Environmental Antigens & HLA Polymorphisms in Hypersensitivity Pneumonitis Patients

Faculty Researcher: Melissa Millerick-May, PhD, Assistant Professor of Medicine at Michigan State University

Project goals: The aim of this multidisciplinary proposal is to determine and isolate antigens important in the etiology of work-related hypersensitivity pneumonitis ( HP), and to identify genetic polymorphisms in HLA class I and II alleles in individuals with HP for the purposes of generating preliminary data to be used to respond to NIH Program Announcement PA-07-300 “Hypersensitivity Pneumonitis — Early Identification and Mechanisms of Disease (R01)”. Information gained from the preliminary work jointly funded by the MSU College of Human Medicine, and the UM-COHSE will increase the likelihood of success of a planned application to NIH to address two of the objectives of the NIH RFA.

Background Hypersensitivity pneumonitis (HP) is a respiratory disease that is the result of an immune response to inhaled organic antigen or low molecular weight chemicals. The prevalence has been reported to be as great as 5-15% among individuals known to have been exposed to antigens associated with the disease.  Diagnosis is difficult as initially symptoms are similar to other respiratory diseases. Treatment involves the use of corticosteroids and complete removal of the patient from the environment in which the antigen is present. The latter is difficult as in the majority of cases the offending antigen cannot be identified through review of the clinical history or from the results the standard serum-HP antigen panel available from a clininal medical laboratory. Clinicians currently have no access to testing that allows them to individualize the panel to actual patient exposure. Identification of the specific causal antigen is the first step in developing workplace controls to prevent the disease. As the majority of individuals exposed to antigens found to be important in HP do not develop disease, researchers have also begun to investigate genetic factors potentially impacting susceptibility. The Human Leukocyte Antigen (HLA) system has recently been determined to be important in individuals with Pigeon Breeders HP. Farmer’s lung is the classic work-related HP. HP has been of particularly concern in Michigan in the last 15 years after HP was described among machinists with exposure to metal working fluids.

Methods We propose to conduct a case-referent study which will compare genetic polymorphisms in the HLA system, and IgG antibody response to antigen important in the case’s HP against a referent group of household members and co-workers. Subjects with HP will be identified from 1) hospital disease reporting records, and 2) patient referrals to the MSU Clinical Center. Two referents will be identified for each case, and will be members of the household and/or co-workers, depending on the suspected location of the offending antigen. Environmental samples will be collected from within and around the case’s workplace and home, with locations determined from a sampling protocol developed from responses to standardized workplace/home questionnaires. The aim of the protocol is to maintain continuity in sampling between cases; however, it will be necessary to tailor the protocol to each case in order to improve the odds of collecting the offending antigen important in the cases’ disease. All cases and referents will have serum analyzed for HLA — A, -B, -DP, -DQ, -DR polymorphisms. Due to budget limitations, only those referents whose associated case reacts to suspected antigens will have their serum tested for IgG antibodies to the standard HP panel and/or antigens collected in environmental samples. Antigens collected will be prepared for use in an immunodiffusion assay with the individual’s (case) serum. Isolation of the specific etiologic antigen from the likely multiple antigens contained in the original assay preparation will take place using standard microbiologic techniques until pure cultures are achieved. Isolates will be tested against the case’s serum using the aforementioned immunodiffusion assay, and extracts producing discrete antigen that previously reacted with sera will be tested utilizing 16S sequencing techniques, potentially allowing for the determination of the full range of microorganisms present in the environmental sample that may not have been cultured/culturable using standard microbiologic techniques. Statistical tests which include descriptive and bivariate analysis and logistic regression will be conducted.

Publications resulting from this project:
Millerick-May ML, Mulks MH, Gerlach J, Flaherty KR, Schmidt SL, Martinez FJ, LeVeque RM, Rosenman KD. Hypersensitivity pneumonitis and antigen identification–An alternate approach. Respiratory Medicine. 2016;112:97-105. doi: 10.1016/j.rmed.2015.09.001.

Grants resulting from this project:
NIH.R01AI121178. Improved Method for Identifying Causative Antigen in Hypersensitivity Pneumonitis. 2016-2021.