Whole Exome Sequencing to Estimate Alloreactivity Potential Between Donors and Recipients in Stem Cell Transplantation

Whole exome sequencing was performed on HLA-matched stem cell donors and transplant recipients to measure sequence variation contributing to minor histocompatibility antigen differences between the two. A large number of nonsynonymous single nucleotide polymorphisms were identified in each of the nine unique donor-recipient pairs tested. This variation was greater in magnitude in unrelated donors as compared with matched related donors. Knowledge of the magnitude of exome variation between stem cell transplant recipients and donors may allow more accurate titration of immunosuppressive therapy following stem cell transplantation.Comment: 12 pages- main article, 29 pages total, 5 figures, 1 supplementary figur

Sequence homology between HLA-bound cytomegalovirus and human peptides: A potential trigger for alloreactivity.

Human cytomegalovirus (hCMV) reactivation may often coincide with the development of graft-versus-host-disease (GVHD) in stem cell transplantation (SCT). Seventy seven SCT donor-recipient pairs (DRP) (HLA matched unrelated donor (MUD), n = 50; matched related donor (MRD), n = 27) underwent whole exome sequencing to identify single nucleotide polymorphisms (SNPs) generating alloreactive peptide libraries for each DRP (9-mer peptide-HLA complexes); Human CMV CROSS (Cross-Reactive Open Source Sequence) database was compiled from NCBI; HLA class I binding affinity for each DRPs HLA was calculated by NetMHCpan 2.8 and hCMV- derived 9-mers algorithmically compared to the alloreactive peptide-HLA complex libraries. Short consecutive (≥6) amino acid (AA) sequence homology matching hCMV to recipient peptides was considered for HLA-bound-peptide (IC5

HCMV cross-reactivity potential of 77 SCT DRP from whole exome sequencing data using sliding window match analysis.

<p>HCMV cross-reactivity potential of 77 SCT DRP from whole exome sequencing data using sliding window match analysis.</p

Human-CMV short sequence homology in GVHD tissue specific peptide and gene distribution from GTEx analysis (n = 18).

<p>GVHD incidence denotes the specific organs affected in each patient; Peptides, lists the number of unique peptide-HLA complexes matched between human and CMV peptide library; the column, <i>Genes</i> lists the source genes for the aforementioned peptides; <i>GVHD tissue specific peptides</i> lists the number of peptides which bind HLA with an IC50 <500nM, and are expressed in tissues affected by GVHD; <i>GVHD tissue gene expression</i> denotes the number of genes expressed at an RPKM >10 corresponding to the GVHD tissue specific peptides. Note: *- All patients with an asterisk following their numeric representation experienced CMV reactivation prior to GVHD (except Patients 67 and 71, <i>de novo</i> CMV infected) and patients without an asterisk experienced GVHD prior to CMV reactivation. **- human peptides may have overlapping areas of homology yielding a higher number of matches. Immunogenic CMV genes in this context refer to the genes associated with HCMV ORF-specific CD8+ T memory cell responses measured in frequency among CMV seropositive patients previously [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0178763#pone.0178763.ref036" target="_blank">36</a>].</p

CMV+GVHD cross reactivity potential.

<p>Patient-specific peak CMV peptide matches intersecting peak alloreactive peptides (IC50<500 nM) as a cross-reactivity potential stratified by donor type contained within each DRP alloreactive peptide library.</p

GVHD tissue-specific immunogenic CMV peptide matches, CMV reactivation before GVHD patients.

<p>Patient-specific process of cross comparison along with the source genes, IC50 values to indicate inverse binding affinity and GVHD organ specific tissue involvement. *Note: Twelve patients with CMV reactivation/infection before GVHD onset exhibited previously identified immunogenic CMV peptide matches with gene expression specific to the tissues affected by GVHD (above); The filter of immunogenicity provides a connection to T cell reactivity shown <i>in vitro</i> to the listed CMV genes in a separate patient population [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0178763#pone.0178763.ref036" target="_blank">36</a>]; Patient 79 with muscle/fascia GVHD showed no muscle-specific previously <i>known immunogenic</i> CMV peptide matches but still had three relevant CMV peptide matches expressed in the skeletal muscle (not shown); Tissues in parentheses were also affected by GVHD but without immunogenic matches/expression by patient; Patients 67 and 71 experienced <i>de novo</i> CMV infection; All 13 patients exhibited multiple CMV peptide matches with unknown immunogenicity.</p

Matrix depicting T cell clonal cross-reactivity between CMVp-HLA and mHA-HLA.

<p>*-indicates response of the alloreactive T cell clone to a viral pathogen peptide,bound to the same HLA as the mHA and vice versa. For example, TC₁ recognizes, mHA₁ HLA + CMVp₂HLA, TC_1CMV recognizes, mHA₁ HLA + CMVp₁ HLA, and so on.</p