Investigational Treatments in Phase I and II Clinical Trials: A Systematic Review in Asthma

Inhaled corticosteroids (ICS) remain the mainstay of asthma treatment, along with bronchodilators serving as control agents in combination with ICS or reliever therapy. Although current pharmacological treatments improve symptom control, health status, and the frequency and severity of exacerbations, they do not really change the natural course of asthma, including disease remission. Considering the highly heterogeneous nature of asthma, there is a strong need for innovative medications that selectively target components of the inflammatory cascade. The aim of this review was to systematically assess current investigational agents in Phase I and II randomised controlled trials (RCTs) over the last five years. Sixteen classes of novel therapeutic options were identified from 19 RCTs. Drugs belonging to different classes, such as the anti-interleukin (IL)-4Rα inhibitors, anti-IL-5 monoclonal antibodies (mAbs), anti-IL-17A mAbs, anti-thymic stromal lymphopoietin (TSLP) mAbs, epithelial sodium channel (ENaC) inhibitors, bifunctional M3 receptor muscarinic antagonists/β2-adrenoceptor agonists (MABAs), and anti-Fel d 1 mAbs, were found to be effective in the treatment of asthma, with lung function being the main assessed outcome across the RCTs. Several novel investigational molecules, particularly biologics, seem promising as future disease-modifying agents; nevertheless, further larger studies are required to confirm positive results from Phase I and II RCTs.


Introduction
The 2022 Global Initiative for Asthma (GINA) report [1] describes asthma as a heterogeneous disease, often characterised by chronic airway inflammation, with a history of respiratory symptoms, including wheeze, shortness of breath, chest tightness, and cough that vary over time and in intensity, along with variable expiratory airflow limitation.
The long-term goals of asthma management are to achieve symptom control, reduce the risk of exacerbations and mortality, preserve lung function, and minimise drug-related side effects [1]. The stepwise approach used for pharmacological treatment in asthma mandates an iterative cycle of assessment, adjustment of pharmacological and nonpharmacological treatment, and review of the therapeutic response [1].
Over the last 30 years, inhaled corticosteroids (ICS) have been the mainstay of asthma treatment, with the long-acting β 2 -adrenoceptor agonist (LABA) formoterol/ICS combination serving as the preferred controller and/or reliever therapy, depending on asthma severity [2]. Nevertheless, this therapeutic option has become increasingly unattractive due to its inability to alter the natural course of the disease, including asthma progression [3]. Although ICS are clinically efficacious in most asthmatics, a considerable subset of patients (3-10%) remain uncontrolled despite optimal therapeutic adherence and proper

Review Question
The question of this systematic review was to assess whether some of the current investigational agents in Phase I and II clinical trials (CTs) might be suitable for effective treatment of asthmatic patients.

Search Strategy
The protocol of this synthesis of the current literature has been registered to the international prospective register of systematic reviews (PROSPERO, Protocol ID: CRD42022336605), and performed in agreement with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocols (PRISMA-P) [15], with the relative flow diagram reported in Figure 1. This study satisfied all the recommended items reported by the PRISMA 2020 checklist [16].
The PICO (Patient problem, Intervention, Comparison, and Outcome) framework was applied to develop the literature search strategy and question, as previously reported [17]. Namely, the "Patient problem" included asthmatic patients; the "Intervention" regarded investigational agents in Phase I and II CTs; the "Comparison" was performed with respect to placebo (PCB) and/or active comparators; the assessed "Outcomes" were lung function, symptoms control, blood eosinophil count (BEC), fractioned exhaled nitric oxide (FENO), exacerbations and hospitalisations, the use of rescue medications, and quality of life (QoL).
A comprehensive literature search was performed for Phase I and II CTs, written in English and investigating the impact of investigational treatments in patients with asthma. The search was performed in ClinicalTrials.gov in order to provide relevant studies available within the past 5 years (from May 2017 to May 2022).
The term "asthma" was searched for the disease, "Interventional Studies (Clinical Trials)" was selected for the study type, "Terminated" and "Completed" were chosen for The term "asthma" was searched for the disease, "Interventional Studies (C Trials)" was selected for the study type, "Terminated" and "Completed" were c for the recruitment status, and "Early Phase I", "Phase I", and "Phase II" were se in the Additional Criteria of the Advanced Search in the ClinicalTrials.org database

Study Selection
Randomised controlled trials (RCTs) reporting results concerning the efficac file of investigational treatments vs. PCB and/or active comparators were included systematic review.

Study Selection
Randomised controlled trials (RCTs) reporting results concerning the efficacy profile of investigational treatments vs. PCB and/or active comparators were included in the systematic review.
Two reviewers independently checked the relevant studies identified from ClinicalTrials.gov. The studies were selected in agreement with previously mentioned criteria, and any difference in opinion regarding eligibility was resolved by consensus.

Data Extraction
Data from included studies were extracted and checked for study references, a NCT number identifier, study duration, treatments and comparators with doses and regimen of administration, number and characteristics of analysed patients, age, gender, smoking habit, forced expiratory volume in 1 s (FEV 1 ), peak expiratory flow (PEF), Asthma Control Questionnaire (ACQ) score and other outcomes related to the impact on symptoms, BEC, FENO, asthma exacerbations, hospital admissions, rescue medication use, Asthma QoL Questionnaire (AQLQ) score, St George's Respiratory Questionnaire (SGRQ) score, and study quality assessment via the Jadad Score [18] and Cochrane Risk of Bias 2 (RoB 2) [19].

Endpoints
The co-primary endpoints of this systematic review were the impact of investigational treatments on lung function and symptoms control.
The secondary endpoints were the impact of investigational treatments on blood eosinophil count, FENO, exacerbations and hospitalisations, the use of rescue medications, and QoL.

Strategy for Data Synthesis
Data from original papers were extracted and reported via qualitative synthesis, and the statistical significance was set at p < 0.05.

Quality Score
The summary of the risk of bias for each included randomised trial was analysed via the Jadad score [18] and Cochrane RoB 2 [19]. The weighted assessment of the overall risk of bias was analysed via the Cochrane RoB 2 [19] using the robvis visualisation software [20,21]. The Jadad score, with a scale of 1-5 (with a score of 5 being the best quality), was used to assess the quality of the papers concerning the likelihood of bias related with randomisation, double blinding, withdrawals, and dropouts [18]. Studies were considered of low quality at Jadad score < 3, of medium quality at Jadad score = 3, and of high quality at Jadad score > 3. The weighted assessment of the risk of bias was analysed via the Cochrane RoB 2 tool [19] by using the robvis visualisation software [20,21].
Two reviewers independently assessed the quality of individual studies, and any difference in opinion about the quality score was resolved by consensus.

Study Characteristics
Of the 101 records identified in the ClinicalTrials.gov database, 75 documents were excluded due to inconsistency between the study title and the PICO framework or because no results were available. Among the remaining CTs, 19 RCTs were deemed eligible for the systematic review.

IL-4Rα Inhibitor
Antagonising the IL-4 receptor α subunit (IL-4Rα) interferes with the downstream IL-4/IL-13 signalling, which is central to the pathogenesis of asthma [55]. As a matter of fact, IL-4 regulates the proliferation and survival of T helper 2 (Th2) cells as well as immunoglobulin E (IgE) synthesis, while IL-13 is implicated as a key effector in AHR, mucus hypersecretion, ASM alterations, and subepithelial fibrosis [56].

Anti-IL-5 mAbs
Targeting BEC reduction through the inhibition of IL-5 represents an established therapeutic option in severe asthma [57]. Depemokimab (GSK3511294) is a subcutaneously administered anti-IL-5 mAb, designed for improved affinity and long-acting IL-5 suppression compared to the currently approved anti-IL-5 mAbs, and it has been evaluated in a first-in-human Phase I RCT [30,31] enrolling mild to moderate asthmatic patients with BEC ≥ 200 cells/µL at screening.

Anti-IL-17A mAbs
Increased expression of the Th17-derived cytokine IL-17A has been observed in sputum, airway tissue biopsies, and serum from asthmatic patients [58][59][60][61][62] and was positively associated with a more severe asthma phenotype [59,63,64] and neutrophilic inflammation [65]. Considering that Th17-high patients are less sensitive or even unresponsive to ICS [59,66] and that asthma progression differs from more treatable Th2 types of the disease [67], developing an effective therapy targeting Th17/IL-17A axis would overcome a major unmet need in severe asthma.
A Phase II RCT [22] investigated the subcutaneously administered anti-IL-17A mAb CJM112 300 mg when added to existing therapy in patients with inadequately controlled moderate to severe asthma, with low serum IgE and BEC. The effect of CJM112 treatment on trough FEV 1 was not different from PCB, but a significant (p < 0.05) improvement was observed in the ACQ6 score (mean difference (MD) −0.22 units (80%CI −0.41-−0.04)) and the ACQ7 score (MD −0.23 units (80%CI −0.40 to −0.06)) vs. PCB. A higher proportion of patients receiving CJM112 had a decrease of ≥0.5 units in the ACQ7 score compared with PCB (71.7% vs. 52.8%) [22].
A Phase II RCT [32,33] investigated the efficacy of subcutaneous itepekimab 300 mg administered alone or in combination with dupilumab 300 mg to patients with moderate to severe asthma, who progressively reduced and discontinued background therapy of inhaled corticosteroid/long-acting β2 adrenoceptor agonist (ICS/LABA) over 12 weeks. Itepekimab significantly (p < 0.05) improved trough FEV 1 compared to PCB (MD 140 mL (95%CI 10-270)) and it was as effective as dupilumab, but no improvement was seen upon treatment with the combination therapy. Itepekimab did not increase post-bronchodilator FEV 1 vs. PCB, but when combined with dupilmab, the improvement was significant (p < 0.05) (MD 130 mL (95%CI 10-250)) and comparable to that of dupilumab administered alone [32,33].
The percentage of patients with an event indicating a loss of asthma control was lower in the itepekimab (22.0%) and combination therapy (27.0%) groups vs. PCB (41.0%). The corresponding odds ratio (OR) for the comparison of itepekimab vs. PCB was significant (p < 0.05) (OR 0.42 (95%CI 0.20-0.88)) and similar to the OR for dupilumab vs. PCB; no difference was detected in the ORs for the comparison between combination therapy and PCB, itepekimab monotherapy, and dupilumab monotherapy. Itepekimab alone and combined with dupilumab significantly (p < 0.05) improved ACQ5 score vs. PCB (MD −0.42 units (95%CI −0.73-−0.12) and MD −0.32 units (95%CI −0.63-−0.01), respectively), and the effect was similar to that observed with dupilumab [32,33].
The BEC significantly (p < 0.05) decreased upon treatment with itepekimab administered alone or combined with dupilumab vs. PCB, and the effect was significantly (p < 0.05) different from that induced by dupilumab monotherapy, which, as expected [32], transiently induced blood eosinophilia. The FENO level was significantly (p < 0.05) lowered in the itepekimab group, although the magnitude of reduction was lower than that observed in the combination therapy and dupilumab groups. Patients treated with itepekimab administered alone or combined with dupilumab showed a significant (p < 0.05) improvement in their AQLQ score vs. PCB (MD 0.45 units (95% CI 0.14-0.77) and MD 0.43 units (95% CI 0.11-0.75), respectively), with an effect comparable to that of dupilumab [32,33].
In a Phase II RCT [40,41], a single dose of etokimab administered at 300 mg/100 mL via intravenous infusion did not improve FEV 1 compared to PCB in severe eosinophilic asthma; no data are available for symptoms control.
The reduction in peripheral BEC following etokimab treatment was similar to that observed with PCB, and no difference was detected in FENO levels. The number of asthma exacerbations experienced by patients treated with etokimab was no different from those treated with PCB [40,41].
A Phase II RCT [23] reported that intravenously administering melrilimab 10 mg/kg to patients with moderate to severe asthma and allergic fungal airway disease for 12 weeks did not improve their FEV 1 and ACQ5 score compared to PCB. No differences between melrilimab and PCB were observed with respect to the change from baseline in BEC, FENO level, and AQLQ score [23].
Another Phase II RCT [24] showed that melrilimab 10 mg/kg administered for 16 weeks to moderately severe asthmatic patients who gradually reduced and discontinued background therapy with fluticasone propionate/salmeterol (FP/SAL) 500/50 µg, did not improve trough FEV 1 and morning and evening PEF vs. PCB. The reduction in ACQ5 score was similar with both melrilimab and PCB, but the percentage of patients who experienced loss of asthma control was lower in the group treated with melrilimab (67.0%) than with PCB (81.0%). No differences between the two treatment groups were observed in the percentage of night-time awakenings due to asthma symptoms requiring rescue medication use and in the daytime asthma symptom score [24].
The effect induced on BEC and FENO level was similar in the melrilimab and PCB groups. The percentage of patients with an asthma exacerbation requiring OCS and/or hospitalisation was higher with melrilimab (13.0%) than with PCB (7.0%). No differences between the two groups were observed in terms of daily use of rescue medications and SGRQ total score [24].

Anti-TSLP mAbs
Similar to IL-33, TSLP is mainly an epithelium-derived alarmin, which plays an upstream role in the initiation of type-2-driven immune responses [71]. In asthma, the number of cells expressing TSLP messenger ribonucleic acid (mRNA) within the airway epithelium and submucosa is markedly increased compared to healthy controls [72]. In a subset of patients with severe asthma, TSLP expression remained enhanced, independent of treatment with high-dose ICS or OCS [73]. Therefore, targeting TSLP signalling represents an intriguing therapeutic strategy in asthma [74].
In a Phase I RCT [53,54] the anti-TSLP mAb fragment ecleralimab (CSJ117) 4 mg was administered via a dry powder inhaler (DPI) for 12 weeks to patients with mild atopic asthma, who exhibited an early asthmatic response (EAR) and late asthmatic response (LAR) to a common inhaled allergen. Ecleralimab did not induce an attenuation in the EAR, as documented by the maximum percentage fall in FEV 1 or as time-adjusted area under the curve (AUC), and numerically increased the minimum of the absolute in FEV 1 compared to PCB. During the LAR, ecleralimab significantly (p < 0.05) reduced the maximum percentage decrease in FEV 1 (MD −8.42% (90%CI −15.66-−1.18)) from pre-allergen inhalation challenge and the time-adjusted AUC fall in FEV 1 (MD −7.18% (90%CI −11.92-−2.44)), compared to PCB. Patients in the ecleralimab group showed a strong trend towards a significant (p = 0.05) increase in the minimum absolute FEV 1 during LAR vs. PCB (MD 0.27% (90%CI 0.00-0.55)) [53,54]. No data are available for symptoms control [53,54].

MABAs
Bifunctional M 3 receptor muscarinic antagonists/β 2 -adrenoceptor agonists (MABAs) are dimeric molecules that simultaneously block M 3 muscarinic receptors while activating β2 receptors, and thus may be readily co-formulated with anti-inflammatory agents [81,82], simplifying dosing schedules and improving patient adherence to medication. A Phase I/II RCT [42,43] reported that in asthmatic patients, the inhaled MABA CHF6366 significantly (p < 0.05) improved the change from pre-dose in FEV 1 on day 1 when administered at 160 µg, but not at 40 µg, 80 µg, and 240 µg, compared to the effect induced by PCB, while no difference was detected in the change from pre-dose in FEV 1 on day 7; no data are available for symptoms control [42,43].

DP 2 Antagonists
Evidence suggests that preventing the activation of the prostaglandin D2 receptor (DP2) pathway improves symptoms of asthma and pulmonary function, and impairs any change in eosinophil shape, while indirectly inducing a reduction in the number of exacerbations in severe asthmatic patients [83].
There was a numerical reduction in the annualised rate of severe asthma exacerbations compared to PCB [36,37].

Selective BTK Inhibitors
Bruton's tyrosine kinase (BTK) is a member of the Tec family of tyrosine kinases involved in the high-affinity receptor for IgE (FcεRI)-dependent mast cell production of cytokines and degranulation [84,85], and in the IgE-mediated activation of human basophils [86]. BTK inhibitors could be useful to treat pathological mast cell responses of asthma [87].
A Phase II RCT [25] reported that orally administering remibrutinib (LOU064) 100 mg to inadequately controlled asthmatic patients did not induce an improvement in trough FEV 1 and in morning and evening PEF compared to PCB. Changes in the ACQ5 score, in the asthma symptom score, and in the number of puffs of SABA taken daily were not different between remibrutinib and PCB groups [25].

ENaC Inhibitors
An imbalance in ion transport across the airway epithelium has been implicated in asthma pathogenesis. Dysfunctions in the cystic fibrosis transmembrane conductance regulator and epithelial sodium channel (ENaC) cause changes in the airway surface liquid permeation, leading to modifications of mucus rheological properties and impairment. Blocking ENaC may reduce airway water reabsorption and increase mucus moist, therefore it is considered a potential target for the treatment of asthma [88].
A Phase I RCT [26] investigated the ENaC inhibitor BI 443651 100 µg, 400 µg, and 1200 µg administered via soft mist inhaler (SMI) to patients with mild asthma following a bolus methacholine (MCh) challenge. In the single-blind, double-dummy Part 1 of the RCT, no difference was detected between BI 443651 and PCB in terms of absolute change from baseline in maximum FEV 1 reduction. In the double-blind, double-dummy Part 2 of the RCT, only BI 443651 administered at 1200 µg significantly (p < 0.05) improved the maximum FEV 1 reduction vs. PCB (MD −157 mL (90%CI −266-−47)). No data are available for symptoms control [26].

Pan-JAK Inhibitors
According to in vitro studies performed on inflammatory cells isolated from asthmatic patients, pan-JAK inhibitors reduced cytokine levels and showed an additive effect on lymphocyte inhibition when combined with ICS [89]. Lung inflammation was improved upon treatment with pan-JAK inhibitors in animal models of airway inflammation [90][91][92].
A Phase II RCT [27] reported that the pan-JAK inhibitor TD-8236 administered at 150 µg and 1500 µg via DPI did not improve the FEV 1 AUC from 3 to 8 h and the maximum percentage decline in FEV 1 from 3 to 8 h following inhaled allergen challenge compared to PCB. No data are available for symptoms control [27].

Anti-Fel d 1 mAbs
The secretoglobulin Fel d 1 is the major cat allergen, eliciting IgE-mediated allergic symptoms in up to 95% of individuals with a cat allergy [93,94], such as sneezing, runny nose, nasal obstruction, conjunctivitis, and/or asthma [95]. REGN1908-1909 is an anti-Fel d 1 cocktail of two IgG4 mAbs, REGN1908 and REGN1909, with a high affinity for and noncompetitive binding to distinct epitopes of Fel d 1, which prevents the allergen cross-linking of IgE-FcεRI complexes on mast cells and basophils and the consequent degranulation and release of inflammatory mediators [96,97].

Synthetic Amino-Benzothiazoles
The synthetic amino-benzothiazole dexpramipexole was first developed as a treatment for amyotrophic lateral sclerosis (ALS) and during the development program, a marked targeted depletion of BEC was observed in ALS patients; therefore, dexpramipexole holds promise for asthma and eosinophil-associated diseases [98].
In the EXHALE Phase II RCT [28,29], dexpramipexole (KNS-760704) orally administered at 37.5 mg, 75 mg, and 150 mg BID for 12 weeks was investigated in patients with poorly controlled moderate to severe eosinophilic asthma with an absolute BEC of ≥300 cells/µL. No differences were observed between dexpramipexole 37.5 mg and 75 mg and PCB in trough FEV 1 and post-bronchodilator FEV 1 , while dexpramipexole 150 mg showed a numerical improvement in both outcomes vs. PCB at the end of the treatment period, and a significant increase in trough FEV 1 at weeks 16/18 vs. PCB. The effect of treatment on the ACQ6 score was similar to that observed in the PCB group [28,29].

Antifungal Triazoles
Respiratory fungal infections complicate lung diseases and, particularly in severe asthma, up to 70.0% of patients are sensitised to at least one fungal allergen [5,99,100]. In a Phase I RCT [38,39], a single dose of inhaled PC945 5 mg did not induce a change in FEV 1 (defined as >15.0% change from baseline, measured 10 min after receiving PCB) in mild asthmatic patients, and no acute bronchospasm was observed.

Probiotics
Probiotics exhibited anti-inflammatory properties to modulate immune functions and were characterised by good tolerance and safety [44]. According to preliminary results of a Phase II/III RCT [44,45] in severe uncontrolled asthma, a change from baseline in ACQ score was similar in patients receiving the orally administered Probiotical ® and PCB; no data are available on lung function. A significant (p < 0.05) reduction in the percentage of sputum eosinophils was observed between baseline and after 3 months of therapy in the Probiotical ® group (0.5% (95%CI 0.0-2.3) vs. 0.1% (95%CI 0.0-0.5)) compared to the PCB group (4.5% (95%CI 1.5-9.3) vs. 2.4% (95%CI 1.2-9.4)) [44,45].

Risk of Bias
The traffic light plot for the assessment of each included RCT is reported in Figure 2A, and the weighted plot for the assessment of the overall risk of bias by domains is shown in Figure 2B.

Discussion
An investigational medication is defined as a drug and/or formulation that has been approved for clinical testing by either the U.S. Food and Drug Administration (FDA) or the European Medicines Agency (EMA), but has not gained marketing authorisation yet [101,102]. Over the last five years, results from 19 Phase I and II RCTs on investigational agents for the treatment of asthma reported data from sixteen classes of investigational agents. Specifically, these investigational drugs included AZD1402, BI 443651, CHF6366, CJM112, depemokimab, dexpramipexole, ecleralimab, etokimab, GB001, itepekimab, All of the included RCTs (100.0%) had a low risk of bias in missing outcome data. For 18 RCTs (94.7%), there was a low risk of bias for the randomisation process, and for 17 RCTs (89.5%), the bias due to deviations from intended intervention was low. For two RCTs (10.5%), there were some concerns in the domain of bias due to deviations from intended intervention, and for one RCT (5.3%) there were some concerns for the randomisation process.
For 14 RCTs (73.7%), no information was available with regard to the risk of bias in the measurement of the outcome and selection of the reported results, as no full text articles concerning the studies have been published yet.

Discussion
An investigational medication is defined as a drug and/or formulation that has been approved for clinical testing by either the U.S. Food and Drug Administration (FDA) or the European Medicines Agency (EMA), but has not gained marketing authorisation yet [101,102]. Over the last five years, results from 19 Phase I and II RCTs on investigational agents for the treatment of asthma reported data from sixteen classes of investigational agents. Specifically, these investigational drugs included AZD1402, BI 443651, CHF6366, CJM112, depemokimab, dexpramipexole, ecleralimab, etokimab, GB001, itepekimab, melrilimab, PC945, REGN1908-1909, remibrutinib, TD-8236, velsecorat, indacaterol acetate, and a probiotic. Overall, the quality of the studies was good, although often data were not published in full text articles; thus, scarce information was available to adequately perform the RoB assessment.
The investigational anti-IL-4Rα inhibitor AZD1402, the anti-IL-5 mAb depemokimab, the anti-IL-17A mAb CJM112, the anti-TSLP mAb ecleralimab, the ENaC inhibitor BI 443651, the MABA CHF6366, and the anti-Fel d 1 mAb REGN1908-1909 were proven effective in the treatment of asthma, although data almost exclusively regarded the assessment of lung function, and thus did not allow conclusions regarding symptoms control and the secondary endpoints of this systematic review. The effectiveness of the LABA indacaterol was confirmed even when delivered using the formulation with maleate salt, which demonstrated an effect that was comparable to the currently marketed indacaterol acetate on FEV 1 , PEF, and rescue medication use reduction. Among the investigational anti-IL-33 mAbs, only itepekimab, but not etokimab and melrilimab, effectively improved asthma outcomes compared to PCB, but generally there was no further improvement observed when itepekimab was combined with dupilumab. Treatment with the SGRM velsecorat was generally superior to PCB when administered at higher doses.
Overall, investigational agents did not show superiority to active controls, with the exception of itepekimab, which significantly reduced BEC compared to dupilumab monotherapy, and velsecorat, which induced a significantly greater improvement in FEV 1 vs. PCB compared to that produced by FF vs. PCB.
The main efficacy outcome assessed by the RCTs included in this systematic review was FEV 1 . In this respect, BI 443651, depemokimab, ecleralimab, indacaterol maleate, itepekimab, REGN1908-1909, and velsecorat produced a statistically significant improvement in lung function compared to PCB, thus representing promising add-on therapies for asthma in the future. It is also worth mentioning the synthetic amino-benzothiazole dexpramipexole, which was found to markedly reduce BEC across all the administered doses in patients with moderate to severe eosinophilic asthma, despite no significant improvement in lung function, relative to PCB [28,29].
The anti-IL-33 mAbs etokimab and melrilimab, the DP2 antagonist GB001, the selective BTK inhibitor remibrutinib, the pan-JAK inhibitor TD-8236, and the antifungal triazole PC945 induced an effect on lung function that was similar to PCB.
Although FEV 1 is generally recognised by the research community and regulatory agencies to be a suitable variable for airflow obstruction assessment [103], it is not the most relevant endpoint for testing investigational anti-inflammatory agents, including the DP2 antagonist GB001 and the pan-JAK inhibitor TD-8236, particularly for short-term assessment. Thus, for such treatments, other efficacy endpoints should be considered in future studies. Although probiotics utilised in dietary supplements reside in a sub-category under the general umbrella term of "foods" rather than drugs, according to both the FDA [104] and the European Food Safety Authority (EFSA) [105], the probiotic Probiotical ® investigated in a Phase II RCT (NCT03341403) [44,45] for uncontrolled severe asthma was included in this systematic review and was treated as investigational agent. The hypothesis of such RCT [44,45] was that Probiotical ® could have an impact in asthmatic patients who were not optimally controlled, reducing the local and systemic inflammatory state and then improving QoL and asthma control [44,45]. This hypothesis was also supported by the evidence that certain probiotic strains have anti-inflammatory and immunomodulatory effects in pre-clinical models of asthma [106,107] and RCTs of adult asthma [108,109]. Interestingly, although dietary supplements are not subjected to the pre-market approval requirement for drugs, an investigational new drug application must be submitted to the FDA if the clinical investigation is intended to evaluate whether a dietary supplement is useful in diagnosing, curing, mitigating, treating, or preventing a disease, under the Code of Federal Regulations Part 312 [110]. In contrast, in the EU, there is still no specific regulation covering probiotics, pre-biotics, synbiotics, or postbiotics, but as suggested by The International Scientific Association of Probiotics and Prebiotics consensus statement, the definition of such products requires a health benefit; thus, it is expected that the use of any of these terms would require a health claim approval [111].
In any case, the daily administration of Probiotical ® showed some improvement in sputum eosinophil count after 3 months of therapy, but in agreement with the current scientific evidence, the use of probiotics as adjuvant therapy for asthma is not yet conclusive [112]. Three meta-analyses carried out to explore the potential effects of probiotics in preventing allergic diseases and asthma led to conflicting outcomes due to a high degree of heterogeneity among the studies, mostly concerning the design, the characteristics of included patients, the analysed variables, and the used probiotic strains [113][114][115].
The assessment of efficacy outcomes reported by the RCTs included in this systematic review indicates that not only were some of the investigational agents superior to PCB from a statistical point of view, but they also elicited clinically relevant effects compared to PCB in asthmatic patients, as reported in Table 2. As a matter of fact, itepekimab 300 mg Q2W, indacaterol maleate 150 µg QD, and velsecorat 720 µg QD overcame the Minimal Clinical Important Difference (MCID) [103] threshold for trough FEV 1 or risk of asthma exacerbation. Interestingly, velsecorat 720 µg QD, itepekimab 300 mg Q2W, and itepekimab 300 mg Q2W + dupilumab 300 mg Q2W were borderline to reach the MCID threshold for ACQ or AQLQ. Indeed, these promising results need to be confirmed by Phase III studies.
A main limitation of this systematic review is that most of the included studies (11 RCT, 57.9%) had a registry record on ClinicalTrials.gov and/or EU Clinical Trial Register but no associated publication, and thus, sponsors and principal investigators are exclusively responsible for the scientific accuracy of the provided results, which may be inconsistent across all the provided studies.
Additionally, findings for three RCT of the included studies were retrieved from grey literature which were not formally and rigorously peer reviewed, and thus should be carefully interpreted due to potential publication bias [116].
There is a strong pharmacological need to look beyond current therapeutic strategies and consider further promising biological drugs for asthma that are under development and for which results have not been posted on clinical trial registries and are not available in current literature.
Asthma remission is a complex condition that can be clinically defined as a sustained absence of symptoms, optimisation or stabilisation of lung function, and no use of OCS for exacerbation treatment [117], but controversy remains regarding the threshold of each item used to assess the asthma remission itself [118]. Although these terms do not necessarily imply the absence of airway pathology, a recent point of view suggested that asthma remission may be an achievable goal, at least in asthmatic patients with the T2 phenotype [117].
In conclusion, novel investigational agents, such as biologics, may have the potential to promote disease modification. Clearly, further larger studies are needed to confirm positive results from Phase I and II RCTs. So far, most of the investigated therapies have been evaluated as add-on options to current treatment, but it would be extremely advantageous for new therapies to be effective enough to replace current pharmaceutical options in order to simplify regimens of administration.  Author Contributions: All the authors (L.C., M.A., A.F., E.P., B.L.R., P.R. and A.C.) made substantial