RESUMEN
Alloreactive NK cells lyse target cells lacking self-HLA-C or the HLA-B-Bw4 epitope. Prior to haploidentical stem cell transplants, donor alloreactivity toward the patient is evaluated by natural killer (NK) cloning followed by testing of the clones in the (51)Cr-release assay. As only a few percent of NK clones are alloreactive, a large number of NK clones must be established and evaluated. This approach is laborious and time consuming, with a complete evaluation taking up to 6 weeks. We developed a flow cytometry-based cytotoxicity assay utilizing CD107a expression on 12-day polyclonally expanded NK cells and showed that NK alloreactivity mediated by inhibitory and activating KIR can be detected by measuring CD107a expression following incubation with targets lacking the appropriate class I epitope. The percentage of alloreactive NK cells varied greatly between individuals and was easily estimated by the CD107a assay. For each epitope (C1, C2, Bw4), donors were found who did not have alloreactivity, although alloreactivity was predicted by the current rules thought to govern alloreactivity. The data emphasize the importance of demonstrating alloreactivity in a functional assay.
Asunto(s)
Pruebas Inmunológicas de Citotoxicidad/métodos , Citotoxicidad Inmunológica , Citometría de Flujo/métodos , Reacción Injerto-Huésped/inmunología , Prueba de Histocompatibilidad/métodos , Células Asesinas Naturales/inmunología , Proteínas de Ciclo Celular/metabolismo , Línea Celular Transformada , Células Cultivadas , Epítopos de Linfocito B/metabolismo , Genotipo , Antígenos HLA/metabolismo , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Interleucina-2/inmunología , Ligandos , Activación de Linfocitos , Proteína 1 de la Membrana Asociada a los Lisosomas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores KIR/genética , Receptores KIR/metabolismo , Factores de TiempoRESUMEN
Natural killer (NK)-cell alloreactivity can be exploited in haploidentical hematopoietic stem cell transplantation (HSCT). NK cells from donors whose HLA type includes Bw4, a public epitope present on a subset of HLA-B alleles, can be alloreactive toward recipients whose cells lack Bw4. Serologically detectable epitopes related to Bw4 also exist on a subset of HLA-A alleles, but the interaction of these alleles with KIR3DL1 is controversial. We therefore undertook a systematic analysis of the ability of most common HLA-B alleles and HLA-A alleles with Bw4 serologic reactivity to protect target cells from lysis by KIR3DL1-dependent NK cells. All Bw4(-) HLA-B alleles failed to protect target cells from lysis. All Bw4(+) HLA-B alleles with the exception of HLA-B*1301 and -B*1302 protected targets from lysis. HLA-A*2402 and HLA-A*3201 unequivocally protected target cells from lysis, whereas HLA-A*2501 and HLA-A*2301 provided only weak protection from lysis. KIR3DL1-dependent alloreactive NK clones were identified in donors with HLA-A*2402 but not in donors with HLA-B*1301 or -B*1302. These findings clarify the HLA types that donors and recipients need in haploidentical HSCT and other NK allotherapies in order to benefit from NK alloreactivity.