Biology subjects

Reed, J.

Publications and source records attributed to Reed, J..

24 records · Page 2Linked to original sources

bioRxiv2020

Chromatin conformation capture (Hi-C) sequencing of patient-derived xenografts: analysis guidelines

Sequencing of patient-derived xenograft (PDX) mouse models allows investigation of the molecular mechanisms of human tumor samples engrafted in a mouse host. Thus, both human and mouse genetic material is sequenced. Several methods have been developed to remove mouse sequencing reads from RNA-seq or exome sequencing PDX data and improve the downstream signal. However, for more recent chromatin conformation capture technologies (Hi-C), the effect of mouse reads remains undefined. We evaluated the effect of mouse read removal on the quality of Hi-C data using in silico created PDX Hi-C data with 10% and 30% mouse reads. Additionally, we generated two experimental PDX Hi-C datasets using different library preparation strategies. We evaluated three alignment strategies (Direct, Xenome, Combined) and three processing pipelines (Juicer, HiC-Pro, HiCExplorer) on the quality of Hi-C data. Removal of mouse reads had little-to-no effect on data quality than the results obtained with Direct alignment strategy. Juicer pipeline extracted the most useful information from PDX Hi-C data. However, library preparation strategy had the largest effect on all quality metrics. Together, our study presents comprehensive guidelines on PDX Hi-C data processing.

genomics↗

bioRxiv2020

Haplotype Diversity and Sequence Heterogeneity of Human Telomeres

Telomeres are regions of repetitive nucleotide sequences capping the ends of eukaryotic chromosomes that protect against deterioration, whose lengths can be correlated with age and disease risk factors. Given their length and repetitive nature, telomeric regions are not easily reconstructed from short read sequencing, making telomere sequence resolution a very costly and generally intractable problem. Recently, long-read sequencing, with read lengths measuring in hundreds of Kbp, has made it possible to routinely read into telomeric regions and inspect their structure. Here, we describe a framework for extracting telomeric reads from single-molecule sequencing experiments, describing their sequence variation and motifs, and for haplotype inference. We find that long telomeric stretches can be accurately captured with long-read sequencing, observe extensive sequence heterogeneity of human telomeres, discover and localize non-canonical motifs (both previously reported as well as novel), and report the first motif composition maps of human telomeric diplotypes on a multi-Kbp scale.

genomics↗

bioRxiv2019

Reduced Plasma Ascorbic Acid Levels in Recipients of Myeloablative Conditioning & Hematopoietic Cell Transplantation

Hematopoietic cell transplantation (HCT) conditioned using myeloablative conditioning (MAC) is complicated by end organ injury due to endothelial dysfunction and graft versus host disease. Mucositis and oxidant injury results in micronutrient deficiency. Ascorbic acid (AA) levels were measured in 15 patients undergoing HCT conditioned with MAC (11 allogeneic and 4 autologous HCT). Ascorbate levels declined post conditioning to 27.3 ({+/-}14.1) by day 0 (p <0.05 compared with baseline), reaching a nadir level of 21.5 ({+/-}13.8) on day 14 (p <0.05) post-transplant. Patients undergoing allogeneic HCT continued to have low AA levels to day 60 post transplant, whereas recipients of autologous HCT recovered plasma AA levels to normal. The role of AA in maintaining endothelial function and hematopoietic as well as T cell recovery is provided, developing the rationale for repletion of vitamin C following HCT.

physiology↗

bioRxiv2018

Risk stratification of allogeneic stem cell recipients with respect to the potential for development of GVHD via their pre-transplant plasma lipid and metabolic signature

The clinical outcome of allogeneic hematopoietic stem cell transplantation (SCT) is strongly influenced from the complications arising during the post-transplant immune restoration and has been well studied and described. However, the metabolic status of the recipient pre-transplant also has the potential to influence this outcome and has never been studied before and has the potential to enable risk stratification with respect to the development of transplant associated complications such as graft vs. host disease (GVHD). In order to better understand this aspect of transplant related complications we investigated the pre-transplantation metabolic signature to assess the possibility of pre-transplant risk stratification. This pilot study was composed of 14 patients undergoing myeloablative conditioning followed by either HLA matched related, unrelated donor, or autologous stem cell transplantation. Blood samples were taken prior to transplant and the plasma was comprehensively characterized with respect to its lipidome and metabolome via LCMS and GCMS. The results indicated a significantly pro-inflammatory metabolic profile in patients who eventually developed Graft vs. Host Disease (GVHD). The data revealed 5 potential pre-transplant biomarkers (1-monopalmitin, diacylglycerol (DG) 38:5, DG 38:6, 2-aminobutyric acid, and fatty acid (FA) 20:1) that demonstrated high sensitivity and specificity towards predicting post-transplant GVHD development. The predictive model developed demonstrated an estimated predictive accuracy of risk stratification of 100%, with an Area under the Curve of the ROC of 0.995 with 100%. The likelihood ratio of 1-monopalmitin (infinity), DG 38:5 (6.0) and DG 38:6 (6.0) also demonstrated that a patient with a positive test result for these biomarkers pre-transplant will likely have very high odds of developing GVHD post-transplant. Collectively the data demonstrates the possibility of using pre-transplant metabolic signature for risk stratification of SCT recipients with respect to development of GVHD.

biochemistry↗

bioRxiv2018

T Cell Repertoire Evolution After Allogeneic Bone Marrow Transplantation: An Organizational Perspective

High throughput sequencing (HTS) of human T cell receptors has revealed a high level of complexity in the T cell repertoire. In an attempt to correlate T cell reconstitution with clinical outcomes several measures of T cell repertoire complexity have emerged. However, the associations identified are of a broadly statistical nature, not allowing precise modeling of outcomes based on T cell repertoire development in clinical contexts such as following bone marrow transplantation (BMT). Previous work demonstrated that there is an inherent, mathematically definable order observed in the T cell population that is conserved in a diverse group of donors, and which is perturbed in recipients following BMT. Herein, we use a public database of human leukocyte antigen matched related-donor and recipient T cell receptor (TCR) {beta} sequences to further develop this methodology. TCR {beta} sequencing from unsorted T cells and sorted T cell subsets isolated from peripheral blood samples from BMT donors and recipients show remarkable conservation and symmetry of VJ segment usage in the clonal frequencies, linked to the organization of the gene segments along the TCR locus. This TCR {beta} VJ segment translational symmetry is preserved post-transplant, and even in cases of acute GVHD (aGVHD), suggesting that GVHD occurrence represents a polyclonal donor T cell response to recipient antiges. We also observe that the complexity of the repertoire is significantly diminished after BMT and is not restored even years out post-transplant. The results here provide a new method of quantifying and characterizing post-transplant T cell repertoire reconstitution by further analyzing the mathematical rules governing TCR usage in the context of BMT. This approach may allow for a new means to correlate clinical outcomes with the evolving T cell repertoire post-transplant.

immunology↗

bioRxiv2018

Determining the Quantitative Principles of T Cell Response to Antigenic Disparity: The Case of Stem Cell Transplant Donor-Recipient Exome Wide Mismatching and HLA Bound Alloreactive Peptides

Alloreactivity compromising clinical outcomes in stem cell transplantation is observed despite HLA matching of donors and recipients. This has its origin in the variation between the exomes of the two, which provides the basis for minor histocompatibility antigens (mHA). The mHA presented on the HLA class I and II molecules and the ensuing T cell response to these antigens results in graft versus host disease. In this paper, results of a whole exome sequencing study are presented, with resulting alloreactive polymorphic peptides and their HLA class I and HLA class II (DRB1) binding affinity quantified. Large libraries of potentially alloreactive recipient peptides binding both sets of molecules were identified, with HLA-DRB1 presenting an order of magnitude greater number of peptides. These results are used to develop a quantitative framework to understand the immunobiology of transplantation. A tensor-based approach is used to derive the equations needed to determine the alloreactive donor T cell response from the mHA-HLA binding affinity and protein expression data. This approach may be used in future studies to simulate the magnitude of expected donor T cell response and risk for alloreactive complications in HLA matched or mismatched hematopoietic cell and solid organ transplantation.

immunology↗