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macrophages. Use of
ex vivo
donor T-cell manufacturing offers a strategy to
resolve these potential limitations because purified immune cell popula-
tions can be exposed to high concentrations of cytokines and other factors
(such as rapamycin) in an extracorporeal manner that dissociates biologic
agent effect from toxicity.
In 1999, we initiated the first clinical trial of allogeneic CD4
+
Th2 cell ther-
apy at the Experimental Transplantation and Immunology Branch, Center
for Cancer Research, National Institutes of Health
[97]
. This protocol rep-
resented a clinical translation of our murine experiments using the tech-
nique of Th2 cell allograft augmentation: that is, patients received the full
complement of donor T cells contained within the G-CSF mobilized, HLA-
matched sibling, hematopoietic cell product and also received
ex vivo
-gen-
erated donor Th2 cells. Transplantation was performed after intensive host
immune ablation using fludarabine-containing outpatient induction and
inpatient preparative chemotherapy. In this first-generation clinical trial,
Th2 cells were manufactured by the following method: (a) CD4 cells were
isolated by steady-state apheresis and subsequent elutriation and negative
selection; (b) CD4 cells were costimulated at day 0 of culture and also for a
second time at day 12 of culture; and (c) T cells were expanded in the pres-
ence of IL-4 and IL-2 (no rapamycin). The resultant T-cell product was par-
tially shifted toward a Th2 phenotype relative to culture input donor CD4
+
T cells; however, such polarization was incomplete relative to the extreme
polarity that can be achieved in murine models. And, because of the rela-
tively prolonged T-cell culture duration that did not include rapamycin, the
differentiation status of the T-cell products was most consistent with rela-
tively late-stage T
EM
cells (including the characteristic of having high levels
of cytokine secretion at the time of infusion).
236
Donor Th2 cells were transfused in a phase I dose-escalation manner at
5, 25, or 125 × 10
6
Th2 cells per kilogram of recipient body weight, with an
expanded cohort of patients treated at the intermediate dose in a phase II
manner (
n
= 3,
n
= 19 and
n
= 6 subjects were transplanted per successive
cohort, respectively). There were no clinical toxicities attributable to Th2
cell infusion. Post-transplant immune monitoring determined that Th2-cell
recipients expressed a mixed pattern of both Th1 and Th2 cytokine produc-
tion post-transplant. However, relative to a protocol control cohort that did
not receive Th2 cells, monocyte inflammatory cytokines IL-1α and TNF-α
were not reduced in Th2-cell recipients; in addition, acute GVHD was not sig-
nificantly reduced in Th2-cell recipients. In sum, these results demonstrated
the feasibility, safety and apparent biologic activity of Th2-cell allograft aug-
mentation, yet also identified the need for further improvements in this
strategy if the goal of improving transplant outcomes might be realized.
In a second protocol (NCT00079625) performed in the setting of allogeneic
HCT for treatment of chemotherapy-refractory metastatic breast cancer,
we evaluated the safety and feasibility of “T-cell exchange” using
ex vivo
manufactured donor CD4
+
Th2 and CD8
+
Tc2 cells
[98]
. Patients received
immune-depleting chemotherapy followed by an HLA-matching sibling
peripheral blood hematopoietic cell allograft that was T-cell-depleted
(final T-cell content: 1 × 10
5
T cells per kilogram of recipient body weight);
at the time of infusion of the T-cell-depleted allograft, patients also
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