Genome binding/occupancy profiling by high throughput sequencing
Summary
Here, we define the landscape and dynamics of active regulatory DNA in cutaneous T cell lymphoma (CTCL) by Assay of Transposase-Accessible Chromatin (ATAC-seq). Analysis of 111 human CTCL and control samples revealed extensive chromatin signatures that distinguished leukemic vs. non-leukemic (host) CD4+ T cells in CTCL patients, vs. CD4+ T cells in healthy donors. We identify three dominant patterns of transcription factor (TF) activation that drive leukemia regulomes, as well as TF deactivations that alter host T cells in CTCL patients. Clinical response to histone deacetylase inhibitors (HDACi) is strongly associated with a concurrent gain in chromatin accessibility. HDACi causes distinct chromatin responses in leukemic and host CD4+ T cells, reprogramming host T cells toward normalcy. These results provide a foundational framework to study personal regulomes in human cancer and epigenetic therapy.
Overall design
We examined chromatin structure using ATAC-seq in purified human CD4+ T cells in 30 samples from 10 health donors and 81 samples from 14 patiets with cutaneous T cell leukemia (CTCL), treated with HDACi anti-cancer drugs, such as Vorinostat and Romidepsin. Each donor provided cells per time point via a single standard 5 ml blood draw, and CD4+ T cells were isolated applying RosetteSep Human CD4+ T Cell Enrichment Cocktail (Experimental Procedures). Because CTCL is typically characterized by a dominant CD4+ T cell clone bearing an unique T cell receptor, we purified CTCL leukemic cells from patients (defined by CD4+, CD26-, and T cell receptor V-beta clone+) vs. host CD4+ T-cell (defined by CD4+, CD26-, V-beta clone-) from the same patients by fluorescence activated cell sorting (FACS). Leukemic, host and bulk T cells were obtained from 10 out of 15 patients who had detectable V-beta clone, and only bulk T cells were obtained for the remaining 5 patients without detectable V-beta clone. Although number and proportion of leukemic and host cells varies depending on the stage and drug response of each individual, especially at a later stage of drug treatment, we were able to obtain at least 50,000 CD4+ T cells per sample.
Please note that the indicated 'Day' (in the sample title) for Normal donors means when the blood was drew. For example “Normal_Donor1_Day38_Rep2” means this sample was drew on day 38. Day for Patients means days after the first drug treatment. For example “BulkCTCL_Patient1366_Romi_Day28” means this is a Bulk cell sample from Patient 1366, who was treated with Romidepsin, and the blood was drew on Day 28 according to the first day of treatment, and on that day, the patient was also treated with the same drug.
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