In the age of big data, thoughtful management and harmonization of clinical metadata and sample processing in translational research is a critical for effective data generation, integration, and analysis. These steps enable the cutting edge discoveries and enhance overall conclusions that may come from complex multi-omic translational research studies. The focus of my thesis has been on harmonizing the clinical metadata collected as part of the lung Pre Cancer Genome Atlas (PCGA) in addition to expanding the use of banked samples. The lung PCGA study included longitudinal collected samples and data from participants in a high-risk lung cancer-screening program at Roswell Park Comprehensive Cancer Center (Roswell) in Buffalo, NY. Clinical metadata for this study was collected over many years at Roswell and subsets of this data were shared with Boston University Medical Campus (BUMC) for the lung PCGA study. During the study, additional clinical metadata was acquired and shared with BUMC to complement the analysis of genomic profiling of DNA and RNA, as well as protein staining of tissue. With regards to the PCGA study, my thesis has two aims: 1) Curate the clinical metadata from received from Roswell during the PCGA study to enhance both its accessibility to current investigators and collaborators and reproducibility of results 2) Test methods to isolate DNA from remnant samples to expand the use of banked samples for genomic profiling. We hypothesized that the accomplishment of these goals would allow for increased use of the clinical metadata, enhanced reproducibility of the results, and expansion of samples available for DNA sequencing
The clinical metadata received from Roswell was consolidated into a singular source that is continually updated and available for export for future research use. These metadata management efforts led to increased use among the members of our laboratory and collaborators working with the lung PCGA cohort. Additionally, the curation of metadata has allowed for improved analysis, reproducibility, and increased awareness of the current inventory of remaining samples.
During the process of lung PCGA clinical metadata curation, physical inventory of the remaining samples revealed remnant organic phase samples. Therefore, in addition to my work associated with clinical metadata, the second goal of my thesis focuses on DNA isolation from remnant banked biological samples from the lung PCGA cohort. In the first phase of the lung PCGA, nucleic acid isolation of RNA was intended to be collected exclusively from fresh frozen endobronchial biopsy samples, and formalin-fixed paraffin embedded (FFPE) biopsy samples were to be used for DNA isolation. DNA isolation from the FFPE samples was unsuccessful. However, from the RNA isolation, the remaining organic phase was banked and could potentially serve as a source of DNA. The organic phase of this isolation contained cell debris, proteins, and, as previously mentioned, DNA. We hypothesized that current protocols for organic phase DNA isolation might yield adequate quantities of DNA for genomic profiling. Utilizing immortalized cell culture lines to establish methodology, numerous organic phase DNA isolation protocols were tested. During subsequent validation using the remaining organic phase samples from the lung PCGA cohort, the protocol yielded varied results, suggesting that further optimization to increase DNA purity is required. The ability to isolate DNA from these valuable samples will enhance progress in the lung PCGA study. The aims of this thesis involving curation of clinical metadata and generation of additional DNA samples for DNA profiling has had significant impact on the PCGA study and future expansions of this work.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/41714 |
| Date | 23 November 2020 |
| Creators | Potter, Austin |
| Contributors | Mazzilli, Sarah, Beane, Jennifer |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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