+Model

TRACLI-2958; No. of Pages 3 ARTICLE IN PRESS

Disponible en ligne sur

ScienceDirect
www.sciencedirect.com

Transfusion Clinique et Biologique xxx (2017) xxx–xxx

Research perspectives

Editorial: Next-generation sequencing technology a new tool for killer cell

immunoglobulin-like receptor allele typing in hematopoietic stem cell
transplantation
Éditorial : séquençage nouvelle génération, un nouvel outil pour typer les allèles killer cell
immunoglobulin-like receptor en greffes de cellules souches hématopoïétiques
B. Maniangou a,b,∗ , C. Retière a,b , K. Gagne a,b,c
aÉtablissement français du sang (EFS) Pays-de-la-Loire, laboratoire de recherche, 34, boulevard Jean-Monnet, 44011 Nantes, France
b Inserm U1232, CNRS, CNRS ERL, centre de recherche en cancérologie et immunologie Nantes Angers (CRCINA), équipe 1, 8, quai Moncousu, 44007 Nantes,
France
c Établissement français du sang Pays-de-la-Loire, laboratoire HLA, 34, boulevard Jean-Monnet, 44011 Nantes cedex, France

Abstract
Killer cell Immunoglobulin-like Receptor (KIR) genes are a family of genes located together within the leukocyte receptor cluster on human
chromosome 19q13.4. To date, 17 KIR genes have been identified including nine inhibitory genes (2DL1/L2/L3/L4/L5A/L5B, 3DL1/L2/L3), six
activating genes (2DS1/S2/S3/S4/S5, 3DS1) and two pseudogenes (2DP1, 3DP1) classified into group A (KIR A) and group B (KIR B) haplotypes.
The number and the nature of KIR genes vary between the individuals. In addition, these KIR genes are known to be polymorphic at allelic level
(907 alleles described in July 2017). KIR genes encode for receptors which are predominantly expressed by Natural Killer (NK) cells. KIR receptors
recognize HLA class I molecules and are able to kill residual recipient leukemia cells, and thus reduce the likelihood of relapse. KIR alleles of
Hematopoietic Stem Cell (HSC) donor would require to be known (Alicata et al. Eur J Immunol 2016) because the KIR allele polymorphism may
affect both the KIR+ NK cell phenotype and function (Gagne et al. Eur J Immunol 2013; Bari R, et al. Sci Rep 2016) as well as HSCT outcome
(Boudreau et al. JCO 2017). The introduction of the Next Generation Sequencing (NGS) has overcome current conventional DNA sequencing
method limitations, known to be time consuming. Recently, a novel NGS KIR allele typing approach of all KIR genes was developed by our team
in Nantes from 30 reference DNAs (Maniangou et al. Front in Immunol 2017). This NGS KIR allele typing approach is simple, fast, reliable,
specific and showed a concordance rate of 95% for centromeric and telomeric KIR genes in comparison with high-resolution KIR typing obtained
to those published data using exome capture (Norman PJ et al. Am J Hum Genet 2016). This NGS KIR allele typing approach may also be used in
reproduction and to better study KIR+ NK cell implication in the control of viral infections.
© 2017 Elsevier Masson SAS. All rights reserved.

Keywords: High-resolution killer cell immunoglobulin-like receptor typing; Allele polymorphism; Next-generation sequencing; Natural killer cells; Hematopoietic
stem cell transplantation

Résumé
Les gènes Killer cell Immunoglobulin-like Receptor (KIR) sont une famille de 15 gènes, localisés chez l’homme sur le bras long du chromosome
19. Ces gènes KIR peuvent être inhibiteurs (2DL1/L2/L3/L4/L5A/L5B, 3DL1/L2/L3) ou activateurs (2DS1/S2/S3/S4/S5, 3DS1) et sont organisés
en deux groupes d’haplotypes : haplotype A ou B. Le nombre et la nature des gènes KIR présents varient selon les individus. De plus, ces gènes
KIR sont connus pour être polymorphes au niveau allélique (907 allèles décrits en juillet 2017). Les gènes KIR codent pour des récepteurs KIR

∗ Corresponding author. EFS Pays de la Loire, laboratoire de recherche, 34, boulevard Jean-Monnet, 44011 Nantes, France.
E-mail addresses: [email protected] (B. Maniangou), [email protected] (C. Retière), [email protected] (K. Gagne).

http://dx.doi.org/10.1016/j.tracli.2017.07.005
1246-7820/© 2017 Elsevier Masson SAS. All rights reserved.

Please cite this article in press as: Maniangou B, et al. Editorial: Next-generation sequencing technology a new tool for killer
cell immunoglobulin-like receptor allele typing in hematopoietic stem cell transplantation. Transfusion Clinique et Biologique (2017),
http://dx.doi.org/10.1016/j.tracli.2017.07.005
+Model
TRACLI-2958; No. of Pages 3 ARTICLE IN PRESS
2 B. Maniangou et al. / Transfusion Clinique et Biologique xxx (2017) xxx–xxx

inhibiteurs ou activateurs, exprimés principalement sur les cellules tueuses naturelles (NK). Les récepteurs KIR ont pour ligands les molécules
HLA de classe I et sont capables de lyser les cellules leucémiques résiduelles des patients après greffe de cellules souches hématopoïétiques
(CSH). Le contenu en allèles KIR de chaque donneur de CSH nécessiterait d’être connu (Alicata et al. Eur J Immunol 2016) car ce polymorphisme
allélique KIR peut affecter le phénotype et la fonction des cellules NK KIR+ (Gagne et al. Eur J Immunol 2013; Bari R, et al. Sci Rep 2016) ainsi
que le devenir des greffes de CSH (Boudreau et al. JCO 2017). L’arrivée de nouvelles technologies de séquençage à haut débit (NGS) a permis
d’aller au-delà des limites des techniques de séquençages conventionnelles, connues pour prendre plus de temps car spécifique d’un seul locus KIR.
Récemment, une nouvelle approche NGS de typage allélique de tous les gènes KIR en entier a été développée par notre équipe nantaise à partir
de 30 ADNs de référence (Maniangou et al. Front in Immunol 2017). Cette approche NGS.KIR est simple, rapide, fiable, spécifique et a montré
une concordance des résultats alléliques KIR proche de 95 % avec ceux effectués sur les mêmes ADN dans une étude de l’exome aux États-Unis
(Norman PJ et al. Am J Hum Genet 2016). Cette approche NGS de typage des allèles KIR peut aussi être utilisée en reproduction et pour étudier
plus finement l’implication des cellules NK KIR+ dans le contrôle des infections virales.
© 2017 Elsevier Masson SAS. Tous droits réservés.

Mots clés : Typage allélique KIR ; Polymorphisme allélique ; Séquençage nouvelle génération ; Cellules tueuses naturelles NK ; Greffes de cellules souches
hématopoïétiques

Killer cell Immunoglobulin-like Receptor (KIR) genes are a cases because KIR genes are polymorphic at the allelic level.
family of genes located together within the leukocyte recep- Currently, 907 KIR alleles are described in the Immuno Poly-
tor cluster on human chromosome 19q13.4. To date, 17 KIR morphism Database for KIR genes (IPD – KIR). KIR allelic
genes have been identified including: nine inhibitory genes variation may affect the NK KIR+ cell phenotype and function
(2DL1/L2/L3/L4/L5A/L5B, 3DL1/L2/L3), six activating genes as demonstrated for both KIR3DL1/S1 [8] and KIR2DL2/L3 [9]
(2DS1/S2/S3/S4/S5, 3DS1) and two pseudogenes (2DP1, 3DP1) genes. Indeed, the KIR3DL1/S1 gene is one of the most polymor-
[1]. These genes are further classified into group A (KIR A) and phic KIR and the allotypes can display high or low cell surface
group B (KIR B) haplotypes [2]. KIR A haplotypes consist of receptor density as well as intracellular retention [10]. Recently,
a fixed number of genes, whereas the KIR B haplotypes have the importance of KIR3DL1/HLA-B subtype combinations in
variable gene content with one or more KIR B-specific genes AML patients treated with HSCT has also been demonstrated
such as: KIR2DS1, S2, S3, S5, KIR2DL2, and KIR2DL5. KIR [11]. Alicata et al. have further suggested that KIR allele typing
genes encode for receptors, which are predominantly expressed should become a standard practice when selecting HSC donors
by Natural Killer (NK) cells. Functionally, these receptors [12]. Clearly, the arguments above provide a basis for contin-
may be inhibitory or activating and have also been identified uing research on the impact of KIR allele polymorphisms on
in subsets of CD4, CD8 and ␥␦ T cells [3,4]. NK cells are HSCT outcome. However, tools to study KIR allele polymor-
innate immune components able to kill target cells and produce phisms of all KIR genes are currently lacking because most of
cytokines without requiring prior sensitization. After allogeneic them are locus specific, targeting only limited polymorphism and
Hematopoietic Stem Cell Transplantation (HSCT), NK cells are often time consuming. The introduction of the Next Gener-
are the first lymphocyte population to reconstitute the recipi- ation Sequencing (NGS) has considerably changed the vision
ent’s immune system before T cell reconstitution [5]. During of biologists about the immune system exploration within the
this period, NK cells are able to kill residual recipient leukemia field of immunology [13]. This NGS technology has the capac-
cells, and thus reduce the likelihood of relapse. The Graft-versus- ity to increase sequencing throughput by attaching millions of
Leukemia (GvL) effect is thought to be mediated mostly by KIR DNA fragments to a solid surface or support, and simultaneously
receptors expressed on NK cells in the early months post-HSCT. sequencing them. Current NGS platforms generally involve two
In 2002, Ruggeri et al. were the first group to report a great- steps: template preparation and sequencing. In the first step, the
est beneficial effect of donor-recipient KIR ligand mismatching DNA sample is fragmented to construct libraries, depending on
[6]. Patients with acute myeloid leukemia (AML) were split into the platform to be used. The fragment libraries are subsequently
two groups and treated with T-cell depleted HLA-haploidentical ligated to end templates with specific adaptor oligonucleotides.
HSCT. In those grafted with a KIR ligand mismatch, the relapse They are then dual-indexed and injected for sequencing either
rate was 0% at 5 years, compared to those grafted with matched in paired-end or single-end configurations, again depending on
donors, who had a relapse rate of 75% over the same time period. the NGS platform used. Post-sequencing, the data is analyzed
Clearly, these results were of great interest to the field. In 2010, using bioinformatic pipelines. This step is very important and yet
Cooley et al. further showed a beneficial effect of HSC donors constitutes one of the main limitations in NGS. Bioinformatic
having centromeric B KIR ligand motifs in terms of relapse pre- pipelines allow alignment of the sequence reads with a reference
vention after unrelated HSCT in AML patients [7]. On the basis genome, and subsequent assignment of allelic variants. To date,
of these results, this group proposed an algorithm, which enables two NGS.KIR approaches have been developed: the exome cap-
the choice of HSC donors, based on their KIR gene content [7]. ture method [14] and the Nantes NGS.KIR method [15]. The
However, this algorithm has limitations. For example, it can- exome capture method developed by the Parham laboratory at
not reliably predict a better KIR+ NK cell alloreactivity in all the Stanford University School of Medicine is interesting and

Please cite this article in press as: Maniangou B, et al. Editorial: Next-generation sequencing technology a new tool for killer
cell immunoglobulin-like receptor allele typing in hematopoietic stem cell transplantation. Transfusion Clinique et Biologique (2017),
http://dx.doi.org/10.1016/j.tracli.2017.07.005
+Model
TRACLI-2958; No. of Pages 3 ARTICLE IN PRESS
B. Maniangou et al. / Transfusion Clinique et Biologique xxx (2017) xxx–xxx 3

resulted in the discovery of thirty-one new KIR alleles from only interactions between KIR3DL1 and HLA-A and HLA-B. J Immunol Baltim
15 individuals. However, this exome approach was not specific Md 2007;178:33–7.
[4] Battistini L, Borsellino G, Sawicki G, Poccia F, Salvetti M, Ristori G, et al.
for investigation of the KIR locus, but also for HLA sequences
Phenotypic and cytokine analysis of human peripheral blood gamma delta
and other genes. It is also time consuming and cannot easily be T cells expressing NK cell receptors. J Immunol 1997;159:3723–30.
implemented routinely. The process generates a large amount of [5] Triplett BM, Horwitz EM, Iyengar R, Turner V, Holladay MS, Gan K,
data that requires analysis with performant algorithms that take et al. Effects of activating NK cell receptor expression and NK cell
into account KIR, HLA and other polymorphic genes. In com- reconstitution on the outcomes of unrelated donor hematopoietic cell trans-
plantation for hematologic malignancies. Leukemia 2009;23:1278–87,
parison, the Nantes NGS.KIR approach, recently developed is
http://dx.doi.org/10.1038/leu.2009.21.
simple, fast, reliable, specific for KIR locus, validated in research [6] Ruggeri L, Capanni M, Urbani E, Perruccio K, Shlomchik WD,
and could be routinely employed for KIR allele typing before Tosti A, et al. Effectiveness of donor natural killer cell alloreactivity
HSCT with ease [15]. In the long term, the Nantes NGS.KIR in mismatched hematopoietic transplants. Science 2002;295:2097–100,
approach is planned to be used to evaluate the impact of KIR http://dx.doi.org/10.1126/science.1068440.
[7] Cooley S, Weisdorf DJ, Guethlein LA, Klein JP, Wang T, Le CT, et al. Donor
allele polymorphisms on HSCT outcome. It can also be used to
selection for natural killer cell receptor genes leads to superior survival
study KIR+ NK cell involvement in control of viral infections after unrelated transplantation for acute myelogenous leukemia. Blood
and reproduction. 2010;116:2411–9, http://dx.doi.org/10.1182/blood-2010-05-283051.
[8] Gagne K, Willem C, Legrand N, Djaoud Z, David G, Rettman P, et al. Both
Funding the nature of KIR3DL1 alleles and the KIR3DL1/S1 allele combination
affect the KIR3DL1 NK-cell repertoire in the French population. Eur J
Immunol 2013;43:1085–98, http://dx.doi.org/10.1002/eji.201243007.
This work was supported by EFS Pays de la Loire and [9] Bari R, Thapa R, Bao J, Li Y, Zheng J, Leung W. KIR2DL2/2DL3-E(35)
by grants from International Research Group on unrelated alleles are functionally stronger than -Q(35) alleles. Sci Rep 2016;6:23689,
Hematopoeitic stem cell Transplantation (IRGHET), Étab- http://dx.doi.org/10.1038/srep23689.
lissement Français du Sang (EFS 2016-09), l’Agence de la [10] Gardiner CM, Guethlein LA, Shilling HG, Pando M, Carr WH, Rajalingam
BioMedecine (ABM), La Ligue contre le Cancer, Association R, et al. Cell surface phenotypes defined by the DX9 antibody are
due to KIR3DL1 gene polymorphism. J Immunol 2001;166:2992–3001,
Recherche et Transfusion (ART), Leucémie Espoir Atlan- http://dx.doi.org/10.4049/jimmunol.166.5.2992.
tique Famille (LEAF) and Nantes Atlantique Greffe de Moelle [11] Boudreau JE, Giglio F, Gooley TA, Stevenson PA, Le Luduec J-B, Shaf-
Osseuse (NAGMO). BM is a PhD student supported by a fer BC, et al. KIR3DL1/HL A-B subtypes govern acute myelogenous
CIFRE/EFS Pays de La Loire/Nantes Université No. 481/2015 leukemia relapse after hematopoietic cell transplantation. J Clin Oncol
2017, http://dx.doi.org/10.1200/JCO.201670.7059.
grant.
[12] Alicata C, Pende D, Meazza R, Canevali P, Loiacono F, Bertaina A,
et al. Hematopoietic stem cell transplantation: improving alloreactive Bw4
Disclosure of interest donor selection by genotyping codon 86 of KIR3DL1/S1. Eur J Immunol
2016;46:1511–7, http://dx.doi.org/10.1002/eji.201546236.
The authors declare that they have no competing interest. [13] Mori A, Deola S, Xumerle L, Mijatovic V, Malerba G, Monsurrò V. Next
generation sequencing: new tools in immunology and hematology. Blood
Res 2013;48:242–9, http://dx.doi.org/10.5045/br.2013.48.4.242.
References [14] Kidd JM, Sharpton TJ, Bobo D, Norman PJ, Martin AR, Car-
penter ML, et al. Exome capture from saliva produces high
[1] Trowsdale J. Genetic and functional relationships between MHC and NK quality genomic and metagenomic data. BMC Genomics 2014;15:262,
receptor genes. Immunity 2001;15:363–74. http://dx.doi.org/10.1186/1471-2164-15-262.
[2] Wilson MJ, Torkar M, Haude A, Milne S, Jones T, Sheer D, et al. Plasticity [15] Maniangou B, Legrand N, Alizadeh M, Guyet U, Willem C, David
in the organization and sequences of human KIR/ILT gene families. Proc G, et al. Killer immunoglobulin-like receptor allele determination using
Natl Acad Sci USA 2000;97:4778–83. next-generation sequencing technology. Front Immunol 2017;8:547,
[3] Thananchai H, Gillespie G, Martin MP, Bashirova A, Yawata N, http://dx.doi.org/10.3389/fimmu.2017.00547.
Yawata M, et al. Cutting edge: allele-specific and peptide-dependent

Please cite this article in press as: Maniangou B, et al. Editorial: Next-generation sequencing technology a new tool for killer
cell immunoglobulin-like receptor allele typing in hematopoietic stem cell transplantation. Transfusion Clinique et Biologique (2017),
http://dx.doi.org/10.1016/j.tracli.2017.07.005