Phase III Multicenter, Double-Blind, Randomized, Placebo-Controlled Trial Evaluating Subcutaneous Immunotherapy with Cupressus arizonica Protein Cup a 1 in Allergic Rhinoconjunctivitis

The primary objective of this clinical trial is the **assessment of efficacy** of specific immunotherapy with the purified and isolated protein Cup a 1, administered subcutaneously in patients sensitized to the allergen under study. This is clinically relevant as it aims to determine the therapeutic potential of Cup a 1 in managing symptoms of moderate to severe **allergic rhinoconjunctivitis**, with or without controlled asthma, in individuals sensitized to Cupressaceae pollen.

Secondary objectives include:

• Analysis of additional efficacy parameters to the combined index of symptom frequency and severity score, along with rescue medication consumption score.
• Evaluation of the variation of follow-up skin tests with Cupressaceae and Cup a 1 extracts.
• Assessment of asthma and allergic rhinitis control using the ACT and the modified ARIA scale.
• Patient's health status assessment using the VAS scale.
• Quality of life evaluation according to rhinitis (ESPRINT-15).
• Evaluation of immunological markers related to efficacy: IL-10, IL-13, INF-γ, IgG4, total IgE, and specific IgE against Cupressus arizonica and Cup a 1.
• Assessment of the degree of tolerance against conjunctival provocation with Cupressaceae extract.
• Evaluation of the safety of the treatment through the registry of adverse events (AA) and adverse drug reactions (AAG).

The clinical trial focuses on evaluating the efficacy of specific immunotherapy with the purified and isolated protein Cup a 1, administered subcutaneously in patients with **allergic rhinoconjunctivitis**. The study population includes both male and female participants aged between 12 and 65 years. Participants are required to have moderate or severe allergic rhinitis or rhinoconjunctivitis, with or without controlled asthma, and must demonstrate sensitization to Cupresaceae. The trial involves individuals with Cupressus or Cup a 1 IgE levels of 2 kU/l or higher and a positive skin prick test for Diater extract of Cupressaceae arizónica and Cup a 1. Participants must exhibit clinically relevant symptoms against Cupresaceae and be willing to comply with all protocol requirements. The trial includes a vulnerable population, and female participants of childbearing potential must have a negative urine pregnancy test and agree to use effective contraception. The sponsor has not provided information regarding the total number of participants in the study.

The clinical trial is a **randomized**, **double-blind**, **placebo-controlled** phase III study designed to evaluate the clinical efficacy and safety of immunotherapy with the purified major allergen **Cup a 1** in patients with moderate to severe **allergic rhinoconjunctivitis**, with or without controlled asthma, sensitized to Cupressaceae. The trial is conducted in parallel groups, where participants are randomly assigned to receive either the investigational product or a placebo. The investigational product, Cup a 1, is administered as a **suspension for injection** via the **subcutaneous route**. The placebo is composed of a 0.9% saline solution with mannitol and histamine, and its appearance, packaging, interval, route, and method of administration are identical to the investigational product.

The trial is expected to last until March 31, 2025, with recruitment having started on May 16, 2022. Participants will be involved in the study for a maximum treatment period of 24 months. The study includes several key visits: an initial screening visit to confirm eligibility, regular follow-up visits to monitor treatment response and safety, and an end-of-study visit to assess the overall outcomes. The primary endpoint is the efficacy of the treatment, evaluated based on symptom frequency and severity scores, along with the Combined Symptom and Medication Score (CSMS) obtained through patient questionnaires during two successive pollen seasons. Secondary endpoints include symptom-free days, medication-free days, and the frequency of adverse events, among others.

Participants are required to meet specific inclusion criteria, such as being between 12 and 65 years of age, having moderate or severe allergic rhinitis or rhinoconjunctivitis, and demonstrating sensitization to Cupressaceae. They must also be willing to comply with all protocol requirements and be available for follow-up throughout the trial duration. Conditions that may lead to early termination from the study include non-compliance with the study protocol, withdrawal of consent, or the occurrence of adverse events that necessitate discontinuation of the investigational product. The trial aims to provide valuable insights into the efficacy and safety of Cup a 1 immunotherapy in the target patient population.

The clinical trial involves the administration of **Cupressus arizonica, protein Cup a 1**, which is a **suspension for injection**. This experimental medication is administered subcutaneously. The maximum daily dose is 2.4 µg, with a total maximum dose of 57.6 µg over a treatment period of 24 weeks. The active substance is a structurally diverse allergen, specifically designed for patients with moderate to severe allergic rhinoconjunctivitis, with or without controlled asthma, sensitized to Cupressaceae. The product is manufactured by DIATER LABORATORIO DE DIAGNOSTICO Y APLICACIONES TERAPEUTICAS S.A.

Another formulation of **Cupressus arizonica, protein Cup a 1** is also used in the trial, with a higher maximum daily dose of 5.6 µg and a total maximum dose of 134.4 µg over the same 24-week period. This formulation is similarly administered as a **suspension for injection** via the subcutaneous route. The purpose of this variation is to assess the efficacy and safety of different dosing regimens of the same allergen in the target patient population.

The placebo used in this study is designed to match the appearance, packaging, interval, route, and method of administration of the investigational medicinal product (IMP). It consists of a 0.9% saline solution with mannitol and histamine. The placebo is administered subcutaneously, following the same schedule as the experimental treatments, to maintain the double-blind nature of the trial. This ensures that any observed effects can be attributed to the active treatment rather than the placebo effect.

The efficacy of the treatment in this clinical trial will be assessed using a combination of primary and secondary endpoints. The primary endpoint involves evaluating the treatment's efficacy based on the symptom frequency and severity score, along with the Combined Symptom and Medication Score (CSMS). This will be obtained through a questionnaire completed by the patient during two successive pollen seasons. Secondary endpoints include various measures such as symptom frequency and severity score comparisons between treatment arms and pollen seasons, score for rescue medication use, symptom-free days, and medication-free days. Additionally, the trial will monitor extra visits and/or emergencies related to the pathology under study.

Further assessments will include a follow-up skin test with Cupressaceae extract and **Cup a 1**, control of asthma and allergic rhinitis assessed by the Asthma Control Test (ACT) and modified ARIA, and a Visual Analog Scale (VAS) for the state of health performed by the patient. The ESPRINT-15 questionnaire will also be evaluated. Immunological markers such as IL-10, IL-13, INF-γ, specific IgE, and specific IgG4 against Cupressus arizonica and **Cup a 1** will be measured. Conjunctival provocation against Cupressaceae will be conducted, and the frequency of adverse events (AEs), serious adverse events (SAEs), and adverse drug reactions (ADRs), particularly systemic events according to the World Allergy Organization (WAO) 2010 classification, will be recorded.