Efficient identification of T‐cell clones associated with graft‐versus‐host disease in target tissue allows for subsequent detection in peripheral blood

Abstract

Summary Graft‐versus‐host disease (GVHD) causes severe morbidity and mortality in allogeneic haematopoietic stem cell transplantation (HSCT) because of destruction of recipient tissues by donor alloreactive T cells. We hypothesized that GVHD‐specific T‐cell clones are expanded within affected tissue of HSCT patients and can also be detected in blood at the time of active disease. A multiplex polymerase chain reaction (PCR) was used to amplify T‐cell receptor (TCR) variable beta (VB) chain rearrangements in skin biopsies from eight allogeneic HSCT patients. Molecular analysis of the complementarity‐determining region 3 (CDR3) of amplified products defined expanded, potentially disease‐associated ‘clonotypes’ and enabled the design of clonotype‐specific PCR assays. We detected immunodominant clones in seven of eight GVHD‐positive skin biopsies. In serial skin biopsies from the same patient, the identical clone was found in each biopsy. In a patient who underwent two successive HSCTs from different donors, distinct clones were identified for each engraftment. Using clonotypic PCR assays, individual tissue‐derived clones could be identified in peripheral blood samples obtained during active GVHD. We hypothesize that clonotypic sequences derived from target tissue can serve as markers for GVHD and may have utility in diagnosis and monitoring response to therapy, as well as enable future therapies targeted against pathogenic clones.

Alloreactive microenvironment after human hematopoietic cell transplantation induces genomic alterations in epithelium through an ROS-mediated mechanism: in vivo and in vitro study and implications to secondary neoplasia

“…There are several possible explanations for that. Although the same allospecific T-cell clones have been identified both in GvHD lesions and peripheral blood, 35 the frequency of alloreactive clonotypes is much lower in the circulation as compared to target tissues. 36,37 In addition, the degree of human leukocyte antigen mismatches in random MLCs results in higher level of T cell activation as compared to the human leukocyte antigen matched setting of the donor/recipient pair.…”

Molecular Analysis of Alloreactive CTL Post-Hemopoietic Stem Cell Transplantation

“…Previously, we characterized the VB utilization spectrum and immunodominant clonotypes among CD8 ϩ T cells in 29 HSCT patients and in biopsies from nine patients posttransplant (8,9). The frequency of the CD8 and biopsy-derived clonotypes in blood posttransplant and the extent of clonotype sharing between patients were determined by sequencing and clonotype-specific PCR.…”

Double‐negative regulatory T cells induce allotolerance when expanded after allogeneic haematopoietic stem cell transplantation

“…As an alternative, various molecular markers of disease activity that do not rely on knowledge of the precipitating antigens have been developed (O'Keefe et al, 2007). We and others have previously shown that the most permissive of these antigens may lead to immunodominant clonal expansions as measured by flow cytometry, and molecular analysis of the T-cell receptor (TCR) variable beta chain (Vb) utilization pattern can serve as an indicator of GvHD (Michalek et al, 2003;Beck et al, 2005;O'Keefe et al, 2007). In our current study, we assessed the Vb spectrum to better quantify the degree of alloreactivity that may develop after HSCT in humans.…”