Molecular Aspects of Allergen-Specific Immunotherapy in Patients with Seasonal Allergic Rhinitis

A systematic review and narrative synthesis of publications was undertaken to analyze the role of component-resolved diagnosis technology in identifying polysensitization for the provision of allergen-specific immunotherapy to patients with seasonal allergic rhinitis. A search of publications was carried out in electronic databases in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The search helped to identify 568 publications, 12 of which were included in this review. Overall, 3302 patients were enrolled. The major finding was that component-resolved diagnostics change the choice of relevant allergens for allergen-specific immunotherapy in at least 50% of cases. Sensitization to allergen components differs with age, type of disease, and overall disease duration. Patients who had both bronchial asthma and allergic rhinitis were sensitized to a larger number of allergens than patients who had bronchial asthma alone.


Introduction
Currently, seasonal allergic rhinitis (AR) presents a global health problem [1]. According to statistics from the World Health Organization (WHO), allergic disorders affect approximately 40% of the global population [2,3] and their incidence continues to grow with increasing disease severity and expanding the spectrum of sensitization to unrelated allergens [4].
Seasonal allergic rhinitis is one of the biggest challenges among all seasonal allergic disorders [3], and it currently affects over 500 million people worldwide [5,6]. According to estimates, AR is observed in 23-30% of Europe's population, in 12-30% of the population of the USA, and 17-35% of the adult population in Russia [7][8][9]. The patients often present allergic reactions to plant food allergens, which might not be cross reactive, making the diagnostics even more complicated [10].
Pollinosis is perhaps the most common seasonal allergic disorder and is prevalent in approximately 30% of the world's population [11]. Most frequently, pollinosis manifests a consensus document devoted to molecular diagnostics with an exceptional focus on CRD as a tool for practicing allergists [52,53]. CRD has proven to be a reliable diagnostic instrument that is used for distinguishing between allergens that cause clinically significant allergic rhinitis and allergens associated with symptomless sensitization [54].
Sensitization to the allergen does not always witness allergy activity and severity, and thus, elucidating the causal allergen CRD is used, which helps to identify the major and minor allergens. When significant sensitization to major allergens is identified, one can conclude about the importance of this allergen and allergy severity [55].
The composition of causal allergens may change over time [56]. Since the immune response diminishes with aging, many patients note a reduction in the severity of allergy symptoms. However, sensitization to new allergens may occur, and the sensitization profile may expand due to the presence of comorbidities or a long duration of the allergy [57]. Nevertheless, it is worth noting that ASIT may prevent the appearance of new sensitization [58].
CRD determines objective criteria for the initiation of ASIT and helps to predict whether it will be effective [46]. Because ASIT is a long-lasting and resource-consuming treatment approach, precise diagnostics, appropriate selection of patients, and identification of major sensitizing allergens are needed to optimize treatment planning, including financial considerations [59,60]. Most patients receiving ASIT have elevated serum levels of allergen-specific IgE within the first few months after treatment initiation [61], which has no consequences in terms of recurrence or aggravation and presents a natural response to ASIT [62]. Progressive reduction of allergen-specific IgE is commonly observed after 6-12 months and corresponds with a decrease in the numbers of residential IgE-secreting long-lived plasma cells in the bone marrow [63]. Two weeks following the initiation of ASIT are accompanied by elevated serum levels of allergen-specific IgG4 and by the secretion of Il-10-producing regulatory B-cells [19,37]. The levels of IgG4 in blood serum remain elevated for more than a year [61,64]. The allergen-specific IgA produced during SLIT may be the source of highly effective blocking antibodies on the surface of mucous membranes [65]. Such blocking antibodies inhibit allergen-IgE binding, which leads to decreased basophil histamine release [61].
A relationship between specific IgE and the effectiveness of ASIT may be attributed to specific IgG, which is the marker of ASIT effectiveness. Shamji and coworkers reported a close correlation between IgG levels and the clinical response to ASIT. The levels of specific IgE and IgG were interrelated [66]. In general, specific IgE is used to monitor the effectiveness of immunotherapy, as its reduction reflects the development of immune tolerance [67]. IgE serum levels decrease and are maintained at low levels for years following successful ASIT [39,40]. Thus, it appears reasonable to suppose that the higher the IgE levels are, the more clinically significant the allergen for an individual patient [65], and the more causative allergens are used for ASIT, the more effective the results will be [68]. For this reason, baseline-specific IgE levels to allergens could serve as reliable biomarkers of effectiveness in patients with allergic rhinitis [23,69].
Thus, it is essential to understand what predictors impact the effectiveness of ASIT [70], and a careful assessment of allergen profiles with the identification of major and crossreactive allergens is needed [71]. Component-resolved diagnosis (CRD) could help to solve this issue [25,26]. This review analyzes the role of molecular allergy diagnostics in identifying polysensitization for the provision of ASIT to patients with seasonal allergic rhinitis.

Materials and Methods
A comprehensive search of publications in electronic databases was carried out to compile this systematic review. The search was specifically focused on papers devoted to the prospects of CRD in identifying polysensitization in patients with seasonal allergic rhinitis. To meet this goal, the following databases and search engines were utilized: PubMed, Cochrane Library, Scopus, Publons, and Google Scholar. For our systematic review of the literature, we adhered to the guidelines set forth by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRIS-MA). The flow diagram of the selection of studies for inclusion in this review is presented in Figure 1.

Materials and Methods
A comprehensive search of publications in electronic databases was carried o compile this systematic review. The search was specifically focused on papers devo the prospects of CRD in identifying polysensitization in patients with seasonal al rhinitis. To meet this goal, the following databases and search engines were utilized: Med, Cochrane Library, Scopus, Publons, and Google Scholar. For our systematic re of the literature, we adhered to the guidelines set forth by the Preferred Reporting for Systematic Reviews and Meta-Analyses (PRIS-MA). The flow diagram of the sele of studies for inclusion in this review is presented in Figure 1A.

Data Sources and Searches
In the first step of search queries, every publication was evaluated by its title, and in the second step, the publication's abstract was assessed to decide if the study met the inclusion criteria. When the publication was considered suitable, the full text was carefully evaluated. While searching in the PubMed database (accessed on 1 June 2022), the following search criteria were applied: ["Immunotherapy" (MeSH)] or ["Desensitization, Immunologic" (MeSH)] and ["Rhinitis, Allergic, Seasonal" (MeSH)] and ["Polysensitization" (title/abstract;TIAB)] and ["Molecular allergy diagnostics" (MeSH)] and ["Componentresolved diagnostics" (title/abstract; TIAB)]. All searches in PubMed, Publons, Cochrane Library, and Scopus were limited to papers in English published between 1 January 2008 and 31 January 2021. In addition, we looked for papers and articles in Russian and thus addressed eLIBRARY (accessed on 1 June 2022) for this purpose.

Study Selection
Inclusion and exclusion criteria for study selection were set by three reviewers (M.I., Y.S., and N.G.) and are presented in Figure 1. After the selection of publications that met the eligibility criteria, the following data were extracted-(i) last name of the first author and year of publication; (ii) patient age; (iii) sample size; (iv) type of allergy; (v) country; (vi) score on Ottawa-Newcastle scale [27]; and (vii) the main findings.
Type of Participants: studies of patients without prior confirmation of the type of sensitization were not eligible for inclusion.
Type of Exposure: CRD confirms the choice of allergen indication and uses for ASIT. Type of Outcome: the primary outcome was the decision on the correctness of the ASIT protocol and diagnostic accuracy for immunotherapy prescriptions. Secondary outcomes included identifying allergen source types, changing allergen source, and diagnostic concordance before and after CRD.
Study Design: observational studies, such as case reports and case series, case-control studies, randomized control trials, and prospective and retrospective cohort studies were eligible for inclusion.

Data Synthesis and Analysis
In total, 12 papers were considered suitable for inclusion (Table 1). Two coauthors (M.G. and G.A.) extracted the data presented in selected publications. Any differences in opinion were resolved in discussions between A.D., M.I., and R.F. Then, G.B., S.K., L.A., and A.M. drafted the initial version of this paper after considering all suitable papers. The final version was compiled based on feedback received from all coauthors and scores obtained on the Ottawa-Newcastle scale. Finally, the major findings on the place of molecular allergy diagnostics in the identification of polysensitization for the subsequent provision of ASIT were summarized, and the resulting conclusions were elaborated. There was significant heterogeneity in molecular sensitization profiles. None of the patients exactly matched the allergen composition of previous specific immunotherapy, containing Phl p 1, Phl p 2, Phl p 5a/b, and Phl p 6, the selection of which was based on conjunctival and nasal provocation tests.

Results
The initial search conducted in PubMed resulted in 521 publications, while the search in other databases helped to identify 47 additional records. When all duplicates were removed, 568 papers remained, and these were subjected to a thorough check. After the selection of publications, 67 papers were considered eligible based on the inclusion criteria. Application of the exclusion criteria resulted in 12 publications, which were included in this systematic review. Table 1 presents a summary of data retrieved from these publications.

Patient Characteristics
Overall, the patient population included 3302 people. Patients were selected through a skin prick test and clinical history in seven studies [73][74][75][76]78,81,83], specific IgE and clinical history in one study [72], provocation tests and clinical history in two studies [77,82], and purely IgE sensitization or skin prick test in two studies [79,80]. Patients were monosensitized in three studies [77][78][79], polysensitized in two studies [81,82], and mono-and polysensitized in seven studies [72][73][74][75][76]80,83]. One study included only adult participants. Three studies were carried out on a pediatric population, while the remaining eight studies used both children and adults as subjects. The majority of publications were focused on the ability and applications of molecular diagnostics in the accurate selection of allergens for the subsequent provision of ASIT. The studies included a determination of the specific IgE against recombinant allergens of grass, trees, and house dust mites. Changes in the provision of ASIT protocols were analyzed following the application of CRD. According to the data, allergy diagnostics often result in a change in allergens relevant to ASIT.

Evaluation of Component-Resolved Diagnosis
CRD is a tool for the identification of IgE sensitization to a panel of allergens. CRD can distinguish between primary sensitization and cross-reactive sensitization by understanding the major allergens and cross-reacting molecules [72]. This makes the identification of primary sensitization more precise and is very important for the selection of ASIT and the evaluation of its effectiveness [73,74,78]. Inhalant allergens play a particular role in seasonal allergies, and pollen grains of different plant families are the most common inhalant allergens ( Table 2).

Efficiency of Component-Resolved Diagnosis for Use in ASIT
The use of CRD impacts the choice of allergens for ASIT in at least 50% of cases [72][73][74]77,78,80,83]. This is of great importance for patients with polysensitization, as an accurate selection of allergen panels provides not only clinical benefits but also economic advantages [73,75,80,81,83]. CRD enables the detection of cross-reactive allergen components [72,79,82]. As a general rule, these are minor allergen components that are not used in ASIT. It was demonstrated that decreased levels of IgE could serve for monitoring allergen-specific blocking of non-IgE antibodies induced by ASIT [78].

Outcomes
The outcomes of studies included in this review were assessed on the basis of the diagnostic accuracy of CRD for immunotherapy prescriptions. Influence variables included allergenic molecule, marker type, and type of sensitization (monosensitization vs. polysensitization). According to the available evidence, these factors significantly influenced the change in the allergen composition of ASIT. CRD use in clinical practice leads to more detailed information on IgE reactivity at the molecular level that could be helpful in the diagnosis of a respiratory allergy and might help clinicians choose appropriate pollen extracts for ASIT (Table 1).

Discussion
This systematic review demonstrated the importance of using CRD in establishing a precise diagnosis [79,82] and identifying true and cross-reactive allergens [75,77], which ensures the correct selection of allergen composition for allergen-specific immunotherapy in children and adults [76,80,81]. By studying cross-reactive allergens, valuable information on potential sensitization to different sources of allergens and associated clinical symptoms is obtained. Some cross-reactive allergens may cause severe clinical presentation, while exposure to others will not lead to any symptoms [57,87]. The list of protein families causing cross-reactivity with inhalant allergens in seasonal allergic rhinitis is presented in Table 3.
The identification of specific IgE markers of true sensitization and cross-reactive molecules is important for CRD in patients with polysensitization [75]. Cross-reactive carbohydrate determinants (CCD) could be the source of positive IgE detection results with no clinical significance. In a serum-based allergy diagnosis, antibodies of the IgE class are directed against CCDs, therefore, give the impression of polysensitization. Anti-CCD IgE, however, does not seem to elicit clinical symptoms. Diagnostic results caused by CCDs are therefore regarded as false positives [55]. Some commercial tests, including CCD inhibitors such as ALEX, have been applied in clinical practice and have shown advantages in reducing false-positive IgE results without impacting diagnostic sensitivity toward relevant allergens [88]. Table 3. Classification of protein families in seasonal allergic rhinitis [55]. The problem with polysensitized patients is that they are sensitized to multiple sources of allergens, including cross-reactive ones. In Moreno et al.'s study, 78.2% of the subjects had polysensitization, while the rest were monosensitized or had a negative result [75]. Hu et al. showed that more than 93.0% of subjects were sensitive to more than one allergen component [82]. Haidar et al. reported that the majority of patients were polysensitized (62.65%) [83]. Therefore, the question of prescribing multiple allergens for these patients with ASIT, and how effective this treatment will be, has long been discussed. Calderon et al. concluded that single-dose therapy with two unrelated allergens is clinically effective [44]. Darsow et al. reported that CRD would be helpful in deciding on the indication for conventional ASIT in polysensitized patients [77]. In addition, Movérare et al. revealed that CRD use in clinical practice leads to more detailed information on IgE reactivity at the molecular level that could be helpful in the diagnosis of a polyclonal allergy, and might help clinicians choose appropriate pollen extracts for ASIT [72].
The implementation of a component-resolved diagnosis utilizing a range of pollen allergens is necessary for more precise patient characterization. This will enable the selection of a suitable allergen immunotherapy product, thereby allowing for the personalized management of seasonal allergic rhinitis [72][73][74][76][77][78][80][81][82][83]. Many authors report the importance of using CRD to select and influence the effectiveness of ASIT. Moreno et al. argue that when selecting the composition of AIT based solely on the results of SPT (skin prick test), approximately one-third of the patients would be treated with an allergen to which they were not allergic. Thus, according to the results of CRD, more than half of the patients had a change in the composition of ASIT compared to the initial selection [75]. Sastre et al. observed that the composition of AIT before and after molecular diagnosis coincided only in 46% of patients [73]. Letrán et al. reported a change in the ASIT protocol in more than 50% of cases after using CRD [74]. Moreover, Del-Río Camacho et al. described in their study that protocols for ASIT were changed in 54.3% of cases after CRD was conducted [81]. Martinez-Cañavate et al. reported that specialists changed the composition of the prescribed immunotherapy in 52.87% of cases [80]. In addition, Schmid et al. found that CRD might be a useful companion diagnostic to monitor the efficacy of SCIT [78]. These facts confirm the need to use CRD when selecting the composition of allergens for AIT.
High effectiveness of ASIT could only be expected in patients having IgE-antibodies to major allergens [89]. If some pollen allergen has several major allergen components, the extract for ASIT should include all of these components to enable sufficient effectiveness. ASIT will likely be moderately effective if a patient presents with a specific IgE to both major and minor allergens. Meanwhile, the absence of IgE to major allergens suggests a low response to ASIT [84].
Identification of potential ASIT patients with typical symptoms should begin at primary health care centers and include a detailed allergy history, medical examination, and assessment of serum total IgE levels to confirm allergic sensitization. In the next step, specific allergy tests should be carried out to elucidate the causes of seasonal AR. For this purpose, skin prick tests and specific IgE antibodies could be performed. When a diagnosis of seasonal allergic rhinitis is confirmed, a patient is better referred for specialized allergy care to make further tests and ASIT, which is carried out at the remission stage. In addition to a preliminary evaluation of symptoms, disease severity, and quality of life, the set of allergens for ASIT need to be selected by CRD technology. According to the European Academy of Allergy and Clinical Immunology (EAACI) guidelines, total and specific IgE should be used as biomarkers of ASIT effectiveness [50]. The summarized version of EAACI recommendations is presented in Figure 2 [50,90,91].  Medical professionals can clearly understand the diagnostic trajectory of individual with suspected allergic rhinitis by using this algorithm. Furthermore, it allows for the re duction of overdiagnosis errors during the primary medical care stage. This algorithm also serves as a guide for allergology experts, as it incorporates current advice on cutting edge technology for allergy diagnosis and treatment. Besides, the algorithm enables di ferential diagnosis of seasonal allergic rhinitis on the basis of clinical and laboratory crite ria for an optimal choice of allergen-specific immunotherapy. The algorithm reflects how the obtained data can help to monitor the effectiveness of ASIT with respect to laborator findings and also considers the assessment of symptom severity and quality of life.
The review's strengths are that we investigated the impact of several variables o treatment outcomes, including allergenic molecule type, marker type, and sensitizatio type, and discovered that they did have a significant impact on the ASIT protocol. Thi Medical professionals can clearly understand the diagnostic trajectory of individuals with suspected allergic rhinitis by using this algorithm. Furthermore, it allows for the reduction of overdiagnosis errors during the primary medical care stage. This algorithm also serves as a guide for allergology experts, as it incorporates current advice on cuttingedge technology for allergy diagnosis and treatment. Besides, the algorithm enables differential diagnosis of seasonal allergic rhinitis on the basis of clinical and laboratory criteria for an optimal choice of allergen-specific immunotherapy. The algorithm reflects how the obtained data can help to monitor the effectiveness of ASIT with respect to laboratory findings and also considers the assessment of symptom severity and quality of life.
The review's strengths are that we investigated the impact of several variables on treatment outcomes, including allergenic molecule type, marker type, and sensitization type, and discovered that they did have a significant impact on the ASIT protocol. This study describes the allergen's major and cross-reactive components at the molecular level, which can aid in the diagnosis of a respiratory allergy and help clinicians select appropriate pollen extracts for ASIT. This review has several limitations. The most significant limitations are financial, as CRD is an expensive method of research that may not be available to all patients. A limitation of some studies is the small number of participants, different sensitization profiles, and a different disease profile when allergic rhinitis is combined with bronchial asthma. In addition, different age profiles were included in some of the studies.

Conclusions
Modern allergology services prioritize the reduction of seasonal allergic rhinitis rates. CRD is highly relevant in this regard for the diagnostic accuracy of CRD for immunotherapy prescription. Such an integrated approach is most appropriate for assessing major and cross-reactive allergen components as well as identifying cases of polysensitization. Furthermore, the search for prognostic biomarkers that can predict the efficacy of ASIT must be continued. Thus, identifying reliable markers enabling the evaluation of treatment response and the prediction of outcomes in patients with different sensitization profiles would be highly desirable. Finally, it appears necessary to keep up with the development of new approaches to allergy diagnostics for choosing the most important parameters influencing the treatment strategy. Funding: This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.

Data Availability Statement:
The data presented in this study are available on request from the corresponding author.

Conflicts of Interest:
The authors declare no conflict of interest.