Immune mechanisms in allergen-specific immunotherapy

Immune mechanisms in allergen-specific immunotherapy

CLINICAL IMMUNOLOGY AND IMMUNOPATHOLOGY Immune Mechanisms 53, s11%s131 (1989) in Allergen-Specific Immunotherapy’ Ross E. ROCKLIN Boehringer l...

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CLINICAL

IMMUNOLOGY

AND

IMMUNOPATHOLOGY

Immune Mechanisms

53, s11%s131 (1989)

in Allergen-Specific

Immunotherapy’

Ross E. ROCKLIN Boehringer lngelheim Pharmaceuticals,

Ridgefield, Connecticut 06877

Allergen-specific immunotherapy has been shown to be clinically effective in patients with seasonal allergic rhinitis and/or asthma. Patients who receive this therapy undergo a number of specific immunologic changes in response to the allergen being administered. These include a “blunting” of the seasonal rise of allergen-specific IgE as well as lowering baseline IgE levels, generation of an allergen-specific IgG response, development of auto-anti-idiotypic antibodies, reduced basophil histamine release in response to allergen, decreased lymphocyte proliferation, lymphokine production in response to allergen, and the generation of allergen-specific suppressor T cells that down-regulate lymphoproliferative responses and IgE synthesis. The mechanism by which allergenspecific immunotherapy produces clinical efficacy is not known. Recent evidence suggests that the development of immunoregulatory responses (suppressor T cells and antiidiotypic antibodies) during immunotherapy may account for the immunologic changes described above but as yet have not been correlated with clinical outcome. Identification of epitopes on allergens that can induce selective T helper/suppressor responses may provide opportunities for producing immunological tolerance and a reduction in the allergic diathesis. o 1989Academic Press. IIK.

INTRODUCTION

Since its introduction by Noon (1) in 1911, “allergic desensitization” (currently termed allergen-specific immunotherapy) has remained a valuable addition to the treatment of patients with severe allergic rhinoconjunctivitis and asthma who are unresponsive to more conservative modes of therapy. Conventional high-dose allergen-specific immunotherapy is administered subcutaneously in weekly increments up to a maximal maintenance dose and then given biweekly or monthly for several years. Identification of the offending allergens is based on skin testing and a carefully obtained clinical history. Whereas significant advances in our knowledge of the pathogenesis of atopic diseases have been recently made, little is known about the mechanism by which immunotherapy is beneficial to allergic patients. Clinical manifestations of hay fever, extrinsic asthma, and other types of immediate hypersensitivity diseases are mediated by IgE antibodies. When cellbound IgE interacts with the allergen, biologically active substances such as histamine, prostaglandins, and leukotrienes are released from membrane phospholipids and the granules of mast cells or basophils. These substances increase vascular permeability, smooth muscle constriction, and the recruitment of inflammatory cells and are generally responsible for the clinical manifestations of immediate hypersensitivity. The majority of studies involving allergen-specific immunotherapy have been designed to evaluate the efftcacy of such therapy, as well ’ Presented as part of a symposium entitled “Autoimmunity 1989, Scottsdale, AZ.

and Immunointervention,”

March 5-8,

s119 0090-1229/89 $1.50 Copyright All rights

0 1989 by Academic Press, Inc. of reproduction in any form reserved.

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as the immunologic alterations that occur in the treated patient. Well-controlled clinical trials have established the efficacy of immunotherapy in patients with allergic rhinitis, Hymenoptera venom hypersensitivity, and extrinsic asthma. In addition, the administration of high-dose immunotherapy is associated with a number of changes in humoral and cell-mediated immunity and which are summarized in Table 1 and discussed below. IgE ANTIBODIES

In early immunotherapy studies, the Prausnitz-Kustner reaction was used to assay changes in serum reaginic activity in allergic individuals receiving ragweed extract injections for periods varying from I to 20 years (2. 3). A significantly lower reaginic titer in treated patients compared to untreated allergic controls was noted. Subsequently, prospective studies observed a reduction in serum reaginic antibody titer in about half the subjects receiving immunotherapy and essentially no change in the serum of untreated patients (4). Moreover, it was noted that the period immediately following the initiation of immunotherapy was marked by a rise in reagin levels. Subsequently, an in vitro method for assaying reaginic activity of human serum based on histamine release from passively sensitized normal leukocytes was developed. The leukocytes were mixed with the serum of an allergic patient, washed, and then incubated with the relevant allergen. The quantity of histamine released was felt to reflect the reaginic titer of the allergic serum. Employing this technique, it was reported that untreated ragweed allergic patients exhibited a seasonal increase in reaginic activity from October to July, followed by a slow decline between seasons (5). Injection therapy with ragweed extract led to an attenuation of the seasonal rise in reaginic activity. The identification of immunoglobulin E as the principal reagin in man was quickly followed by the development of sensitive techniques for its assay (6, 7). Using the RAST, investigators in various laboratories have measured serum levels of IgE to given allergens during immunotherapy and have essentially confirmed the above findings obtained with the leukocyte histamine release and in vivo passive transfer methods. The major change induced by immunotherapy seems to be a dampening effect on the seasonal increase in IgE antibodies, which is a well-documented occurrence in untreated allergic individuals (8). This blunting of the seasonal rise even occurs in some treated patients who do not have a fall TABLE IMMUNOLOGICCHANGES

I

DURING ALLERGEN-SPECIFICIMMUNOTHERAPY

Blunted seasonal rise in allergen-specific IgE antibodies Increase in allergen-specific IgG antibodies Development of auto-anti-idiotypic antibodies Diminished basophil histamine release in some patients Reduction of lymphocyte proliferation and lymphokine production to the immunizing allergen Development of allergen-specific suppressor cells Reduction in the late phase response

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in the baseline levels of their allergen-specific IgE. However, documented decreases in the seasonal rise in the IgE antibody may not correlate with clinical improvement; in any one patient, clinical efficacy may be achieved with a decrease, no change, or an increase in the levels of IgE. While as a group baseline specific IgE levels (out of season) in treated patients have been shown to decline with time to below pretreatment levels, the response may vary in any one patient. Furthermore, total IgE levels do not change significantly during treatment unless the specific IgE levels directed against the particular allergen constitute the major portion of the total. IgG ANTIBODIES

Serum factors capable of abrogating the allergic response appear during immunotherapy. They have traditionally been termed “blocking factors” because of their ability to block the wheal and flare skin response in the Prausnitz-Kustner reaction when mixed with the antigen prior to its intracutaneous administration. Early studies showed that serum of treated allergic patients contained both a heat-sensitive skin-sensitizing reaginic activity and a heat-resistent blocking factor. The latter was not detected prior to the onset of immunotherapy. Subsequent investigations using similar passive transfer experiments confirmed these flndings. However, the techniques used in these early studies suffered from an unacceptable degree of variability and allowed only a semiquantitative evaluation of blocking antibodies. The development of a reliable technique for the study of in vitro histamine release permitted the indirect measurement of blocking antibodies by their ability to inhibit the allergen-induced release of histamine from sensitized leukocytes. Employing this method, it was reported that the blocking antibody titer rises dramatically during immunotherapy in patients with ragweed pollenosis (3). The magnitude of the increase in blocking antibodies was approximately proportional to the amount of ragweed injected. Within 6 to 12 months of the cessation of immunotherapy, the blocking antibody titer fell toward pretreatment levels. The increase in IgG antibodies correlates well with clinical outcome in groups of pollen- and venom-treated patients (9, 10). That blocking antibodies belong to the IgG class of immunoglobulins has been clearly established, initially be means of immunoprecipitation, and with the aid of highly sensitive and specific radioimmunoassays. Quantitative measurement of blocking antibodies during immunotherapy in a number of unrelated antigen systems has amply corroborated the concept that parenteral immunization stimulates the production of antigen-specific IgG antibodies, and that in the majority of patients, the levels attained are generally much higher than the serum concentrations prior to treatment. IgG subgroup data from a number of laboratories suggest that IgG4 is selectively increased over time in venom- and pollen-treated patients. The significance of this tinding, however, is not clear at the present time since some studies find a correlation of IgG4 levels with clinical efficacy while others do not (I 1). ANTI-IDIOTYPIC

It is generally

ANTIBODIES

accepted that idiotypic-anti-idiotypic

interactions

play both a

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qualitative and a quantitative role in the regulation of IgE antibody levels ( 12). Several recent studies (13-15) indicate that auto-anti-idiotypic antibodies can be detected in the serum of immunotherapy-treated patients and that untreated patients have much lower levels. Auto-anti-idiotypic antibodies could be detected in the serum of patients with allergic rhinitis after about 8 to 10 months of ragweed immunotherapy in one study (16) and 6 months (17) of therapy in another. Cross-reactive idiotypic antibodies recognized by hetero-antisera were detected in one study of rye grass-sensitive allergic patients (14). The anti-idiotypic antibodies that were isolated reacted with idiotypic antibody from 20 of 20 rye-allergic patients. In another study that prospectively followed seven ragweed-allergic patients, cross-reactive anti-idiotypic antibody levels increased continuously over a 3-year period following the onset of immunotherapy (17). These results are summarized in Fig. 1. If these anti-idiotypic antibodies were indeed playing a role in modulating the IgE immune response to an antigen during immunotherapy, then it should be possible to document a relationship between idiotypic and anti-idiotypic antibody levels in these patients. In fact, one study (17) demonstrated an inverse relationship between specific IgE and anti-idiotypic antibody levels in the nonatopic (low IgE Id, high anti-Id) and untreated (high IgE Id, low anti-Id) atopic groups (Fig. 2). The results of the immunotherapy-treated ragweed atopics were, however. 1.2 1.0

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Months Post Immunotherapy FIG. I. Prospective measurement of Ab2 levels in ragweed-allergic patients before and at varying times after ragweed immunotherapy was started (3-36 months). Ab2 levels increased severalfold (2-g) postimmunotherapy. (From Ref. (17) with permission.)

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RAST Ab 2 NON-ATOPIC

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RAST Ab 2 UNTREATED ATOPIC

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RAST

Ab2 TREATED ATOPIC

FIG. 2. RAST (ragweed-specific IgE) and Ab2 (measured by reactivity with SC7H.lG). The ordinant represents *SD about the mean for each variable, AB2 or RAST. (From Ref. (17) with permission.)

variable. In general, their IgE levels were decreased compared with untreated patients but they had significantly higher anti-idiotypic antibody levels. Thus, immunotherapy stimulates the production of anti-idiotypic antibodies in atopic patients and these may be involved in the regulation of the anti-ragweed IgE response. Further studies are necessary in order to determine whether the development of anti-idiotypic antibodies correlates with the clinical outcome in such patients. RESPONSIVENESS

OF MEDIATOR-RELEASING

CELLS

In pollen immunotherapy studies, several patients exhibited a dramatic decrease in leukocyte reactivity to ragweed (18, 19). Cellular reactivity was defined as the maximal mediator-releasing capacity of peripheral blood leukocytes when challenged with an optimal concentration of allergen. The sera of those patients who had a decrease in cellular reactivity could still sensitize normal leukocytes to release most of their histamine upon exposure to the allergen, implying that the alteration in cellular reactivity was not the result of a decrease in serum reaginic antibodies. In general, leukocytes from approximately 30% of patients on immunotherapy will exhibit decreased cell sensitivity. Although a decrease in reactivity of mediator-releasing cells during immunotherapy is an appealing concept to explain its mechanism of action, it does not appear to be a common occurrence and the cellular basis for this hyporesponsiveness is not clear at present.

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LATE PHASE RESPONSE

Some investigators have recently suggested that the clinical efficacy of immunotherapy is related in part to its ability to modify the late phase response (20). They demonstrated that patients on immunotherapy would often lose their late phase skin reaction to specific allergen. A significant inverse correlation between the level of allergen-specific IgG and the magnitude of the late phase response was also noted. Untreated atopics had low levels of allergen-specific IgG and a 94% incidence of late phase reactions, whereas immunotherapy-treated patients had high levels of allergen-specific IgG and a less than 50% incidence of late phase reactions. Behrens et al. (21) have demonstrated that infused allergen-specific IgG could modulate the appearance of the late phase response following challenge in a rabbit model. The chronicity and nonspecific airways hyperreactivity of such conditions as asthma and rhinitis suggest that their immunopathogenesis may be more closely related to the late phase rather than the acute phase reaction. The ability of immunotherapy to down-regulate the late phase reaction is therefore of great interest and may provide new insights into the mechanism by which immunotherapy is effective for management of these conditions. Further studies will require an in-depth analysis of an even greater collection of mast cell/basophil mediators (histamine, kinins, PGD,, proteases) and the kinetics of the cellular infiltrate (basophils, mast cells, eosinophils, neutrophils, and mononuclear leukocytes) associated with this reaction. LYMPHOCYTE

FUNCTIONS

Several studies have examined the effects of conventional immunotherapy on the in vitro responses of T cells isolated from atopic patients. Previous work showed that mononuclear cells from patients undergoing immunotherapy became less responsive in vitro to the allergen, in that lymphocyte proliferation and lymphokine production was reduced (22). The lymphokines measured at that time included migration inhibitory factor (MIF) and lymphocyte mitogenic factor. More recently, interleukin 2 (IL-2) production was measured in patients undergoing immunotherapy (23). Data from this study indicated the IL-2 production in response to mite antigen was increased in untreated asthmatics. In contrast, IL-2 production in response to phytohemagglutinin (PHA) was indistinguishable from that of control subjects. When the patients underwent immunotherapy to dust antigen, IL-2 production was reduced to the levels found in normal subjects. Furthermore, it was noted that the response to IL-2 itself was increased in immunotherapy-treated patients compared with normal or with untreated patients. This was true whether the responder cells were stimulated with PHA or dust antigen. An extension of this work analyzed the proliferative responses of purified T lymphocyte subpopulations during immunotherapy (24). Utilizing purified populations of helper/inducer (T4 + or CD4) and suppressor/cytotoxic (T8 + or CD8) cells, distinct responses of CD4+ T cells to PHA, which were reduced before immunotherapy, increased to normal levels in mite-treated patients. In contrast,

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the CD4+ T cell response to mite antigen was decreased. The proliferative response of the CD8 + T cell population to PHA increased after immunotherapy and this was accompanied by increased proliferation to mite antigen. Studies such as the ones described above have been limited by the use of relatively poorly defined antigens, such as dust or ragweed AgE, for which the primary amino acid structures are unknown. In experimental animals, a number of well-defined antigens have been studied and in some murine systems the actual antigenic epitope or peptide fragment has been documented to bind directly to the appropriate class II MHC (25). Recent studies (26) have examined human T cell recognition of antigen by means of a model that employs mouse allergen-l (MAI), a major allergen found in mouse urine. Mononuclear cells isolated from subjects with a history of allergy to laboratory mice were found to make a significant T cell proliferative response to MA-l. Allergen-specific MHC-restricted human T cell clones were then generated by limiting dilution from lines derived from blood T cells from these individuals. The clones that were isolated and found to respond specifically to MA-l also exhibited cross-reactivity with rat a 2u globulin (RA2uG), a related family of rat liver proteins. Antigen-presenting cells (APCs) were necessary for both the activation of these T cells by allergen and their long-term in vitro growth. When MA-l and irradiated APCs were added to the cultures, significant amounts of IL-2 were liberated into the supernatants with peak IL-2 levels detected 24 hr before maximum T cell proliferation. Most of the clones were phenotyped as being CD3 + CD4 + and served in a helper T cell capacity to increase allergen-specific IgG production by donor B cells. In order to identify the relevant antigenic epitope being recognized by the T cells, the above workers utilized a cDNA probe for RA2uG that was inserted into an SP6 plasmid. They were able to obtain whole length and restriction endonuclease-generated fragments of RA2uG message. Peptides generated in this way were used to obtain preliminary mapping data (27) which have demonstrated a single dominant epitope for T cell activation regardless of the source of HLA specificity of the T cell clone and which was located in the region between amino acids 80 and 110 of the RA2uG primary structure. This process of epitope mapping has important and potentially exciting clinical application in that it may allow the construction of fragments of allergen that are altered slightly to a “tolerogen” configuration. These may be capable of blocking the T cell response to native protein and could herald the development of a new form of immunotherapy for allergic disease. IMMUNE

MECHANISMS

IN ANIMALS

In recent years, significant advances have been made in the understanding of IgE regulation in experimental animals (28). A review of some of these findings may shed light on the immunologic aberrations associated with the atopic state in man and their modification by immunotherapy. Manipulations of the immune response in animals which strive to suppress the IgE antibody system have followed two main pathways. The first approach consists of modifying the production of IgE directed against specific antigens, and the second approach aims at selectively suppressing the IgE class of immunoglobulins by an antibody directed

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against the interleukin which induces its production. Both approaches offer areas of promise for future applications in the management of clinical allergies (Table 2). An experimental system that mimics allergic desensitization in ragweed hay fever was previously developed (reviewed in Ref. (7)). The cellular interactions involved in the regulation of the IgE response against ragweed antigen E in a high-responder strain of mice were then analyzed. After inducing a state of persistent IgE production, the animals were treated with weekly injections of native or urea-denatured ragweed antigen E. The mice receiving native ragweed antigen E did not manifest a significant change in ragweed-specific IgE titer but had an increase in ragweed-specific IgG levels. Those receiving urea-denatured ragweed antigen E showed a reduction in antigen-specific IgE antibody titer with no change in antigen-specific IgG antibody titer. Both regimens were effective in suppressing the IgE response to ragweed antigen E. Using adoptive transfer techniques, it was found that the blunting of the secondary IgE response in mice treated with either regimen resulted from reduced T-helper activity for IgE production. The above findings suggest that immunotherapy is accompanied by diminished T-helper activity for IgE antibody production and that use of modified allergens may be a more effective means of desensitization since it achieves a greater reduction of the ongoing IgE antibody response. Subsequently, it was also found that suppressor T cells were more effective in abrogating the IgE than the IgG response to ragweed antigen when the animals were given large doses of ragweed without adjuvant. It was postulated that the induction of antigen-specific suppressor cells may be an important aspect of the beneficial effect of immunotherapy. Much attention has recently been focused on the ability of lymphokines secreted by helper T cells to induce or suppress IgE synthesis. Key among these has been interleukin 4 (IL-4). a 20,000 MW lymphokine originally termed B cell stimulatory factor (BSF-I) because of its ability to promote the proliferation of antiIgM-stimulated B lymphocytes (29). Initial studies showed that IL-4 selectively enhanced in vitro IgGl and IgE production and suppressed IgG3 and IgG2b secretion by murine B cells stimulated by LPS (30-32). These in vitro studies were extended to show that IL-4 is involved in IgE production in vivo. Mice infected with helminths were found to have an loo-fold increase in serum IgE concentrations within approximately 2 weeks following innoculation with the larvae of N. TABLE EXPERIMENTAL

METHODS

FOR SUPPRESSION

2

OF THE

IgE

RESPONSE

Modified antigens Urea-denatured ragweed antigen E Dinitrophenyl-D glutamic acid (DGL) DNP-polyvinyl alcohol Ovalbumin-DGL Ovalbumin-polyethyleneglycol Anti-idiotypic antibodies IgE-specific soluble suppressor factors Anti-IL-4 antibodies

IN ANIMAL

MODELS

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brasiliensis. That this marked increase in IgE levels was in fact due to IL-4 was demonstrated indirectly by the administration of anti-IL-4, which blunted this rise (33). This finding raises the possibility that anti-IL-4 could be used in vivo as a means to down-regulate IgE production. The finding (34) that IL-4 controls the expression of several isotypes with strikingly different dose-response and time course relationships suggests that agents that interfere with IL-4 secretion, that block its binding to its cellular receptor, or that, like interferon-y, counteract its effects on the B cell may provide means to diminish the allergy-producing effects of antigenic stimulation without compromising other isotype responses. IMMUNE MECHANISMS

IN MAN

Although clinical trials have documented the efficacy of immunotherapy in allergic diseases, studies to unravel the underlying mechanisms are still at an early stage. A number of hypotheses have been formulated, but none of them has been experimentally established to be the major mode of action of immunotherapy (Table 3). A widely held concept is that immunotherapy favors the production of IgG-blocking antibodies over IgE reaginic antibodies. When present in excess, IgG antibodies would interact with the allergen and make it unavailable to IgE antibodies on the surface of mediator-releasing cells, thus interrupting the initial triggering event of the allergic response. Ample evidence has been provided that adequate immunization with ragweed, Hymenoptera venom, and cat allergen extracts is associated with a striking increase in antigen-specific IgG antibodies in most allergic subjects. Whether this relationship is causal or a mere statistical association remains to be determined. In the individual patient, it is not always possible to predict the degree of clinical improvement from measurements of the rise in IgG antibodies. Furthermore, direct evidence for the protective effect of IgG antibodies has been presented only in the insect venom system. Hence, it seems reasonable to conclude that IgG antibodies may contribute to an amelioration of symptoms but do not constitute the only mode of action of immunotherapy. The possibility that immunotherapy induces the production of protective secretory antibodies has been raised. However, there has not been a consistent correlation between the detection of IgG antibodies in nasal secretions and relief of clinical symptoms. Although secretory IgA and IgG antibodies are increased durTABLE PROPOSEDIMMUNE

MECHANISMS

3 IN ALLERGY DESENSITIZATION

Development of IgG “blocking” antibodies Induction of tolerance in IgE-producing B cells Impairment of T-cell help Induction of allergen-specific and/or isotype-specific suppressor factors and/or suppressor cells IgE regulation by auto-anti-idiotypes Decreased reactivity of mediator-releasing cells

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ing immunotherapy, there is no compelling reason to believe that local antibodies bear a causative relationship to the benefits derived from immunotherapy. On the basis of evidence from animal experiments and the initial studies reported in man concerning the generation of ragweed-specific suppressor cells capable of inhibiting lymphocyte proliferation, it has been postulated that a major pathogenetic mechanism of immunotherapy may be the induction of mononuclear suppressor cells. In view of the T cell dependence for the regulation of LgE synthesis, overproduction of specific IgE observed in atopic disease might be a consequence of altered regulatory function. It has been reported that atopic individuals have reduced numbers of T cells with surface membrane receptors for the Fc portion of IgG, and that after successful immunotherapy their numbers reach normal levels (3.5). Cells within the latter subpopulation are known to regulate immunoglobulin synthesis by suppressing T-dependent B cell differentiation. Further, T lymphocytes bearing histamine type-2 receptors and expressing suppressor activity after stimulation with histamine also possess Fc receptors for IgG (36). It is possible, then, that the observed abnormalities in atopic subjects reflect in vivo deficiency in antigen-nonspecific and/or antigen-specific suppressor cell activity, either or both of which may be necessary for the dampening of IgE antibody production. Evidence for this hypothesis has been directly provided by a study that showed that ragweed antigen-specific suppressor T lymphocytes could be detected in ragweed-allergic individuals during immunotherapy for 6 to 12 months, but not prior to therapy (37). When lymphocytes were isolated from treated patients and passed over columns containing insolubilized histamine, antigen-specific suppressor cells that could be activated by ragweed antigen were deleted. These results suggested that antigen-specific suppressor T cells belonging to the subpopulation of lymphocytes bearing histamine receptors were generated during immunotherapy and failure to detect these cells in untreated patients may be a reflection of the underlying defect leading to the atopic diathesis. More recent studies using a different antigen system have confirmed the finding that immunotherapy results in the detection of a suppressor cell population that can recognize a specific allergen (38). In a group of rye grass atopics, it was reported that immunotherapy induced a population of T cells which adhered to allergen-coated plates that were capable of suppressing proliferation to the specific allergen in the responding population. This suppression was reversed by pretreatment of the adherent T cells with anti-CD8 plus complement. The detection of suppressor cells during immunotherapy was recently extended to IgE regulation. It was shown that immunotherapy induced & nova generation of suppressor cells or increased their number and/or activity (39). In this prospective study, five ragweed-allergic individuals were followed before and during immunotherapy. Depletion of CDS+ cells resulted in enhancement of in vitro IgE production once these patients were on immunotherapy for 1 year, whereas prior to immunotherapy no effect was seen. Furthermore, a suppressive effect of antigen was also observed. The addition of ragweed antigen E in vitro to the cultures did not alter IgE synthesis before immunotherapy. but resulted in a decrease in IgE production when the patients’ cells were examined I year after starting immunotherapy. This suppression of IgE synthesis by ragweed antigen E could be

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abolished by eliminating CD8 + cells from the culture. In contrast to its effect on IgE, the addition of ragweed antigen in vitro resulted in augmentation of the anti-ragweed IgG response. Thus, the ability of immunotherapy to induce/ augment suppressor cell activity may be linked to its overall effectiveness clinically, but these correlations require prospective long-term controlled trials to make this determination. Changes in serum IgE concentration do not correlate with clinical improvement during immunotherapy. Nonetheless, immunotherapy suppresses the seasonal increase in allergen-specific IgE antibodies which occurs in untreated subjects. More detailed investigations of the IgE system are needed to assess the effect of immunotherapy on this important aspect of the allergic response. It is hoped that the recently developed in vitro models for the study of IgE biosynthesis by peripheral blood lymphocytes utilizing IL-4 will shed further light on the mechanism underlying immunotherapy. SUMMARY

Multiple factors are probably responsible for the efficacy of immunotherapy. An enhanced T-cell suppressor activity may act in concert with increased antigenspecific IgG antibody in order to block various steps of the allergic response, whereas a diminution in IgE antibody against the offending allergen would eliminate an essential step in the sequence of events leading to the release of inflammatory mediators of immediate hypersensitivity. The modification of allergens through denaturization so that they are less reactive with preformed IgE has resulted in potent preparations that induce a strong IgG response. These preparations permit flexibility in dosage and in frequency of administration as well as produce no greater side effects than aqueous extracts. While these preparations would have a decided advantage over existing extracts, recent information that is accumulating regarding T cell recognition of antigen opens the way for more precise targeting of therapy. That is, determination of the antigenic epitopes that trigger helper T cells should permit the construction of peptides that can “block” T cell recognition and potentially lead to immunologic unresponsiveness. The tolerization of helper T cells that are involved in IgE synthesis over time would theoretically lead to decreased IgE production against the specific allergen and ultimately to a reduction in sensitivity and symptoms. Further, since the antigenic epitopes that are recognized by the T cell receptor for antigen and by the B cell receptor are very often different, then the administration of these peptides may not result in an interaction with preformed IgE that leads to mast cell triggering. Thus, while being potentially more safe and effective than existing antigen preparations, these peptides might also have more selective effects on the immune response to allergen, permitting some responses (IgG) while inhibiting others (IgE). REFERENCES I. Noon, L., Prophylactic inoculation for hay fever. Lancer 1, 1572, 1911. 2. Sherman, W. B.. Stull. A., and Cooke, R. A., Serologic changes in hayfever cases treated over a period of years. J. Allergy 11, 225, 1940.

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3. Loveless, M. H., Immunological studies in pollenosis. 1. The presence of two antibodies related to the same pollen antigen in the serum of treated hay fever patients. J. Imm~nol, 38, 25, 1940. 4. Norman. P. S., Winkenwerder. W. L.. and Lichtenstein. L. M., Immunotherapy of hay fever with ragweed antigen E: Comparisons with whole pollen extract and placebo. .I. Allergy 42. 93 1968. 5. Levy, D. A., Lichtenstein. L. M.. Goldstein. E. 0.. and Ishizaka. K., Immunologic and cellular changes accompanying the therapy of pollen allergy. J. C/in. Invesf. 50, 360, 1971. 6. Gleich, G. I., Jacob, G. L.. Yunginger, .I. W., and Henderson, L. L., Measurement of the absolute levels of IgE antibodies in patients with ragweed hay fever. J. Allergy C/in. Immune/. 60. 188, 1977. 7. Ishizakd, K., and Ishizaka, T.. Mechanisms of reaginic hypersensitivity and IgE antibody response. Immunol. Rev. 41, 60. 1978. 8. Lichtenstein, L. M., lshizakd. K., Norman, P. S., et ~1.. IgE antibody measurements in ragweed hayfever: Relationships to clinical severity and the results of immunotherapy. J. C/in. /rives/. 52, 210. 1973. 9. Starr, M. S., and Weinstock, M.. Studies of pollen allergy. III. The relationship between blocking antibody levels and symptomatic relief following hyposensitization with alpyral in hayfever subjects. Int. Arch. Allergy Appl. Immunol. 38, 514, 1970. IO. Hunt, K. J., Valentine, M. D., Sobotka. A. K., er al.. A controlled trial of immunotherapy in insect hypersensitivity. N. Engl. J. Med. 299, 157. 1978. Il. Nakagawa. T., The role of IgG subclass antibodies in allergic reactions. Immunol. Allergy Pmt. 9, 453. 1987. 12. Blaser. K.. Nakagawa. T., and de Week, A., Investigation of a syngeneic murine model for the study of IgE antibody regulation with isologous anti-idiotypic antibodies. Znr. Arch. AIlergy Appl. Immunol. 64, 42, 1981. 13. Mally. A., and Dresser. D. W.. Anti-idiotypic regulation of timothy grass pollen IgE formation. Immunology 46, 653. 1982. 14. Bose, R., Marsh, D. G., Duchateau, J.. et al.. Demonstration of auto-anti-idiotypic antibody cross reacting with public idiotypic determinants in the serum of rye-sensitive allergic patients. J. Immunol. 135, 2475, 1984. 15. Castracane. J. M., and Rocklin, R. E.. Detection of human auto-anti-idiotypic antibodies (AB2). I. Isolation and characterization of AB2 in the serum of a ragweed immunotherapy-treated patient. Int. .4rch. Allergy C/in. Immunol. 86, 288, 1988. 16. Lebenger, K., Wang. V.. Kin. Y., et al., Kinetics of the auto-anti-idiotypic (anti-anti-ragweed) antibody response to antigen E during ragweed immunotherapy in humans. Clin. Res. 33, 51SA, 1985. [Abstract] 17. Castracane. J. M., and Rocklin, R. E.. Detection of human auto-anti-idiotypic antibodies (AB2). II. Generation of AB2 in atopic patients undergoing allergen immunotherapy. Int. Arch, Allergy Clin.

Immunol.

86, 295. 1988.

18. Lichtenstein. L. M., Norman, P. S., Osler, A. G.. et ul., In vitro studies of human ragweed allergy: Changes in cellular and humoral activity associated with specific desensitization. J. C/in. Invest. 45, 1126, 1966. 19. Melam. H., Pruzansky, J. J., and Patterson, R., Correlations between clinical symptoms, leukocyte sensitivity, antigen binding capacity and P-K activity in the longitudinal study of ragweed pollinosis. J. Allergy 46, 292. 1970. 20. Pienkowski, M.. Norman, P. S.. and Lichtenstein, L. M.. Suppression of late phase skin reactions by immunotherapy with ragweed extract. J. Allergy C/in. Zmmunol. 76, 729. 1985. 21. Behrens, B.. Clark. R.. March, W., er al., Modulation of the late asthmatic response by antigenspecific immunoglobulin G in an animal model. Amer. Rev. Respir. Dis. 130, 1134, 1984. 22. Evans, R., Pence. H., Kaplan, H., and Rocklin, R. E.. The effect of immunotherapy on humoral and cellular responses in ragweed hay fever. J. C/in. Invest. 57, 1378. 1976. 23. Hsieh, K., Altered IL-2 production and responsiveness after hyposensitization to house dust. J. Allergy C/in. Immunol. 76, 188, 1985. 24. Hsieh, K.. Changes of lymphoproliferative responses of T cell subsets to allergen and mitogen after hyposensitization in asthmatic children. J. Allergy C/in. Immunol. 74, 34. 1984.

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25. Guillet, J., Lai. M., Briner, S., et nl., The relation between MHC restriction and the capacity of Ia to bind immunogenic peptides. Science 235, 1353, 1987. 26. Gurka, G., Ohman, J.. and Rosenwasser, L.. Human T cell clones specific for a mouse urinary protein allergen. J. Allergy C/in. Immunol. 79, 169, 1987. [Abstract] 27. Gurka, G., Kalluri. S., Rosenwasser. L.. et al., Allergen-specific T cell clones can be utilized to MAP allergenic epitopes for T cell activation. J. Allergy C/in. Immunol. 81, 195. 1988. [Abstract] 28. Ishizaka. K., IgE-binding factors and regulation of the IgE antibody response. Annu. Rev. Immud. 6, 513, 1988. 29. Howard, M., Farrar, J., Hiltiker, M., Johnson, B., Takasu, K., Hamaoka, T., and Paul, W. E., Identification of a T cell-derived B cell growth factor distinct from interleukin 2. J. Exp. Med. 155, 914, 1982. 30. Isakoson, P. C., Pure, E.. Vitetta, E. S., and Krammer, P. H., T cell-derived B cell differentiation factor(s): Effect on the isotype switch of murine B cells. J. Exp. Med. 155, 734, 1982. 31. Coffman, R. L., and Carty, J.. A T cell activity that enhances polyclonal IgE production and its inhibition by interferon-gamma. J. Immunol. 136, 949, 1986. 32. Coffman, R. L., Ohara, J., Bond. M. W., Carty, J., Zlotnick, E.. and Paul, W. E.. B cell stimulatory factor-l enhances the IgE response of lipopolysaccharide-activated B cells. J. Immunol. 136, 4538, 1986. 33. Finkelman, F. D., Katona, 1. M.. Urban, J. F., Jr., Snapper, C. M., Ohara, J.. and Paul, W. E., Suppression of in vivo polyclonal IgE responses by monoclonal antibody to the lymphokine B-cell stimulatory factor 1. Proc. Natl. Acad. Sci. USA 83, 9675. 1986. 34. Snapper, C. M., Finkelman, F. D., and Paul, W. E.. Differential regulation of IgGl and IgE synthesis by interleukin 4. J. Exp. Med. 167, 183, 1988. 35. Canonica, G. W., Mingari. M. C., Melioli, G., ef al., Imbalances of T-cell subpopulations in patients with atopic diseases and effect of specific immunotherapy. .I. Immunol. 1213,2669, 1979. 36. Beer, D. J., Osband, M. E.. McCaffrey, R. P., et al.. Non-specific suppressor cell function in atopic individuals. N. Engl. J. Med. 306, 454, 1982. 37. Rocklin, R. E.. Sheffer, A. L., Greineder. D. K.. and Melmon, K. L., Generation of antigenspecific suppressor cells during allergy desensitization. N. Engl. J. Med. 302, 1213. 1980. 38. Nagaya. H., Induction of antigen-specific suppressor cells in patients with hay fever receiving immunotherapy. J. Allergy Clin. Immunol. 75, 388, 1985. 39. Tamir. R.. Castracane, J. M., and Rocklin, R. E., Generation of suppressor cells in atopic patients during immunotherapy that modulates IgE synthesis. J. Allergy Clin. Immunol. 79, 591, 1987. Received June 19, 1989; accepted July 13, 1989