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dc.contributor.authorRobinson, Hayley
dc.date.accessioned2019-12-10T08:21:43Z
dc.date.created2019-03
dc.date.issued2018-10-25
dc.date.submitted2019
dc.identifier.citationRobinson, Hayley. 2019. Deep Next-Generation Sequencing Enables Mutational Profiling of Classical Hodgkin Lymphoma. Master's thesis, Harvard Extension School.
dc.identifier.urihttp://nrs.harvard.edu/urn-3:HUL.InstRepos:42004070*
dc.description.abstractEffective treatments are available for classical Hodgkin Lymphoma (cHL); however, for many patients the success of initial treatment is often followed by a high rate of treatment-related conditions including infertility and secondary malignancies. Due to the rarity of the malignant Hodgkin and Reed/Sternberg (HRS) cells in cHL, molecular genotyping has not been successfully implemented. The inability to obtain the mutational profile currently precludes genotyping of cHL which is informative for clinical management in numerous other cancers. The genetic pathways implicated in the pathogenesis and oncogenic activity of cHL have been described and leveraged for the selection of first-line therapeutics. However, a comprehensive evaluation of relevant mutations in the SOCS1 gene remain elusive despite its role in cHL being verified in several studies. To assess the feasibility of next-generation sequencing (NGS) for genotyping in cHL, an NGS panel employing Anchored Multiplex PCR (AMP) was designed to target and sequence three relevant genes: B2M, JAK2, and SOCS1. The primary endpoint was suitable detection of low-lying mutations in the rare HRS cells which could be adequately detected by a clinical-grade laboratory assay. Secondary endpoints were the prevalence of mutations and establishing the limit of detection (LOD). Sufficient average sequencing depth for SOCS1 was achieved in normal feasibility samples at 236 total reads. In cHL cases sequencing coverage over genes B2M, JAK2 and SOCS1 was achieved at an average of 503 reads, 462 reads, and 191 reads respectively, which demonstrated the technical performance of the method. Diagnostic performance was satisfactory with mutations successfully detected in each of the three targeted genes for all sequenced cHL cases, with an established LOD of 5% allelic fraction establishing the analytical performance. Despite the technical barriers imposed by the rarity of tumor cells and the necessity to test DNA extracted from tissue biopsies with variable quality, the application of this method can reliably detect somatic mutations with a sufficient level of high quality sequencing coverage across the coding regions of B2M, JAK2, and SOCS1. The demonstrated levels of technical, diagnostic, and analytical performance are adequate to move this approach forward towards implementation of a clinical-grade laboratory assay.
dc.description.sponsorshipBiotechnology
dc.format.mimetypeapplication/pdf
dash.licenseLAA
dc.subjectclassical Hodgkin lymphoma, next-generation sequencing, genotyping
dc.titleDeep Next-Generation Sequencing Enables Mutational Profiling of Classical Hodgkin Lymphoma
dc.typeThesis or Dissertation
dash.depositing.authorRobinson, Hayley
dc.date.available2019-12-10T08:21:43Z
thesis.degree.date2019
thesis.degree.grantorHarvard Extension School
thesis.degree.grantorHarvard Extension School
thesis.degree.levelMasters
thesis.degree.levelMasters
thesis.degree.nameALM
thesis.degree.nameALM
dc.contributor.committeeMemberDenkin, Steven
dc.contributor.committeeMemberLennerz, Jochen K.
dc.type.materialtext
thesis.degree.departmentBiotechnology
thesis.degree.departmentBiotechnology
dash.identifier.vireo
dash.author.emailhayley.mt@gmail.com


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