HIV; AIDS; vaccine; peptide; T cell;
memory.
INTRODUCTION
An effective vaccine against human immunodeficiency
virus (HIV) will likely need to expand virusspecific
T helper cell, cytotoxic T lymphocyte (CTL),
and antibody responses. T helper cell proliferative responses
against HIV or simian immunodeficiency virus
(SIV) antigens correlate with reduced viral loads and
delayed progression to disease (1–4). This observation
is not surprising given that in addition to their longestablished
role in providing B cell help, T helper cells
play critical roles in generating and maintaining CTL
activity in vivo (5–7). Strong CTL responses are associated
with lower viral loads and neutralizing antibodies
can protect against viral challenges (8 –15).
Two problems that have vexed the HIV vaccine field
are the inability to elicit broadly reactive neutralizing
antibody responses and the relatively short duration of
cellular and humoral immunity elicited with candidate
HIV vaccines (16). We have previously described a
vaccine approach that uses degenerative peptide cocktails,
termed hypervariable epitope constructs (HECs).
HECs are based on hypervariable epitopes of the envelope
glycoproteins of HIV or SIV and are used to
elicit broadly reactive, long-lasting neutralizing antibody
responses (17, 18). In the present study we compared
the ability of a HEC based on the hypervariable
V3 region of HIV with a peptide based on a single
isolate of HIV to similarly expand broadly reactive T
helper cell responses. Our results demonstrate that
immunization of mice with a HEC expanded T cells
that were more broadly reactive than did mice immunized
with the single peptide. Significantly, the T cell
repertoire expanded after HEC immunization was
more durable than the repertoire expanded after immunization
with the single peptide and recognized naturally
processed antigen more readily.
MATERIALS AND METHODS
Design of HECs and Peptide Synthesis
The design and synthesis of the HECs based on the
V3 loop of HIV-1 gp120 and on the V4 loop of SIV gp130
have been described extensively elsewhere (17, 18).
Briefly, in the design of the HECs, sequences obtained
from the Los Alamos Human Retroviruses and AIDS
Database for the two regions of interest were aligned.
The most common amino acids at each position along
the region of interest were selected for addition during
the peptide synthesis, based on the frequency with
which they occurred among clade B strains of HIV-1
(HEC V3) or among strains of SIVmac and SIVsm
(HEC V4). The peptide mixtures were synthesized using
9-fluorenylmethoxycarbonyl (Fmoc) chemistry utilizing
high-capacity (0.7 mmol/g) Knorr resin (Advanced
ChemTech, Inc., Louisville, KY) with 1%
divinylbenzene crosslinker. Standard Fmoc chemistry
was used for the synthesis of the MN V3, RF V3, V4
(142), and V4 (239) peptides. Their sequences are provided
in Table 1. Amino acid analysis ensured the
presence of the expected amino acids in all peptides
and HECs.
Immunization of Mice
Balb/c mice 6–8 weeks old were housed in conventional
facilities and immunized twice with 100 mg of
peptide or HEC, the second immunization occurring
Clinical Immunology
Vol. 101, No. 2, November, pp. 152–157, 2001
doi:10.1006/clim.2001.5114, available online at http://www.idealibrary.com on
1521-6616/01 $35.00
Copyright © 2001 by Academic Press
All rights of reproduction in any form reserved.
152
three weeks after the first. They were immunized with
peptides or HECs mixed at a 1:1 ratio with the adjuvant
Montanide ISA-51 (Seppic, Paris, France) and
PBS, in a final volume of 100 ml/mouse. This oil-based
adjuvant has been approved for use in humans. Subcutaneous
immunization was performed at the base of
the tail.
Quantitation of Antigen-Specific T Helper Cells
Two weeks and two months after a second immunization,
mice were sacrificed and their spleens and
draining lymph nodes were harvested. Isolated cells
(2 3 106/ml) were left unstimulated or stimulated with
indicated peptides or HECs at a final concentration of
10 mg/ml. Purified whole inactivated SIVmac251 virus
(ABI, Columbia, MD) was used at 5 mg/ml. An irrelevant
negative control peptide derived from Type D
simian retrovirus was used as an additional negative
control. Typically there was little difference in the frequency
of CD691 T cells left unstimulated or stimulated
with the negative control peptide. After 16 h in a
37°C CO2 incubator, cells were collected, washed with
PBS, and stained with anti-CD4-FITC and anti-
CD69-PE monoclonal antibodies (Caltag, Burlingame,
CA) according to the manufacturer’s recommendations.
The frequency of T helper cells specific for each
antigen was determined by calculating the percentage
of CD41 T cells that were also CD691 (a marker of T
cell activation). Typically, 100,000 events were collected
from splenocyte samples and 25,000 events from
lymph node samples. Expression of CD69 induced by
stimulation with irrelevant peptide was subtracted
from all values. The frequency of antigen-specific
splenocytes and lymph node cells for each animal was
calculated individually. These two values were added
to arrive at a total frequency of antigen-specific T cells
for each animal. Statistical significance among different
immunization and stimulation conditions was determined
with a Mann–Whitney test using GraphPad
Instat 2.03 for Mac, GraphPad Software (San Diego,
CA).
RESULTS
Broadly Reactive and Durable T Cell Memory
Elicited with a Hypervariable Epitope Construct
Hypervariable epitope constructs are degenerative
peptide cocktails that are made in a single synthesis
reaction and that are based on hypervariable B cell and
T helper cell epitopes. Their design and ability to elicit
broadly reactive neutralizing antibodies in mice, rabbits,
and nonhuman primates have been described extensively
elsewhere (17, 18). In the current study we
wished to determine whether immunization of mice
with a HEC based on the V3 loop of HIV-1 clade B
viruses could similarly expand broadly reactive T
helper cell responses. We compared T cell responses
elicited by the V3 HEC to those elicited with a single
peptide based on the V3 loop of the HIV-1MN strain of
TABLE 1
Peptide and HEC Immunogens
Note. The predicted peptide sequences contained within the HEC V3 based on the third hypervariable loop of HIV-1
gp120 and the HEC V4 based on the fourth hypervariable loop of SIV gp130 are indicated. The HIV-1 HEC V3 is predicted
to contain 64 different peptide variants and the SIV HEC V4 to contain 32,768 peptide variants. Related peptide
sequences based on the HEC V3 and HEC V4 epitopes are also provided.
OVERCOMING ORIGINAL (ANTIGENIC) SIN 153
virus. The sequences of the HEC V3 and the MN V3
peptide are illustrated in Table 1; the sequence of the
V3 loop of the HIV-1RF strain of virus, which was used
to assess reactivity to a heterologous epitope, is also
provided. The HEC V3 is predicted to contain 64 different
peptides, which represent the most commonly
occurring epitope variants based on available in vivo
sequence data.
Mice were immunized twice with the HEC V3 or with
the MN V3 peptide and T cell responses were measured
against HEC V3, MN V3, or RF V3 2 weeks and 2
months after the second immunization. We determined
the number of peptide-specific T helper cells using flow
cytometry by quantitating the number of CD41 T cells
that were also CD691 after 16 h of stimulation in vitro
with the indicated peptides. We assessed antigen specificity
by measuring expression of CD69 rather than
secretion of IFN-g. This marker of early T cell activation
was chosen because, in contrast to viral infections
that strongly polarize the T helper cell response toward
Th1 cytokine profile (and IFN-g secretion), we were not
sure that the majority of antigen-specific T cells expanded
by peptide immunization in oil-based adjuvant
would similarly result in a polarized Th1 response.
As expected, immunization of mice with the MN V3
peptide expanded a high frequency (mean frequency,
4.9%) of T cells present 2 weeks after the second immunization
that were specific for the MN V3 peptide
(Fig. 1A). This robust T cell response was not surprising,
given the known immunogenicity of this region of
the HIV-1 envelope protein. Despite the high frequency
of T cells specific for the MN V3 peptide, 10-fold fewer
T helper cells reacted with the RF V3 peptide (mean
frequency, 0.4%) or with related peptides contained
within the HEC V3 (mean frequency, 0.5%). By comparison,
whereas immunization of mice with the HEC
V3 expanded a lower frequency of T cell specific to itself
(mean frequency, 0.8%), there were comparable frequencies
of T cells specific for both the MN V3 and RF
V3 peptides (mean frequencies of 0.6 and 0.8%, respectively).
As measured 2 weeks after the secondary immunization,
there was not a statistically significant
quantitative difference in the ability of MN V3- vs HEC
V3-immunized mice to recognize the heterologous RF
V3 peptide. Noteworthy is the observation that only a
small fraction (approximately 10%) of the T cell repertoire
expanded with and specific to the MN V3 peptide
reacted with the RF peptide, whereas a comparable
number of T cells expanded with and specific to the
HEC V3 reacted with the RF peptide. These data suggest
that immunization with the MN V3 peptide expanded
a larger repertoire of T cells with a more limited
breadth of reactivity, whereas immunization with
the HEC V3 elicited a smaller repertoire of T cells that
possessed a greater breadth of reactivity.
The high frequency of T cells specific for MN V3 in
MN V3-immunized mice was rapidly lost over the
course of 6 weeks (Fig. 1A). There was an approximately
10-fold reduction in the number of T cells
present 2 weeks vs 2 months after immunization
(mean frequencies of 4.9 vs 0.4%, P , .001). The
modest reactivity to the RF V3 peptide present at 2
weeks declined approximately 2-fold (mean frequencies
of 0.4 vs 0.2%). In contrast, the frequencies of T
cells specific for the HEC V3 or the MN V3 or RF V3
peptides were statistically comparable 2 weeks and 2
months after immunization with the HEC V3 (0.8 vs
1.5%, 0.6 vs 1.2%, 0.8 vs 1.2%, respectively, Fig. 1B).
Indeed, 2 months after the secondary immunization
there were statistically greater frequencies of T helper
cells specific for the MN V3 and RF V3 peptides in the
FIG. 1. Primary immunization with a mixture of peptides expanded
a more broadly reactive and durable CD41 T cell repertoire
than did immunization with a single peptide. Mice were immunized
twice with a single peptide (A) corresponding to the HIV-1MN sequence
of the V3 loop of HIV-1 (MN V3) or with a mixture of peptides
(B) based on the V3 loop of HIV-1 (HEC V3). Two weeks or two
months after the second immunization, splenocytes and lymph node
cells were stimulated in vitro with the HEC V3, MN V3, or RF V3
antigens. The total frequency of CD41CD691 T cells present in each
mouse is depicted as a filled circle. A horizontal line represents the
mean frequency of antigen-reactive T cells present in each group.
The experiments were repeated twice, and the combined results are
depicted. Ten mice immunized with MN V3 were analyzed at 2 weeks
and 17 mice at 2 months; 9 mice immunized with HEC V3 were
analyzed at 2 weeks and 11 mice at 2 months.
154 ANDERSON ET AL.
HEC-immunized mice (P , .05 and P , .01, respectively).
These data confirm that immunization with the
HEC V3 expanded a repertoire of T helper cells with
broader reactivity than did immunization with a single
peptide. Moreover, they indicate that the memory response
elicited by the HEC V3 was more durable than
that expanded by the MN V3 peptide.
T Cells Expanded with a HEC Recognize a Naturally
Processed Viral Epitope
We wanted to demonstrate that the broad T cell
reactivity elicited by immunization with the HEC
based on the hypervariable V3 loop of HIV-1 held true
for other hypervariable epitopes as well. Moreover, we
wanted to determine whether T cells expanded with
HEC immunization could recognize naturally processed
viral antigen. To address these issues, we immunized
mice with a HEC based on a hypervariable
epitope of the V4 loop of SIV (HEC V4). For comparison,
we used a peptide based on the V4 loop epitope
derived from the SIVmac239 strain of virus, V4 (239).
Two weeks after secondary immunizations with the
HEC V4 or the V4 (239) peptide, we evaluated the
frequency of T cell specific for the V4 (239) peptide, a
related heterologous peptide, V4 (142), and for naturally
processed peptide derived from whole inactivated
SIVmac251 virus.
As expected, the V4 (239) peptide induced a greater
frequency of T cell specific for itself (4.2%) than did the
HEC V4 (1.8%) (Fig. 2). However, consistent with our
previous results, a greater frequency of T cells present
in HEC V4-immunized mice recognized the V4 (142)
peptide (2.0%) than in V4 (239)-immunized mice (0.7%)
(P , .05). Most importantly, the HEC-immunized
mice on average contained 10-fold greater numbers of
T cells that could recognize naturally processed SIVmac251
peptide antigen (2.3%) than did mice immunized
with a single peptide (0.3%) (P , .05). Thus,
comparable results were obtained in two different viral
systems based on two different epitopes.
DISCUSSION
Based on cloning of rearranged T cell receptor genes
in primary and secondary responses, repeated protein
immunizations were found to limit the diversity of the
responding T cell repertoire (19). These results suggest
that not all reactive T cell clonotypes present to respond
in a primary response are ultimately involved in
memory responses. Moreover, they imply that a less
diverse memory repertoire is likely composed of T cells
that are less cross-reactive. Similarly, studies of numerous
viral infections of humans and mice have demonstrated
that CTL responses to given viral epitopes
can be very focused (20, 21).
These data can help explain the phenomena of original
antigenic sin that was originally described in the
1950s. While characterizing the antibody responses of
individuals vaccinated with an influenza vaccine,
many vaccinated individuals subsequently became infected
with a heterologous strain of influenza. To the
surprise of immunologists studying the humoral responses
to influenza in these patients, exposure to the
second heterologous virus boosted immunity against
the first (vaccine) strain of the virus, with only modest
responses directed against the heterologous virus. This
phenomena was termed original antigenic sin and was
demonstrated in both humans and animals (22, 23).
More recently, Klenerman and Zinkernagel and colleagues
have also demonstrated the phenomenon of
original antigenic sin using lymphocytic choriomeningitis
virus (LCMV) infection of mice (24). The authors
examined secondary (memory) CTL responses to
LCMV after infection with a variant LCMV. Secondary
infection of mice with a variant virus elicited a response
to the parental virus (and its epitopes) with
which the mice were first infected; this held true for
various parental and variant viruses and their
epitopes, as well as for both dominant and subdominant
epitopes.
Original antigenic sin could represent a major obstacle
to the generation of efficacious HIV vaccines (25).
For example, the only candidate HIV vaccine in Phase
III clinical studies involves multiple immunizations
with a vaccine composed of recombinant envelope proteins
from only two strains of virus. It is not unreasonable
to predict that this vaccination approach will
serve to expand and focus B cell and T helper cell
repertoires specific to epitopes derived from these two
strains of virus. Given the tremendous number of HIV
variants circulating throughout the human population,
FIG. 2. Immunization with a mixture of peptides expands a
broadly reactive repertoire of CD41 T cells that can recognize naturally
processed viral antigen. Mice were immunized twice with either
a single peptide based on an epitope in the hypervariable V4 loop of
the SIVmac239 strain of virus (V4 (239)) or with a HEC based on this
epitope (HEC V4). The frequency of antigen-specific CD41 T cells
present 2 weeks after the second immunization for each mouse is
depicted as a filled circle; the mean frequency is depicted as a
horizontal line.
OVERCOMING ORIGINAL (ANTIGENIC) SIN 155
however, it is unlikely that patients will encounter
vaccine strains of virus. If original antigenic sin is
operational in these vaccinated individuals, it may significantly
delay or preclude antibody and T helper cell
responses elicited against infecting, heterologous
strains of HIV.
Our data demonstrate that immunization with a single
peptide (MN V3 or V4 (239)) expands a higher
frequency of T cells specific for that peptide than does
immunization with a cocktail of related peptide variants
(HEC V3 or HEC V4). However, the T cell repertoire
expanded with a single peptide contained relatively
few T helper cells capable of recognizing related
heterologous peptides compared with the repertoire
expanded with a HEC. These data demonstrate that
when the immune system is primed with a large array
of epitope variants, it expands a broadly reactive T
helper cell repertoire. Consistent with these results, we
have previously characterized antibody responses with
similarly broad reactivities in nonhuman primates immunized
with HIV- and SIV-based HECs (17, 18).
Our data also demonstrate that T cells expanded
with the HEC V3 with specificity for related singlepeptide
epitopes (MN V3 or RF V3) persisted longer in
vivo than did T cells expanded with the MN V3 peptide.
At present we are unsure how to explain the enhanced
durability of the T helper cell memory response expanded
by the HEC V3. Based on murine CTL responses
to viral infections in vivo, it has been suggested
that the size of the final memory population is
proportional to the clonal burst size of the responding
CTL repertoire (26). These data would predict that
greater frequencies of T helper cell should be detected
2 months after the secondary immunization with the
MN V3 peptide because it would have expanded a
greater frequency of T cells 2 weeks after the secondary
immunization. Our data, however, do not support this
prediction. One explanation may be that the broadly
reactive T cell repertoire expanded by the HEC V3 can
be stimulated and thus maintained more readily in
vivo because individual T cells can respond to low
levels of multiple different endogenous peptides
present on the surface of antigen-presenting cells in
vivo.
Ultimately, these data imply that vaccines that seek
to expand broadly reactive T helper cell (or antibody)
repertoires should do so by priming and expanding the
immune system with multiple peptide or recombinant
protein variants rather than with one or a few variants.
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Received August 14, 2001; accepted with revision August 22, 2001; published online October 8, 2001
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