Granulocyte Colony-Stimulating Factor Effectively Mobilizes TCR γδ and NK Cells Providing an Allograft Potentially Enhanced for the Graft-Versus-Leukemia Effect for Allogeneic Stem Cell Transplantation

Abstract

Allogeneic hematopoietic stem cell transplantation (HSCT) is a potential cure for patients with hematological malignancies but substantial risks of recurrence of the malignant disease remain. TCR γδ and NK cells are perceived as potent innate effector cells in HSCT and have been associated with post-transplant protection from relapse in clinical studies. Immunocompetent cells from the donor are crucial for patient outcomes and peripheral blood stem cells (PBSC) are being increasingly applied as graft source. G-CSF is the preferential mobilizing agent in healthy donors for PBSC grafts, yet effects of G-CSF on TCR γδ and NK cells are scarcely uncovered and could influence the graft composition and potency of these cells. Therefore, we analyzed T and NK cell subsets and activation markers in peripheral blood samples of 49 donors before and after G-CSF mobilization and—for a subset of donors—also in the corresponding graft samples using multicolor flowcytometry with staining for CD3, CD4, CD8, TCRαβ, TCRγδ, Vδ1, Vδ2, HLA-DR, CD45RA, CD197, CD45RO, HLA-DR, CD16, CD56, and CD314. We found that TCR γδ cells were mobilized and harvested with an efficiency corresponding that of TCR αβ cells. For TCR γδ as well as for TCR αβ cells, G-CSF preferentially mobilized naïve and terminally differentiated effector (TEMRA) cells over memory cells. In the TCR γδ cell compartment, G-CSF preferentially mobilized cells of the nonVδ2 types and increased the fraction of HLA-DR positive TCR γδ cells. For NK cells, mobilization by G-CSF was increased compared to that of T cells, yet NK cells appeared to be less efficiently harvested than T cells. In the NK cell compartment, G-CSF-stimulation preserved the proportion of CD56dim NK effector cells which have been associated with relapse protection. The expression of the activating receptor NKG2D implied in anti-leukemic responses, was significantly increased in both CD56dim and CD56bright NK cells after G-CSF stimulation. These results indicate differentiated mobilization and altering properties of G-CSF which could improve the effects of donor TCR γδ and NK cells in the processes of graft-versus-leukemia for relapse prevention after HSCT.

Perturbed NK-cell homeostasis associated with disease severity in chronic neutropenia

“…The outstanding question is at what point the NK cell maturational trajectory is interrupted in these patients. NK cell progenitors are functionally impaired in G-CSF mobilized stem cell products, albeit peripheral CD56 bright/dim ratios appear to be unaffected 55,56 . We found normal absolute numbers of CD56 bright with non-aberrant phenotypes, suggesting that the homeostatic defect is downstream of progenitor stages, primarily affecting the CD56 dim NK cell compartment.…”

Donor lymphocyte infusion for prophylaxis and treatment of relapse in pediatric hematologic malignancies after allogeneic hematopoietic stem cell transplant

“…26 Furthermore, G-CSF may mobilize a high number of HLA-DR + TCR gamma-delta cells, increase the CD56dim subpopulation of NK cells, and upregulate the expression of the activating receptor NKG2D, all of which are associated with antitumor effects and peripheral tolerance. 11,12 A retrospective, single-center study reported that G-CSFprimed DLI improved the rate of conversion into full donor chimerism and reduced CIR without increasing the incidence of GVHD. 9 By contrast, other retrospective studies 27,28 and a metaanalysis involving 585 patients 29 have reported no significant difference between G-CSF-primed DLI and SS-DLI in terms of their effectiveness in preventing disease progression or GVHD development.…”

“…With the accumulation of clinical DLI experience and the changes in the relevant guidelines, 2009 onward, the policy for serial DLI shifted toward dose escalation. Considering the potential advantages of G-CSF-primed DLI indicated in several immunologic studies, 9,11,12 since 2017, we have been routinely cryopreserving an excessive amount of peripheral blood stem cell (PBSC) product obtained on day 0 of HSCT for future use in G-CSF-primed DLI. In studies focusing on SS-DLI, starting DLI doses ranging from 1 × 10 5 to 1 × 10 6 CD3 + cells/kg were used for prophylactic and preemptive purposes after matched sibling or unrelated donor HSCT and a higher initial dose (1 × 10 7 CD3 + cells/kg) was used for frank relapses.…”

“…10 G-CSF stimulates various immune cell populations (eg, plasmacytoid T cell–derived CD11c − dendritic cells, HLA-DR + TCR gamma-delta cells, and CD56dim subpopulation of NK cells). 11,12 Whether the polarization of immune cell subpopulations and nonlymphocytic components increases the GVL effect, reduces the GVHD risk, and provides survival advantages in a clinical setting is a highly debated topic.…”

Gamma delta T-cell reconstitution after allogeneic HCT: A platform for cell therapy

“…Conversely, a more detailed analysis by Minculescu et al. showed that G-CSF preferentially mobilized naïve and terminally differentiated effector (TEMRA) γδ T cells over memory cells with a preferential increase in the non-Vδ2 subset while also increasing the proportion of HLA-DR expressing γδ T cells ( 67 ). Regarding the impact on TCR repertoire, it has been shown that G-CSF was associated with TCR repertoire disturbances in the form of alteration of distribution and clonality of some TRG and TRD subfamilies, suggesting a potential immune modulatory effect ( 70 ).…”