Genotyping killer immunoglobulin-like receptors using MALDI-TOF mass spectrometry for stem cell transplant donor selection

Abstract

The highly polymorphic receptors known as killer immunoglobulin-like receptors (KIR) are one of several families of receptors involved in regulating the activity of natural killer (NK) cells, a component of the innate immune system specialized to kill host cells that have become transformed or virally infected [1Lanier L.L. Curr Opin Immunol. 2003; 15: 308-314Crossref PubMed Scopus (293) Google Scholar]. KIR are diverse in terms of the number, type, and combination of genes present in individuals, and display extensive polymorphism at the nucleotide level. The ligands for NK receptors include the polymorphic human leukocyte antigen (HLA) class I cell surface molecules. Recent evidence suggests that interactions between an individual’s KIR molecular profile and HLA class I ligands may play an important role in stem cell transplant outcome; NK activation may be of benefit in hematopoietic transplants as donor NK cells in a KIR/HLA mismatched transplant have shown the ability to attack host leukemias and reduce graft versus host disease and graft rejection [2Ruggeri L. Capanni M. Urbani E. et al.Science. 2002; 295: 2097-2100Crossref PubMed Scopus (2598) Google Scholar, 3Gagne K. Brizard G. Gueglio B. et al.Hum Immunol. 2002; 63: 271-280Crossref PubMed Scopus (132) Google Scholar, 5Shilling H.G. McQueen K.L. Cheng N.W. Shizuru J.A. Negrin R.S. Parham P. Blood. 2003; 101: 3730-3740Crossref PubMed Scopus (219) Google Scholar. Thus assessment of KIR genotypes may become part of the donor selection process. As part of a larger study to analyze the role of donor and recipient KIR and HLA ligand profiles in stem cell transplantation, we are developing a novel, high-throughput single nucleotide polymorphism (SNP)-based KIR genotyping methodology. In this assay, the masses generated in a SNP-based primer extension reaction are analyzed on a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF). The method uses 384-well microarray chips, and is both highly accurate and rapid [4New MassARRAY Homogenous MassEXTEND (hME) Assay. Sequenom. http://www.sequenom.com.Google Scholar]. We present data using this novel typing method to demonstrate that the method is capable of accurately defining KIR genotypes in a panel of 100 donor recipient pairs from the NMDP. We are collaborating with the NMDP on an analysis of the KIR/HLA profiles of this panel and their relationship to hematopoietic transplant outcomes.