A new tool for rapid and reliable diagnosis of HLA loss relapses after HSCT

Ahci, Müberra; Toffalori, Cristina; Bouwmans, Evelien E.; Crivello, Pietro; Brambati, Chiara; Pultrone, Cinzia; Stempelmann, Karin; Bost, Douglas A.; Mazzi, Benedetta; Beelen, Dietrich; Ciceri, Fabio; Mulder, Wietse; Fleischhauer, Katharina; Vago, Luca

doi:10.1182/blood-2017-05-784306

Cited by 35 publications

(21 citation statements)

References 25 publications

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“…Another advantage of qPCR HC for studying relapse is its flexibility in adding additional assays to target markers of specific interest. For instance, it is possible to include markers for polymorphic HLA to allow discrimination between classical relapses and those that have specifically undergone genomic loss of mismatched HLA [59], a condition described in particular after SCT from haploidentical donors, by side-by-side comparison between chimerism results outside and inside HLA [60,61].…”

Section: Discussionmentioning

confidence: 99%

Clinical Utility of Quantitative PCR for Chimerism and Engraftment Monitoring after Allogeneic Stem Cell Transplantation for Hematologic Malignancies

Ahci

Stempelmann

Buttkereit

et al. 2017

Biology of Blood and Marrow Transplantation

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Although quantitative PCR (qPCR) has been explored for chimerism monitoring after allogeneic stem cell transplantation (SCT), evidence regarding its clinical utility compared with standard short tandem repeat (STR) is still limited. We retrospectively studied commercial qPCR and STR chimerism with respective positivity thresholds of .1% and 1% in 359 peripheral blood (PB) and 95 bone marrow (BM) samples from 30 adult patients after first HLA-matched SCT for myeloid malignancies or acute lymphatic leukemia. Concordance between the 2 methods was 79.5%, with all discordant samples positive in qPCR but negative in STR. Of the latter, sporadic qPCR positivity without clinical correlates was seen mostly in BM samples early post-transplant. In 7 of 21 patients with available follow-up samples in the first months after transplantation, qPCR but not STR revealed low levels (<1%) of sustained host chimerism in PB, reflecting delayed engraftment or persistent mixed chimerism (PMC). These conditions were associated with donor-recipient cytomegalovirus (CMV) serostatus and early CMV reactivation but not with immunosuppressive regimens or clinical outcome. qPCR predicted all 8/8 relapses with samples in the 6 months before onset by sustained positivity in both PB and BM compared with 1/8 relapses predicted by STR mainly in BM. The response kinetics to donor lymphocyte infusions for the treatment of PMC or relapse was shown by qPCR but not STR to be protracted over several months in 3 patients. Our results demonstrate the superior clinical utility of qPCR compared with STR for monitoring subtle changes of host chimerism associated with different clinical conditions, making a case for its use in the clinical follow-up of transplant patients.

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Section: Discussionmentioning

confidence: 99%

Clinical Utility of Quantitative PCR for Chimerism and Engraftment Monitoring after Allogeneic Stem Cell Transplantation for Hematologic Malignancies

Ahci

Stempelmann

Buttkereit

et al. 2017

Biology of Blood and Marrow Transplantation

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“…We showed in this study that the SS‐SBT method could detect the 6pLOH phenomenon by precise calculations of the depth ratios, supporting that this high‐resolution method is considered superior to the conventional methods such as PCR‐SSOP assay, single nucleotide polymorphism (SNP) array analysis, simple HLA typing method using qPCR, and HLA typing kit using NGS . This method is also a cost‐saving method compared to genome‐wide techniques, because SS‐SBT can work with a smaller amount of genomic DNA (1 ng per sample) than other methods such as SNP array analysis (250 ng per sample) .…”

Section: Hla Allele and Read Sequence Information Of The Aplastic Anementioning

confidence: 55%

Detection of 6pLOH in an aplastic anemia patient by in phase HLA genotyping

Kikkawa

Shiina

Shigenari

et al. 2020

HLA

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Recent studies have reported loss of heterozygosity in the chromosome 6p arms (6pLOH) of acquired aplastic anemia (AA) patients, and in tumor cells trying to escape the autoimmune system. We thus sought to establish detection methods for LOH to investigate the mechanisms underlying AA and tumor immunity. Herein, we report our evaluation of 6pLOH in a patient with severe AA patient using super‐high resolution, single‐molecule, sequence‐based typing (SS‐SBT). The highest ratios of 6pLOH detection were observed during the patient's treatment with granulocyte colony stimulating factor (G‐CSF). This result suggested that most of the neutrophil precursor cells stimulated by G‐CSF already had LOH in the HLA lesion. The SS‐SBT method is a simple NGS method that provides complete HLA allele coverage.

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“…By this mechanism, leukaemic cells escape being killed by alloimmune cell immunotherapy/GvL (Vago et al, ). As haploidentical aHSCT has been on a sharp increase in recent years, observations of post‐transplant somatic HLA mutation (namely LOH) are common and of diagnostic value (Ahci et al, ). In a perspective manner, knowledge of the mechanisms by which the GvL effect helps to combat haematological malignancies by directly targeting the mismatched HLA expressed on leukaemic cells may be applied to the process of donor selection in particular cases (Ikegame et al, ).…”

Section: Discussionmentioning

confidence: 99%

HLA‐B gene somatic insertion/deletion mutations in patients with acute myelogenous leukaemia

Königova

Skoumalova

Onderkova

et al. 2018

Int J Immunogenetics

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Loss of heterozygosity is considered to be the most common type of tumour-specific somatic mutation of the human leucocyte antigens (HLA) genes in patients with haematological malignancies. Nevertheless, subtle DNA sequence changes, namely short insertions/deletions, may also abolish the expression of HLA molecules and interfere with routine HLA typing. Two male patients with acute myelogenous leukaemia (AML) were indicated for the search of a suitable donor for allogeneic haematopoietic stem cell transplantation (aHSCT). The patients and their relatives were initially HLA typed by serological and DNA techniques at a low-resolution level. The HLA high-resolution (HR) type was obtained by means of sequencing-based typing (SBT). In both cases, anomalous frameshifts in the sequence were observed in the HLA-B gene, namely in exon 3 (Case 1, heterozygous deletion of two bases) and exon 4 (Case 2, heterozygous insertion of two bases). In the second case, the insertion variant was associated with a loss of HLA-B8 expression. To reveal whether these sequence patterns may be caused by somatic mutations in the malignant cells, blood sample in remission (Case 1) and buccal swab sample (Case 2) were collected from the patients. In an important manner, the SBT in these germline samples revealed common HLA-B*07:02,*15:01 (Case 1) and HLA-B*08:01,*35:02 (Case 2) types with no evidence for the sequence alteration observed in the initial samples. In conclusion, the insertion/deletion sequence variants of the HLA-B gene in two patients were limited to the initial blood samples with a substantial proportion of AML cells and thus may be attributed to the somatic mutation in the malignant cells. HLA somatic mutations should be taken into account in patients with haematological malignancies to prevent HLA mistyping and inappropriate selection of an aHSCT donor.

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A new tool for rapid and reliable diagnosis of HLA loss relapses after HSCT

Cited by 35 publications

References 25 publications

Clinical Utility of Quantitative PCR for Chimerism and Engraftment Monitoring after Allogeneic Stem Cell Transplantation for Hematologic Malignancies

Clinical Utility of Quantitative PCR for Chimerism and Engraftment Monitoring after Allogeneic Stem Cell Transplantation for Hematologic Malignancies

Detection of 6pLOH in an aplastic anemia patient by in phase HLA genotyping

HLA‐B gene somatic insertion/deletion mutations in patients with acute myelogenous leukaemia

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