Repertoire and Properties of Anti-drug Antibodies Involved in Immediate Hypersensitivity in the Operating Room

Study Overview
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Brief Summary
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Acute per-anesthetic hypersensitivity reaction (HSA-PA) is a rapidly occurring systemic reaction following injection of a drug during anesthesia (mortality between 3 and 9%). The substances responsible for these reactions in France are curare in 60% of cases, followed by antibiotics. The main mechanism mentioned is an immediate systemic hypersensitivity immune reaction mediated by IgE antibodies (anaphylaxis). NeuroMuscular Blocking Agents (NMBA; curare) relax skeletal muscles to facilitate surgeries and permit intubation, but lead to adverse reactions: (a) severe hypersensitivity reactions (anaphylaxis) thought to rely on pre-existing anti-NMBA antibodies; (b) complications due to postoperative residual curarization. Identification of patients at risk remains suboptimal due to the lack of adequate tools to detect anti-NMBA antibodies. A capturing agent exists for only one out of the four most used NMBAs, allowing reversal of profound curarization. Case reports suggested that it might also ameliorate an ongoing anaphylaxis due to that NMBA. Based on strong preliminary results, our study proposes to characterize anti-drugs antibody repertoires in patients with various NMBA or antibiotics-anaphylaxis, describe activation pathways leading to anaphylaxis, develop and validate diagnostic and therapeutic molecules to ameliorate patient screening, NMBA-anaphylaxis and reverse profound neuromuscular block.

Detailed Description
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Acute per-anesthetic hypersensitivity reaction (HSA-PA) is a rapidly occurring systemic reaction following injection of a drug during anesthesia (mortality between 3 and 9%). The substances responsible for these reactions in France are curare in 60% of cases, followed by antibiotics. The main mechanism mentioned is an immediate systemic hypersensitivity immune reaction mediated by IgE antibodies (anaphylaxis). NeuroMuscular Blocking Agents (NMBA; curare) used in the clinic (e.g., atracurium, suxamethonium, rocuronium, vecuronium) belong to the curare family and block the nicotinic acetylcholine receptor to cause muscle relaxation. They share the presence of quaternary ammonium groups that are required for binding the acetylcholine receptor, and thus for their biological activity. Even though NMBA are relatively safe molecules, two main types of adverse reactions have been reported. The most striking are immediate and severe anti-drug reactions at NMBA infusion with 1/10,000 patients suffering from severe hypersensitivity reactions, i.e. anaphylaxis. Anaphylaxis is classically considered to rely on IgE antibodies against the culprit compound, and to involve massive histamine liberation by mast cells and basophils, following antigen-induced aggregation of IgE receptor (FcεRI)-bound specific IgE antibodies1. The clinical diagnosis of anaphylaxis to NMBA is based on this IgE paradigm. NMBA quaternary ammonium cations are considered their major allergenic epitopes2, even if patients are rarely hypersensitive to several NMBA, even closely related NMBA like rocuronium and vecuronium that derive from each other, suggesting that antibodies may bind NMBA-specific structures other than quaternary ammoniums. Antibodies may even interact with NMBA as a whole epitope due to NMBA very reduced size; NMBA indeed vary from 300-1,000 Da. Intriguingly, 10-20% of patients who experience NMBA-induced anaphylaxis do not present with any biological signs of IgE-dependent immune activation, suggestive of other potential mechanisms. 4 An IgG antibody pathway has recently been described in NMBA-induced human anaphylaxis, leading to Platelet-Activating Factor release, neutrophil and platelet activation, which may aggravate anaphylaxis in combination with the IgE-pathway or underlie anaphylaxis in the absence of specific IgE. The other most common adverse reaction is not allergic in nature, and is due to prolonged presence of NMBA following surgery, i.e. residual curarization. This very frequent complication (up to 83% of the patients) leads to adverse postoperative pulmonary events, pharyngeal dysfunction, urgent tracheal reintubation and prolonged stay in post-anesthesia care units that lead to a high economic burden13. The first option for pharmacological neuromuscular blockade reversal, prostigmine (used for atracurium reversal), is restricted to situations where half of the spontaneous reversal is effective. The second one is a synthetic cyclodextrin, sugammadex, that captures rocuronium or vecuronium in vivo within minutes following injection. Instant blockade reversal might be necessary, as in difficult intubation scenarios, explaining that it has been considered economically advantageous to administer sugammadex despite a cost of $100/dose, leading to a $400-$3,000 economy per patient. Based on strong preliminary results, our study proposes to characterize anti-drugs antibody repertoires in patients with various NMBA or antibiotics-anaphylaxis, describe activation pathways leading to anaphylaxis, develop and validate diagnostic and therapeutic molecules to ameliorate patient screening, NMBA-anaphylaxis and reverse profound neuromuscular block.

Official Title
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Repertoire and Properties of Anti-drug Antibodies Involved in Immediate Hypersensitivity in the Operating Room

Conditions
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Antibiotic Allergy, Hypersensitivity, Immediate, Peri-operative Injury, Antibody Hypersensitivity

Intervention / Treatment
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* Diagnostic Test: IgG group
* Diagnostic Test: IgE group
* Diagnostic Test: IgG group + IgE group


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Common to both groups: Age ≥ 18 years old and ≤ 70 years old Patient having presented an allergic reaction in the operating room in the last 10 years, regardless of the grade of severity, the causative agent or the type of anesthesia Patient having benefited from an allergy-anaesthesia consultation in the investigator center For IgE group: - presence of at least one positive skin test to one of the suspected agent (curare and/or antibiotic) during the allergist consultation For the IgG group: - presence of circulating IgG-type antibodies directed against one of the suspected agents (curare, antibiotic, antiseptic), identified in the routine biological tests carried out at the time of the shock and / or the consultation of allergology. Exclusion Criteria: Common to both groups: Absence of written informed consent Protected person: under guardianship or curatorship Patient without social security Pregnancy of breast feeding Ongoing immunosuppressive or chemotherapy Acute heart failure Patient included in another interventional research with an exclusion period For IgE group: Taking oral or injectable anticoagulants Taking a double antiplatelet aggregation Previous sternotomy Previous thoracic radiation therapy Known allergy to local anesthesic or ioda Previous major sternal cutaneous lesions Prerequisites for carrying out the sternal sample in order to carry out a myelogram (IgE group): Leukocytes > 4 giga/L in the 2 months preceding inclusion Platelets > 100,000 / mm3 in the 2 months preceding inclusion For the IgG group Orthostatic hypotension Realization of a blood donation in the 6 to 8 weeks preceding inclusion Prerequisites for performing the blood sample (IgG group): - Hemoglobin > 12.5 g/dL for women and > 13 g/dL for men in the 2 months preceding inclusion

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 70 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Basic Science
Allocation: Non-Randomized
Intervention Model: Parallel Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Other: IgG group<br>Patient in the IgG group will benefit from a blood sampling of 5 ml (for realization of a basophil activation test) and a blood sampling of 250 ml | Diagnostic Test: IgG group<br>* Will be performed : a blood sampling of 5 ml (for realization of a basophil activation test) a blood sampling of 250 ml<br>|
| Other: IgE group<br>Patient in the IgE will benefit from a blood sampling of 5 ml (for realization of a basophil activation test) and a bone marrow sampling (approx 2 ml) | Diagnostic Test: IgE group<br>* Will be performed : a blood sampling of 5 ml (for realization of a basophil activation test) a bone marrow sampling<br>|
| Other: IgG group + IgE group<br>Patient in the IgG + IgE will benefit from a blood sampling of 5 ml (for realization of a basophil activation test), a blood sampling of 250 ml and a bone marrow sampling (approx 2 ml) | Diagnostic Test: IgG group + IgE group<br>* a blood sampling of 5 ml (for realization of a basophil activation test) a blood sampling of 250 ml a bone marrow sampling<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Obtain sequences of gene producers (VH and VL pairs) coding for the variable part of anti-drug antibodies, thanks to the isolation of B lymphocytes of antibodies directed against the responsible molecule, from blood cells or bone marrow |  | up to 10 years | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Production of recombinant antibodies based on obtained sequences | Purification system (AKTA 25L, Cytiva) | up to 10 years | 
| Evaluate antibodies properties, such as specificity | ELISA | up to 10 years | 
| Evaluate antibodies properties, such as avidity | Interferometry (Octet, ForteBio) | up to 10 years | 
| Ability of antibodies expressed as N297A mutated human IgG1 to reduce or block target curare-induced neuromuscular blockade without an immune response triggered by the antibody drug | Measure of muscle strength of mice | up to 10 years | 
| Ability of antibodies expressed as N297A mutated human IgG1 to reduce or block target curare-induced neuromuscular blockade without an immune response triggered by the antibody drug | Measure of motricity of mice | up to 10 years | 
| Ability of antibodies expressed as N297A mutated human IgG1 to reduce or block target curare-induced neuromuscular blockade without an immune response triggered by the antibody drug | Measure the survival rate of mice | up to 10 years | 
| Ability of antibodies expressed as N297A mutated human IgG1 to reduce or block target curare-induced neuromuscular blockade without an immune response triggered by the antibody drug | Evaluate the absence of occurrence of anaphylactic shock | up to 10 years | 
| Core temperature loss greater than 3°C within 60 minutes after target drug injection only in mice pre-injected with anti-drug IgE or IgG | Rectal probe thermometer for mice | up to 10 years | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Hypersensitivity, IgE, IgG, repertoire, NMBA, anaphylaxis