Abstract
Spectratyping assays are well recognized as the clinical gold standard for assessing the T cell receptor (TCR) repertoire in haematopoietic stem cell transplant (HSCT) recipients. These assays use length distributions of the hyper variable complementarity-determining region 3 (CDR3) to characterize a patient's T cell immune reconstitution post-transplant. However, whilst useful, TCR spectratyping is notably limited by its resolution, with the technique unable to provide data on the individual clonotypes present in a sample. High-resolution clonotype data are necessary to provide quantitative clinical TCR assessments and to better understand clonotype dynamics during clinically relevant events such as viral infections or GvHD. In this study we developed and applied a CDR3 Next Generation Sequencing (NGS) methodology to assess the TCR repertoire in cord blood transplant (CBT) recipients. Using this, we obtained comprehensive TCR data from 16 CBT patients and 5 control cord samples at Great Ormond Street Hospital (GOSH). These were analyzed to provide a quantitative measurement of the TCR repertoire and its constituents in patients post-CBT. We were able to both recreate and quantify inferences typically drawn from spectratyping data. Additionally, we demonstrate that an NGS approach to TCR assessment can provide novel insights into the recovery of the immune system in these patients. We show that NGS can be used to accurately quantify TCR repertoire diversity and to provide valuable inference on clonotypes detected in a sample. We serially assessed the progress of T cell immune reconstitution demonstrating that there is dramatic variation in TCR diversity immediately following transplantation and that the dynamics of T cell immune reconstitution is perturbed by the presence of GvHD. These findings provide a proof of concept for the adoption of NGS TCR sequencing in clinical practice.
Highlights
Haematopoietic stem cells transplantation (HSCT) utilizes three different stem cell sources; Bone Marrow (BMT), Peripheral Blood (PBSCT) or Umbilical Cord (CBT)
Since the speed at which the immune system recovers after HSCT has a direct bearing on outcome [12] and given that CBT has been shown to result in impaired T cell immune reconstitution [13], we recently demonstrated that the omission of serotherapy can lead to a rapid thymicindependent T cell expansion following CBT [14]
We investigated T cell receptor (TCR) clonal distribution profiles using both the Gini coefficient and the Shannon diversity index for all patients during immune reconstitution
Summary
Haematopoietic stem cells transplantation (HSCT) utilizes three different stem cell sources; Bone Marrow (BMT), Peripheral Blood (PBSCT) or Umbilical Cord (CBT). The stem cell choice usually depends on availability, underlying disease, and clinical status [1]. Initial HSCTs were carried out using bone marrow as a stem cell source, but in 1989, Broxmeyer and colleagues demonstrated that cord blood has similar attributes to bone marrow and contains significant numbers of progenitor cells, and suggested cord blood as a possible alternative source to bone marrow in transplantation [2]. Since CBT has been widely used as a treatment for many diseases, with results from the Cord Blood Transplantation Study (COBLT) demonstrating the high uptake of CBT in the US [4]. Since the incorporation of CBT into standard practice in the UK was recommended [5, 6], the number of CBTs has increased [7]
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