Opportunities and limitations of natural killer cells as adoptive therapy for malignant disease

Abstract

Although natural killer (NK) cells can be readily generated for adoptive therapy with current techniques, their optimal application to treat malignant diseases requires an appreciation of the dynamic balance between signals that either synergize with or antagonize each other. Individuals display wide differences in NK function that determine their therapeutic efficacy. The ability of NK cells to kill target cells or produce cytokines depends on the balance between signals from activating and inhibitory cell-surface receptors. The selection of NK cells with a predominant activating profile is critical for delivering successful anti-tumor activity. This can be achieved through selection of killer immunoglobulin-like receptor-mismatched NK donors and by use of blocking molecules against inhibitory pathways. Optimum NK cytotoxicity may require licensing or priming with tumor cells. Recent discoveries in the molecular and cellular biology of NK cells inform in the design of new strategies, including adjuvant therapies, to maximize the cytotoxic potential of NK cells for adoptive transfer to treat human malignancies.

Keywords: C-type lectin; graft-versus-leukemia; immunotherapy; killer immunoglobulin-like receptors; natural cytotoxicity receptors; natural killer.

Figure 1. NK cell activation by a kinetic segregation model

Activating NK receptors generally use adapter proteins such as Fcεγ, and DAP12, which contain ITAM consensus sequences. These trigger a rise in intracellular Ca²⁺, and degranulation following phosphorylation by tyrosine kinases such as Syk and ZAP70. Similarly the majority of inhibitory NK cell receptors contain an ITIM consensus motif, phosphorylation of which results in recruitment of tyrosine specific phosphatases such as SHP-1, SH2-containing protein tyrosine phosphatise-1 (SHP-1) and SH2-containing inositol pholyphosphate 5-phosphatase (SHIP). These act by dephosphorylating ITAM motifs and SHIP degrades phosphatidylionsitol-3,4,5-trisphospate, leading to inhibition of sustained calcium signalling. The kinetic segregation model of NK cell activation (19) proposes that the normal balance between phosphorylation and dephosphorylation of these receptors is disturbed by physical extrusion of large phosphatases such as CD45 and CD148 from areas of close contact between the NK cell and its target. This leads to phosphorylation by small kinases of both the activating and inhibitory receptors that diffuse into, and are held at the immune synapse, and allows NK cell activation to be dependent on the balance between the number of activating and inhibitory ligands on the target cell. Several important receptor subtypes are not depicted in this diagram (e.g. the activating receptor CD16, which uses the ITAM containing adapter protein Fcεγ, see Table 1). In addition non-ITAM mediated activation also occurs, particularly by the NKG2D-DAP10 complex (102).

Figure 2. KIR genotype and receptor function

Simplified schema of the KIR gene locus (modified from the UCSC genome browser http://genome.ucsc.edu/, hg19 (2009)). At least 37 haplotypes have been identified thus far; however, the KIR genotype can broadly be divided into two groups, depending on whether the person is homozygous for genotype A (AA), in which case they only express one activating KIR (KIR2DS4) or whether they have at least one B haplotype (Bx). The B haplotype is defined by the presence of a variable number of additional activating and inhibitory KIRs (KIR2DL5, KIR2DS1, KIR2DS2, KIR2DS3, KIR2DS5 and KIR3DS1).

Figure 3. KIR ligand mismatch

KIR Ligand mismatch is largely dictated by the ligands of the three inhibitory KIR receptors KIR2DL1, KIR3DL1 and KIR2DL2, which are highly prevalent (97%, 90% and 95% respectively). This model predicts that when when the transplant recipient is missing at least one of the three major classes of HLA ligands for inhibitory KIR, the donor NK cells will recognise the missing ligand in the host, resulting in an augmented graft versus leukaemia effect. Theoretically this can occur in the opposite direction (where the ligand is missing from the donor) but there is little evidence that this leads to worse rates of graft rejection.