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410 Part V: Therapeutic Principles Chapter 26: Immune Cell Therapy 411

Transferred T cells must persist as functional memory T cells and T-CELL THERAPY OF EPSTEIN-BARR
migrate to sites of virus replication to be effective. Therefore, it was VIRUS INFECTION
essential to verify the presence of the transferred T cells in the blood
based on functional or structural properties. In the initial studies of EBV is a ubiquitous γ-herpesvirus (Chap. 82) that persists in immuno-
adoptive therapy with CD8+ CMV-specific T-cell clones, assays of cyto- competent hosts lifelong without causing disease. Some latently infected
lytic activity provided an indirect semiquantitative functional measure B cells express only the EBNA-1 protein, which has glycine-alanine
for the presence of the transferred immune effectors. New approaches repeats that inhibit its translation and processing for presentation to
27
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that use flow cytometric analysis of blood samples have been devel- CD8+ T cells. Infected B cells may activate the latency III program
oped, such as staining with HLA multimers or intracellular staining, to of viral genes that includes EBNA-1, EBNA-2, EBNA-3A, EBNA-3B,
detect cytokines produced after antigen stimulation. These techniques EBNA-3C, LMP-1, LMP-2A, and LMP-2B, and induces cell prolifera-
56
are being employed to enumerate and analyze the function of cells on a tion. In normal hosts, both the CD8+ CTL and CD4+ Th cell response
single-cell level. 11,34 The unique DNA sequences of the rearranged TCR to EBV infection is mainly directed against lytic viral proteins and the
Vα or Vβ genes have also been used to evaluate survival of transferred T EBNA-3A, -3B, and -3C latency proteins. 57,58 The CD4+ Th cell response
27
cells in the first trial of CMV-specific T-cell therapy. Quantitative real- may also contribute to eliminating class II MHC+ EBV-infected cells in
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time polymerase chain reaction with TCR-specific primers that flank vivo and EBV-specific CD8+ and CD4+ T cells cooperate to prevent
the unique complementarity determining region 3 (CDR3) sequence the outgrowth of EBV+ B cells in immunocompetent hosts. 59,60 Thus,
can provide precise quantitation of transferred T cells in blood samples. tumors comprised of proliferating EBV+ B cells can arise in individuals
Advances in high-throughput sequencing and computational analysis with a T-cell deficiency, such as solid-organ or HSCT patients receiv-
of the TCR sequences have now enabled much more sensitive analy- ing intense immunosuppression, especially if T-cell depletion is used as
sis and sequencing of the CDR3 and detection of unique sequences in part of the conditioning regimen. 61,62 Historically, patients with EBV-
transferred T cells. 36,41,42 induced lymphoproliferative disease (EBV-LPD) had a grave prognosis,
responding poorly to antiviral drugs or chemotherapy, although early
detection of EBV reactivation and treatment with mAbs specific for
Selection of Defined Subsets of Cytomegalovirus-Specific CD20 have improved outcomes. Ultimately, restoration of EBV-specific
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T cells for Adoptive Therapy T cells is necessary for control of the virus.
The pool of memory T cells contains both CD45RO+CD62L+ cen-
tral memory (T ), and CD62L– effector memory (T ) subsets that
CM
EM
differ in phenotype, function, and migration. 43–45 Studies in animal Techniques for Isolation and Adoptive Transfer
models have revealed profound differences in the ability of adop- of Epstein-Barr Virus–Specific T Cells
tively transferred T cells from distinct subsets to persist in vivo and The efficacy of T-cell therapy for EBV-LPD was first demonstrated in a
revert to the memory pool. The transfer of effector cells derived from study in which a low dose of unselected donor lymphocytes was admin-
CD8+CD62L+ T or a rare subset of CD62L+ cells termed memory istered to five patients with EBV-LPD after T-cell–depleted allogeneic
CM
64
stem cells (T SCM ) that share cell-surface markers of both naïve T cell HSCT. Complete resolution of EBV-LPD was achieved in all patients,
(T ) and T cells, displayed superior survival in vivo, and/or medi- but this was complicated by GVHD and two patients developed a
CM
N
ated superior antitumor activity compared with more differentiated fatal respiratory failure, demonstrating the importance of select-
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T . 46–49 Studies in mice provide increasing evidence for a progressive ing EBV-specific T cells for therapy. The adoptive transfer of donor
EM
differentiation model of T-cell subsets. Fate mapping studies and sin- EBV-specific T-cell lines derived by in vitro culture with EBV-trans-
gle-cell transfer experiments show that naïve T cells differentiate into formed lymphoblastoid cell lines (LCLs) was effective in 2 of 3 HSCT
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memory and effector subsets (Fig. 26–2), and demonstrated that the recipients with established EBV-LPD without causing GVHD. One
stem cell–like properties of self-renewal and differentiation are pres- patient had progressive LPD with a mutation in the EBNA-3B gene
ent in CD62L+ T CM cells. 50–52 It is uncertain whether the intermediate that eliminated the region encoding the epitopes targeted by the CTL
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phenotype T SCM has self-renewal capability or represents a cell that is of line. To diminish the probability of escape variants, subsequent studies
sufficient frequency to be reliably isolated for immunotherapy. Never- administered donor EBV-specific T-cell lines prophylactically to a cohort
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theless, these findings suggest that selecting less differentiated memory of patients at risk for EBV-LPD after T-cell–depleted allogeneic HSCT.
subsets as a starting population for clinical adoptive T-cell therapy can No GVHD was observed, and there were no cases of LPD, although this
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allow very low numbers of T cells to be effective, and this concept has was expected to occur in 14 percent of the patients. A 9-year followup
53
been validated in animal models and patients with CMV infection. report summarized the results in a total of 114 HSCT recipients, 101
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Given the considerable potential of this approach, methods are now of which safely received prophylactic infusions of EBV-specific CTLs.
being developed for clinically applicable serial selection strategies for None of the 101 patients progressed to EBV-LPD and 11 of 13 patients
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human T subsets. 54 with LPD achieved sustained complete remissions. Thus, transfer of
CM
Naïve T cell Memory T cell Effector T cell Figure 26–2. Linear differenti-
ation of T-cell subsets. The phe-
T N T SCM T CM T EM T E notype of naïve, memory, and
effector subsets is shown and the
linear pathway of differentiation
from a naïve T cell is based on
+
+
+
+
+
CD45RA CD62L + CD45RA CD62L + CD45RO CD62L + CD45RO CD62L – CD45RO CD62L – recent data from fate mapping
+
+
+
CD28 CD95 – CD28 CD95 + CD28 CD95 + CD28 +/– CD95 + CD28 +/– CD95 + studies in murine models. 50,51
hi
CD122 + perforin granzyme B hi
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