Immune reconstitution following haematopoietic stem cell transplantation

Stem cell transplantation is characterized by a subsequent period of immune deficiency, the duration and severity of which varies according to graft manipulation, choice of graft type (donor and source), development of graft-versus-host disease and level of residual thymic activity. This results in significant morbidity and mortality. Supportive therapies have advanced considerably leading to improved outcomes of infective episodes. However, disease relapse remains a considerable obstacle to improvement in overall outcomes. An increased understanding of the role of the immune system in preventing relapse, particularly in the allogeneic setting, has highlighted the importance of the reconstitution process. Successful application of some of the newer therapeutic strategies, such as tumour vaccination approaches, may depend critically on reconstitution of functional immune reactivity. In addition, the development of assays to measure recovery of antigen-specific immunity may allow further refinement of targeted therapies. This article reviews current knowledge of the reconstitution process. The escalation in doses of cytoreductive therapy that have characterized the development of haematopoietic stem cell transplantation (HSCT) protocols results in profound and long-lasting perturbations in immune response. Whilst early recovery of innate immunity [myeloid series and natural killer (NK) cells] results in reconstitution of protective immunity against many bacterial pathogens, both the absolute levels and function of T and B lymphocytes remain abnormal for many months, resulting in prolonged susceptibility to viral pathogens. More recently, the recognition that dose escalation alone does not eradicate the malignancy in many cases and that an associated immune-mediated graft-versus-malignancy effect may be equally important has led to the development of reduced intensity and non-myeloablative conditioning regimens, which aim to reduce the toxicity of the procedure by substituting more immunosuppressive agents in place of the myelosuppressive components of the original regimens. The influence of these developments on the tempo and characteristics of immune repertoire reconstitution are only just becoming evident. Incorporation of T-cell depletion, choice of graft type (both donor and source), development of graft-versus-host disease (GvHD) and level of residual thymic activity can all influence aspects of the reconstitution process. In addition, recent developments in the tools that are available for monitoring the reconstitution of these processes are providing new insights, particularly into the recovery of specific T-cell subsets. There are significant differences between the patterns of immune reconstitution following autologous and allogeneic transplants, relating both to the genetic and immunological disparity between donor and recipient in the latter, and to the influence of the immunosuppressive drugs employed to address these differences. These agents interfere both with quantitative and qualitative aspects of immune recovery. This review focuses on recent advances in the understanding of immune reconstitution in the allogeneic setting. Reference is made to autologous SCT only where it aids in the illustration of general principles, and for more comprehensive reviews the reader is referred elsewhere (Guillaume et al, 1999; Porrata et al, 2001). The reconstituting T-cell repertoire appears to be derived from cells from a number of sources. Contributions are made in varying proportions from expansions of infused mature lymphocytes, of residual T lymphocytes that have survived the conditioning therapy, and from the development of thymic-independent and thymic-dependent naïve T cells. Extrathymic maturation of naïve T cells has been primarily described in sites such as the gut. It commonly results in development of non-classical T cell populations [e.g. T-cell receptor (TCR)γδ, CD4−CD8−TCRαβ] that are not generally considered to re-circulate into peripheral tissues, and appears to play a minimal role in post-transplant reconstitution. Advances in the understanding of the contributions of the thymic-dependent and thymic-independent pathways have been aided by developments in multiparameter flow cytometry, T-cell rearrangement excision circlet (TREC) assays, and TCR spectratype (immunoscope) analysis (Fig 1). More detailed information on the reconstitution of antigen-specific responses has been derived from intracellular cytokine flow cytometry, cytokine secretion assays [enzyme-linked immunospot (ELISPOT), matrix affinity technology] and class I human leucocyte antigen (HLA)-peptide tetramer labelling. Whilst each of these assays has its own particular limitations (which are important to appreciate), their combination can be particularly informative. Methods of monitoring T cell reconstitution. Deletion of the TCRD locus from within the TCRA locus is a common requirement for productive rearrangement of the α-chain (A). The resultant signal joint (sj) T-cell rearrangement excision circlet (TREC) can be quantified by real-time polymerase chain reaction (PCR) allowing assessment of thymic output. Because episomal DNA does not replicate but remains stable over time, the TRECs are diluted out by cell divisions as the T cells age (B). PCR across the rearranged β-chain CDR3 region forms the basis of TCRBV spectratyping (C). The resultant PCR product size distribution profile approximates to Gaussian in neonates. Flow cytometry can identify surface or intracellular molecules, and combination with class I human leucocyte antigen (HLA)-peptide multimers allows detection of peptide-specific T-cell receptor (TCR) (D). Capture and localization of secreted cytokines to the surface of the secreting cell by bi-specific antibodies forms the basis of affinity matrix technology [e.g. interferon (IFN)-γ capture]. The most commonly used tool for assessment of T-cell repopulation has been flow cytometry, and this allows differentiation of the thymic-dependent component by surface immunophenotype. Naïve CD4+ T cells can be identified by expression of CD45RA antigen (often in combination with CD62L or CD11a), whereas identification of naïve CD8+ T cells requires multiple markers. More refined technologies have identified ever increasing complexity in terms of heterogeneity in previously described lymphocyte subsets, both in terms of surface immunophenotype and functional characteristics. The current of available and cytometry allow with technologies increasing the to assessment flow to to or more et al, These which have on CD8+ subsets, that assessment of to surface CD62L and with to functional assays, may be for the analysis of naïve CD8+ T-cell are to complexity in naïve CD4+ in by the identification of recent thymic of these technologies functional and of according to antigen and intracellular (Fig for components and and have all technologies et al, assays that on of or cytokine expression [e.g. et al, and the of class I multimers or that to antigen-specific T cells et al, have the for the of reconstitution of antigen-specific immunity (Fig The of T cells in the TCR of and with only a a TCR of and of these are by and which are rearranged to the of the TCR the of These are for the of the of both the TCRA and T-cell maturation the of DNA are as the are to a functional TCR in DNA et al, The of the DNA are to DNA Because of the of and and the TRECs can be used to overall thymic However, a common requirement for the productive rearrangement of the TCRA locus is of the TCRD locus that it (Fig This rearrangement results in of the signal the locus and the and the resultant a signal joint (sj) This is in of T cells and the resulting can be used to and thymic of T cells. Because episomal DNA does not replicate but remains stable over time, the TRECs are diluted out by cell divisions as the T cells age (Fig The the can be polymerase chain reaction (PCR) assays with to the and by real-time PCR allows assessment of thymic output. levels are as a TRECs T cells or and be by the of T-cell and et al, 2001). The rearrangement the basis for that can be used to T-cell repertoire to of the TCR has on the β-chain of its and the number of region with the TCRA The region is the region of the TCR by the rearrangement of TCR β-chain and The of and region with the of activity and by the (Fig of TCR across the CDR3 allows of the of within an TCRBV et al, et al, et al, T lymphocyte maturation for productive of a requirement for rearranged to with and receptor on the cell only T cells with further to in the peripheral resulting in the CDR3 of T cells of the functional varying by of in spectratype approximates to a Gaussian distribution in et al, et al, with resulting in expansions which in the and of this with increasing TCRBV spectratype analysis has been used to overall TCR repertoire et al, relating to the number of CDR3 and their of These of and a number of have been derived in to These from the the number of to the and to has been as to the most of this et al, et al, et al, et al, can be to B cells. The of and is by and across the CDR3 in an region with of TCR spectratype characterized by of et al, CDR3 of the CDR3 by polymerase chain reaction (PCR) results in size heterogeneity to by analysis of the TCRBV CDR3 have been to over prolonged following transplantation In to antigen-specific T the cells of the innate immune system the to and the and of the immune cells are innate immune lymphocytes that cells an of and and these can be identified multiparameter flow cytometry with antibodies et al, early of or residual mature lymphocytes results in of the that primarily as a of CD4+ T-cell recovery. Whilst are to in T cell the to which expansions and the of the remain has been in transplantation in which T cells into into This appears to be by and levels have been to following transplantation in and with CD4+ T cell levels et al, 1999; et al, et al, 2001). may be in these processes et al, The immunophenotype of the CD8+ T cell expansions that can to early reconstitution and the of T cells that has been to of spectratype this period this et al, et al, et al, 2001). There are a number of that these These and some of the multiple of of peripheral lymphocytes have been in with processes or and has identified T-cell to CDR3 with a number of the that in (Fig et al, These that the post-transplant T-cell expansions are However, T cells in the and can be to from expansions that TCRBV spectratype the patterns in peripheral may not in the the of on the of reconstitution et al, et al, of antigen-specific responses following stem cell transplantation I human leucocyte antigen (HLA)-peptide tetramer identify only CD8+ T cells (A). TCRBV spectratype following transplantation reduced spectratype with to more over subsequent (B). by spectratype analysis can identify the region and CDR3 of the T cells. of these with the expansions in the spectratype from T that T cell expansions can to the of immune reconstitution (B). allows functional identification of both CD4+ and CD8+ T cells according to cytokine secretion (C). for surface antigen expression allows of these populations (D). of in to an increased susceptibility to cell are a of the lymphocytes that are in the early This which is more in the and in the of to the of T-cell reconstitution et al, the of early expansions and with subsequent CD4+ T cell reconstitution of the latter, with of more spectratype may depend more on thymic function CD8+ T-cell reconstitution. reconstitution. The thymic that increasing age has been for many to be an important in the in immune reconstitution that in with The development of has in an increased understanding of the role of the in in a in thymic the residual are to and to This more following transplantation with its prolonged CD4+ of naïve T cells and TCR repertoire as by of more spectratype generally does in and has been with However, the of this can be particularly in with the of TCRBV repertoire the in naïve T cells and of lymphocytes not the or and this is particularly for CD4+ T cells. are characterized by of thymic and more This results in more recovery of CD4+ T which a naïve and a recovery of levels and TCRBV spectratype et al, 2001). This in has been to with more recovery of particular of the TCR repertoire in responses post-transplant has significant differences between recipient and donor in terms of β-chain region and TCRBV spectratype the of the CDR3 of the et al, 2001). These the of most from the naïve repertoire by thymic-dependent This may have significant for to protective immunity against and responses by vaccination in the post-transplant which be to with the of functional in immune where of donor T-cell responses is important for responses in the early post-transplant early vaccination may be However, for that on the of immune responses and thymic-dependent immunological monitoring may for of vaccination of graft-versus-host disease and The development of the of thymic-independent and tempo of thymic-dependent T-cell reconstitution. CD4+ and CD8+ T cell have been to in sites it appears that these can to overall repertoire reconstitution. CD4+ to the immune to such a that it to of the CD4+ the development of in et al, real-time quantitative PCR that the of this with and subsequent of However, such appears in TCRBV spectratype have more such responses to be et al, et al, in some to from the et al, In of the responses are in some et al, 2001). of T lymphocytes class and following transplantation that and as early as graft et al, expansions of and and with from the of these to that be by TCRBV spectratype their to the overall CD8+ T-cell repertoire that of the previously described CD4+ T cell to the CD4+ T-cell The expansions associated with the reconstitution of T-cell repertoire in a et al, allowing of antigen-specific T cells have that with are associated with expansions of T cells which are but significant levels et al, 1999; et al, expansions can be following SCT (Fig and following therapy for or et al, et al, et al, et al, This an important to the of spectratype in the of reconstitution of immune of TCRBV following reduced T-cell repertoire with a subsequent to a more This has been with the abnormal immune function that this However, of spectratype by expansions results in a immune response. as immune as by the tempo of of TCRBV by both and have been to thymic-dependent et al, et al, et al, et al, et al, results in levels of naïve and T cells. This to the levels of TRECs described in donor with may this effect in by a effect on However, it is important to that by lymphocyte both and can in an in naïve T-cell and levels these are with T cell as a Whilst this effect can be by absolute cell it the to be of the limitations of new monitoring in the of in where multiple can influence the of of lymphocyte et al, The influence of the development of on immune reconstitution following allogeneic transplantation has been to from the of associated or from the influence of characteristics or aspects of conditioning protocols that are more commonly associated with its of with disease reduced intensity transplantation that may be associated with more immune reconstitution and T-cell et al, 2001). However, in this series more common in the and levels in this in with the of more thymic In addition, the more donor lymphocyte and An influence of the of of more lymphocytes not be The that may be associated with and or and that this influence many of the differences remains but and further in this are have that of may be by transplantation with graft and et al, T lymphocytes to and donor in the and is to with in some of these et al, et al, 1999; et al, 2001). may against the malignancy such that the relapse may be in T a that not be to be in the in is an for subsequent disease et al, of can this leading to to donor the immunological of have that to donor is associated with of T-cell repertoire and more Gaussian spectratype et al, et al, et al, The this remain of T cell with that activity be to in increased of TCRBV in a to that described for the that of the responses that are of a of the with be to in expansions and to further to this effect these responses are by T cells as generally The of these in some may be by the level of in which these responses are an is not only on the absolute but on the level of expression of the TCRBV within which the post-transplant the lymphocyte has to some to may remain by the tools generally employed for these In addition, the of tools for repertoire may the the be following application of et al, However, CD4+ associated with increased of TRECs in T that be a of thymic-dependent reconstitution following et al, the complexity of the multiple that repertoire expansions and naïve T cell it is not that the overall of on repertoire are and that have been to be outcomes et al, of antigen-specific T cell of immune responses allogeneic transplantation appears to be protective against the development of viral In the of of activity against with against et al, peptide-specific class I has these et al, et al, et al, 2001). It is early to the influence of conditioning regimens, graft and donor In the recovery in the of donor in the of donor transplants, but an influence of T cell by not be et al, 2001). increased of but by and to of all CD8+ T cells in both donor and recipient for to to levels associated with from derived in a subsequent et al, 2001). a recovery in a graft from a The number of T cells within the graft may with the number of et al, 2001). Incorporation of into a reduced intensity transplantation to recovery in SCT et al, The contributions of T-cell and of antigen cells to this effect remains in of responses by has been with this et al, of these responses is to of B cells. in reconstitution of responses are most commonly with protocols levels of T-cell and post-transplant immune employed for Whilst of the on responses has on CD8+ T the importance of antigen-specific CD4+ T cells is The therapy of and a requirement for reconstitution of CD4+ responses for of CD8+ responses et al, et al, and affinity matrix technologies allow further refinement of CD4+ of has been to be associated with reconstitution of CD4+ responses with these et al, of these with application to (both viral and may the for both monitoring and following allogeneic application to and to further into the of disease relapse and allow development of targeted of The importance of cells in the development of immune in has over recent role in immune repertoire recovery following transplantation is only to be The of has been to in following in et al, have been to be for the of leading to the that or of be in the of following allogeneic et al, in populations are to influence the of thymic-independent T cell expansions and TCRBV spectratype levels by a and in a to that identified in neonates. recovery in levels by levels and levels the to some on conditioning and graft characteristics. repopulation can be in the early period with B cell CDR3 by and from the distribution of with within et al, et al, The remain for considerably with of prolonged expansions but overall reconstitution of more becoming et al, These are in with the not of or on The period of immune SCT is to be by a number of the of and The more recovery of repertoire in B cells with the recovery of TCR repertoire and may the of B cells with the in thymic activity. levels to specific have been to with particularly within the et al, immunity appears to be This donor leading to to to transplantation against both agents and such as B cell of results in B the remains and of the drugs used to of innate immunity following transplantation is more that of the immune of cell reconstitution following have been with of T and B cells in the of In analysis has been to quantitative reconstitution of lymphocytes, with further of and subsets. The may in or and have early or in cell in the early advances in the understanding of their receptor and functional importance in activity have in an increased in cell reconstitution. The cell are by of the killer and the et al, or to their These of and in the of is the class I in the of functional activity. with with derived from the of class I It appears that cell receptor specific for autologous class resulting in the of and are in of resulting in a class I that is by the of a as is for T cells. The cell repertoire of an is by the which in number and as as et al, and are in the human in the leucocyte receptor on human and and for allogeneic transplantation focuses on the of and all be of cell following allogeneic transplantation that the the is the of repertoire and that the has most only a effect et al, be into according to the of receptor expression the In the the expression profile that of the donor and stable recovery of cell the of cells The of these cells in with with a of cells donor levels within In the the expression profile but the tempo of of cell This only donor the that of and This with a in recovery of expression cells following autologous led to the that genetic disparity may the recovery of an influence of differences in conditioning therapy not be with outcomes may The patterns of reconstitution that from the donor and a characterized by or or relapse of with in the These further and The functional of the recipient appears to be the influence on thymic-dependent the of stem cell peripheral stem cells and stem cells are all of the reconstitution of naïve T cells and T cell et al, et al, et al, and associated T cell depletion, has been to be associated with increased levels following autologous transplantation and this has been as thymic et al, The of this is but be to to by the levels of which are increased following transplantation and with CD4+ T cell has activity in peripheral T cell populations and et al, 2001). However, to be in on levels et al, 2001). 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In of have been associated with a of increased and transplantation with particularly In this the antibodies against B that both on donor and recipient cells et al, 2001). and of B by the type of CD4+ cells and be to these differences. has been to by B lymphocytes et al, T-cell has been to be the most for preventing following allogeneic However, of T cells immune resulting in increased of and These can all on patterns of reconstitution according to are T lymphocyte CD4+ T cells remain the that has most been to be following of the has been that immune reconstitution following reduced intensity et al, et al, In general the patterns are important differences may the current of conditioning which considerably in and immunosuppressive are to in that may be more on the drugs early that reduced intensity transplantation may be associated with more of the T-cell repertoire and a number of CD4+ and CD8+ cells et al, However, early T-cell recovery to from T the of which most by the T-cell but not by or are An review of current and of immune reconstitution is the of this these can be into general immune repertoire recovery and antigen-specific is the most cytokine of in but the T-cell dose for of in some cases et al, The subsequent thymic toxicity in T transplants, but in T In immune reconstitution improved et al, 2001). In this the and peripheral T cells in and and and et al, 2001). has some as it appears to thymic from et al, lymphocyte can immune the development and of remain et al, to the of associated with the of of T cells. can be into that aim to out the and that for more subsets. CD8+ appears to be of responses with a reduced of et al, this a to immunity remains to be but is some from that CD4+ T cells not of increased of T cells et al, of T cells identified by of or of by following lymphocyte is an et al, et al, 1999; et al, et al, In this it may be to that to with a of early following transplantation et al, immunity can be by of cells that have been or to have or specific activity against an or of choice et al, et al, et al, or by vaccination or The period of immune that allogeneic SCT to in significant morbidity and mortality. have the tools available to more the of immune and to the of to immune recovery. therapies and some However, it is important to that of the immune is and in to the of development of to immunity that to allow for this may of this More specific therapies are for some of the viral that post-transplant and therapies have the of knowledge immune reconstitution may to the immunological of allogeneic transplantation by the and