Effect of in Vitro Blocking the Common Beta Chain on Cell Viability in Asthma

Effect of in Vitro Blocking the Common Beta Chain on the Function of Blood, Bone Marrow and Sputum Cells Collected From Asthmatic Donors.

This study examines in vitro blockade of signaling through the β-chain, on viability, activation and differentiation of eosinophils and their progenitors collected in sputum, blood and bone marrow samples pre and post-allergen challenge from mild atopic asthmatic subjects.

Study Overview

• Status: Completed
• Conditions: Asthma
• Intervention / Treatment: Other: Allergen

Detailed Description

The experiments will use sputum samples induced from subjects with mild asthma, undergoing allergen inhalation challenges. In general, each sample will be composed of >10% neutrophils and >10% macrophages. Samples collected pre-allergen will have a low frequency of eosinophils and lymphocytes (<1%), however the percentage of eosinophils will increase to approximately 12% following allergen challenges. The sputum samples will be processed in DPBS (without dithiothreitol), and the cell suspension will be adjusted to 1 million cells/ml in DMEM with penicillin and streptomycin. A cytospin will be made for differential cell counts. The mixed cell population at 5 million cells/ml will be incubated for 48 hours at 37 degrees Celcius ± β-chain MAb at a concentration of 100 mcg/ml. After 48 hours the cell culture medium will be removed for assay of cytokines and chemokines by ELISA. Cells will be resuspended in PBS and Binding Buffer (BD Pharmingen, Cat no. 556454),stained for assessments by flow cytometry, and analyzed in duplicate.

Experiments will use blood (80 ml) and bone marrow aspirates (5ml) from atopic asthmatics taken pre and 24hr post allergen challenge. Methylcult micro-culture colonogenic assays will be performed to enumerate outgrowth of Eo/Baso-CFU and GM-CFU from CD34+ cells populations collected from the blood and bone marrow samples. Methylcult assays will be performed with CD34+ enriched cell populations in the presence of IL-5,IL-3 and GM-CSF +/- CSL311. Following 14 days culture, colonies will be enumerated.

• Study Type: Observational
• Enrollment (Actual): 10

Contacts and Locations

Study Locations

• Canada
  • Ontario
    • Hamilton, Ontario, Canada, L8N 3Z5
      • McMaster University Medical Center

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 18 years to 65 years (Adult, Older Adult)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

• Sampling Method: Probability Sample

Study Population

Subjects with mild allergic asthma.

Description

Inclusion Criteria:

• Male and female volunteers 18 through 65 years of age.
• General good health
• Mild to moderate, stable, allergic asthma
• History of episodic wheeze and shortness of breath; FEV1 at baseline at least 70% of the predicted value
• Able to understand and give written informed consent and has signed a written informed consent form approved by the investigator's REB
• Positive methacholine challenge
• Positive skin-prick test to common aeroallergens (including cat, dust mite, grass, pollen)
• Positive allergen-induced airway bronchoconstriction (a fall in FEV1 of at least 20% from baseline)

Exclusion Criteria:

• A worsening of asthma or a respiratory tract infection within 6 weeks preceding study entry
• Use of corticosteroids, immunosuppressives, anticoagulants (warfarin or heparin) within 28 days prior to randomization into the study
• Use of nonsteroidal anti-inflammatory drugs (NSAIDs) within 48 hours of dosing or aspirin with 7 days of dosing
• Have chronic use of any other medication for treatment of allergic lung disease other than short- and intermediate-acting ß2-agonists or ipratropium bromide
• Use of caffeine-containing products or medications for 12 hours or alcohol or over the counter drugs including aspirin, cold and allergy medications for 48 hours or inhaled bronchodilators for 8 hours prior to methacholine and allergen challenges
• Use of tobacco products of any kind currently or within the previous 12 months, or smoking history > 10 pack years.
• Lung disease other than mild to moderate allergic asthma
• Unwillingness or inability to comply with the study protocol for any other reason.

Study Plan

How is the study designed?

Design Details

• Observational Models: Cohort
• Time Perspectives: Prospective

Number of groups / cohorts

1

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Allergen challenge; Determination of the effects of a β-chain monoclonal antibody (MAb) on the function of cells naturally activated by in vivo allergen exposure, from donors with allergen-induced asthma	Other: Allergen; Allergen extract is administered by inhalation.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Sputum Collection	Three baseline sample collections will ensure sufficient numbers of cells are collected for proposed outcomes. Post allergen cells will be examined at time points when these populations are known to be at their highest frequency.	7 hours post allergen

Collaborators and Investigators

• Sponsor: Hamilton Health Sciences Corporation
• Collaborators: CSL Limited
• Investigators: Principal Investigator: Gail M Gauvreau, PhD, Hamilton Health Sciences Corporation

Publications and helpful links

Helpful Links

• Study publication

Study record dates

Study Major Dates

• Study Start: November 1, 2012
• Primary Completion (Actual): September 1, 2014
• Study Completion (Actual): September 1, 2014

Study Registration Dates

• First Submitted: December 17, 2012
• First Submitted That Met QC Criteria: December 27, 2012
• First Posted (Estimate): January 3, 2013

Study Record Updates

• Last Update Posted (Actual): July 13, 2017
• Last Update Submitted That Met QC Criteria: July 10, 2017
• Last Verified: July 1, 2017

More Information

Terms related to this study

• Keywords: Asthma; sputum; blood collection; β-chain signaling; allergen inhalation challenges; bone marrow aspirate; -methacholine challenge
• Additional Relevant MeSH Terms: Respiratory Tract Diseases; Immune System Diseases; Lung Diseases; Hypersensitivity, Immediate; Bronchial Diseases; Lung Diseases, Obstructive; Respiratory Hypersensitivity; Hypersensitivity; Asthma
• Other Study ID Numbers: Bc in vitro study

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: No

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No