Genetics of Lung Disease (Exhaled Breath DNA Methylation in Lung Carcinogenesis)
Trial Snapshot
- Phase
- Not Applicable
- Status
- Recruiting
- Enrollment
- 2,000
- Locations
- 1
- Primary Endpoint
- Number of smoker and non-smoker participants demonstrating somatic DNA mutations as evidenced by mutation burden
Study Overview
Brief Summary
The lung is a privileged organ; blood does not reflect most lung processes well, if at all. Therefore, for population scale diagnostics, the investigator team is developing non-invasive portals to the lung, for eventual early detection/risk assessment and diagnostic purposes. However, large macromolecules are not likely suspended nor readily detected in the breath. In particular, genomic DNA in the breath condensate (EBC) is very sparse, and where present, generally highly fragmented, not readily amenable to sequencing based assessments of DNA somatic mutation burden or distribution. Because gDNA (and protein) is challenging to obtain non-invasively from EBC, the study team considered alternative surrogate lower airway specimens. Cough capture is rarely done, and the investigator team is in the process of optimizing its collection. Importantly, the team will be evaluating how much of coughed material is from saliva contamination. Additionally, analyzing material that is target captured by capturing deep lung extracellular vesicles (EVs) using immobilized CCSP/SFTPC antibodies targeting EVs from distal bronchiole Club and alveolar type 2 cells could circumvent the mouth contamination problem, leaving a non-invasive portal to the deep lung suitable for large molecules, and in turn suitable for myriad epidemiologic and clinical applications.
The investigator team proposes (Aim 1) to pursue optimizing cough collection, and testing the efficacy and practicality of partitioning cough specimen for deep-lung specific extra-cellular vesicles (EVs). This cough specimen will be compared to that from invasively collected deep lung samples BAL/bronchial brushings, and to the potential contaminating mouth rinse, all from the same individuals. (Aim 2) The study team initially proposes to examine these cough specimens for somatic mutations by SMM bulk sequencing for single nucleotide variation, developed in the Vijg/Maslov labs. Finally, the investigator team will (Aim 3) test all airway specimens (cough, mouthwash and BAL) for lung surrogacy of cough, using proteins known to be specific for lung, as opposed to oral cavity/saliva, in the Sidoli/proteomics core.
The investigator team envisions that the translational impact of non-invasively obtained DNA or protein markers could allow for more rapid acute clinical diagnoses, and facilitate precision prevention and/or early detection of many acute and chronic respiratory disorders, including lung cancer, asthma and COPD, acute and chronic infectious diseases, and indeed systemic disorders of inflammation and metabolism.
Detailed Description
Hypothesis: Cough can serve as a valid non-invasive surrogate for the lung for large molecules such as DNA and proteins, potentially for a variety of clinical and public health purposes.
Proposal in Brief: For development of an epidemiological and clinically-applicable platform for large molecules captured uniquely from deep lung, the investigator team will pursue a new approach that includes cough collection, and testing the efficacy and practicality of partitioning cough specimen for deep-lung specific extra-cellular vesicles (EVs). This cough specimen will be compared to that from invasively collected deep lung samples BAL/bronchial brushings, and to the potential contaminating mouthrinse, all from the same individuals. The study team initially proposes to examine these crude cough specimens, and then EV partitioned airway samples, for somatic mutations by SMM bulk sequencing for somatic mutations, state-of-art techniques developed in the Vijg/Maslov labs and for proteins known to be specific for lung, as opposed to oral cavity/saliva, in the Sidoli/proteomics core.
Specific Aims:
- Cough will be captured by aerochamber or equivalent/optimized device in 5 healthy volunteers, and in 5 current smokers, in parallel with mouthrinse and with BAL. Lung-specific EVs from these same samples via deep lung capture (Loudig lab) will be performed in parallel.
- Cough will undergo DNA mutation analyses using single molecule mutation sequencing (SMM-seq).
- Cough, mouthwash and BAL will undergo proteomic analysis using LC-MS in our Core facility.
Aims 1 and 2 will be studied in a cross-sectional manner, enrolling a convenience sample of PLWHA smokers attending the Montefiore Medical Center (MMC) Infectious Diseases (ID) Clinic. During Aim 1, group EBC DNA methylation and microRNA and metabolomics patterns will be determined by high resolution tagged bisulfite genomic sequencing of three tumor suppressor genes (DAPK, RASSF1A, PAX5B), according to smoking status (current, former, never), and lifelong dose (pack-years) in a sample of HIV-infected smokers. Aim 2 will pilot the comparison of EBC DNA methylation and microRNA and metabolomics patterns from HIV infected individuals to that from uninfected individuals, collected in other projects funded by other mechanisms. Aim 3 will employ a prospective design, collecting two specimens over time from a subset of the overall study sample to examine the stability of EBC DNA and microRNA and metabolomics patterns methylation patterns over time in HIV-infected smokers.
Study Design
- Study Type
- Observational
- Observational Model
- Case Control
- Time Perspective
- Cross Sectional
Eligibility Criteria
- Ages
- 21 Years to — (Adult, Older Adult)
- Sex
- All
- Accepts Healthy Volunteers
- No
Inclusion Criteria
- •Age: minimum age of 21 years
- •Gender: Male and Female adults
- •Ethnicity: All ethnic groups and races
- •Subjects undergoing bronchoscopy for diagnostic purposes or therapy
- •Subjects without a known diagnosis of lung cancer who are not scheduled for lung tissue collection procedures
- •Subjects with a known or suspected diagnosis of asthma or COPD and are scheduled for a visit at Montefiore Asthma or COPD Center and individual practices, and/or in-hospital with exacerbation
Exclusion Criteria
- •Bleeding diathesis or known coagulopathy precluding clinically indicated biopsy (e.g., INR>1.3, PTTr>1.3), thrombocytopenia <50,000, uremia with serum creatinine >3.0
- •Unstable angina
- •Recent myocardial infarction (within 3 months),
- •Uncontrolled congestive heart failure or severe pulmonary hypertension (mean PAP>75 mmHg)
Arms & Interventions
Bronchoscopy subjects, current or former smokers
Bronchoscopy subjects >=21 yo, current or former smokers
Intervention: Observational only, all subjects; measure DNA mutation and proteomic survey. (Other)
Bronchoscopy subjects, never smokers
Bronchoscopy subjects >=21 yo, never smokers
Intervention: Observational only, all subjects; measure DNA mutation and proteomic survey. (Other)
Outcomes
Primary Outcomes
Number of smoker and non-smoker participants demonstrating somatic DNA mutations as evidenced by mutation burden
Time Frame: Up to 30 minutes for collection of all airway samples
Somatic DNA analysis will be conducted using Single Molecule Mutation sequencing (SMM-seq) to identify mutations in EV-partitioned cough samples. The number will be determined by statistical testing using Fishers Exact Test to determine if there is a nonrandom association between the two variables
Aggregate Median Mutation Rate in smoker and non-smoker participants
Time Frame: Up to 30 minutes for collection of all airway samples
Somatic DNA analysis will be conducted using Single Molecule Mutation sequencing (SMM-seq) to identify mutations in EV-partitioned cough samples. The aggregate median mutation rate (somatic mutation burden) in the cough from the group of smokers, as compared to that of non-smoking individuals, will be determined by statistical T-test.
Number of smoker and non-smoker participants demonstrating altered protein expression
Time Frame: Up to 30 minutes for collection of all airway samples
Proteomic analysis will be conducted via LC-MS on the collected and EV-partitioned cough samples, MW samples, and BAL samples, respectively. To evaluate cough surrogacy for the deep lung (BAL) specimen, Spearman correlations of the most highly expressed 80 proteins among the three specimen types will be compared. The number of participant specimens demonstrating Spearman inter-tissue correlation values in excess of their corresponding threshold (0.3) will be tabulated.
Proteomic signature comparison in smoker and non-smoker participants
Time Frame: Up to 30 minutes for collection of all airway samples
Proteomic analysis will be conducted via LC-MS on the collected and EV-partitioned cough samples, MW samples, and BAL samples, respectively. The proteomic signature of the most highly expressed 80 proteins in each of three specimen types will be compared between the current smoker and the non-smoker participants using PCA.
Secondary Outcomes
No secondary outcomes reported